CA2632221A1 - Confocal imaging methods and apparatus - Google Patents

Confocal imaging methods and apparatus Download PDF

Info

Publication number
CA2632221A1
CA2632221A1 CA002632221A CA2632221A CA2632221A1 CA 2632221 A1 CA2632221 A1 CA 2632221A1 CA 002632221 A CA002632221 A CA 002632221A CA 2632221 A CA2632221 A CA 2632221A CA 2632221 A1 CA2632221 A1 CA 2632221A1
Authority
CA
Canada
Prior art keywords
line
radiation
sites
rectangular
detector array
Prior art date
Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
Granted
Application number
CA002632221A
Other languages
French (fr)
Other versions
CA2632221C (en
Inventor
Wenyi Feng
Theofilos Kotseroglou
Mark Wang
Alexander Triener
Diping Che
Robert Kain
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Illumina Inc
Original Assignee
Illumina, Inc
Wenyi Feng
Theofilos Kotseroglou
Mark Wang
Alexander Triener
Diping Che
Robert Kain
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Family has litigation
First worldwide family litigation filed litigation Critical https://patents.darts-ip.com/?family=37808340&utm_source=google_patent&utm_medium=platform_link&utm_campaign=public_patent_search&patent=CA2632221(A1) "Global patent litigation dataset” by Darts-ip is licensed under a Creative Commons Attribution 4.0 International License.
Application filed by Illumina, Inc, Wenyi Feng, Theofilos Kotseroglou, Mark Wang, Alexander Triener, Diping Che, Robert Kain filed Critical Illumina, Inc
Publication of CA2632221A1 publication Critical patent/CA2632221A1/en
Application granted granted Critical
Publication of CA2632221C publication Critical patent/CA2632221C/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64Fluorescence; Phosphorescence
    • G01N21/645Specially adapted constructive features of fluorimeters
    • G01N21/6456Spatial resolved fluorescence measurements; Imaging
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64Fluorescence; Phosphorescence
    • G01N21/6428Measuring fluorescence of fluorescent products of reactions or of fluorochrome labelled reactive substances, e.g. measuring quenching effects, using measuring "optrodes"
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64Fluorescence; Phosphorescence
    • G01N21/645Specially adapted constructive features of fluorimeters
    • G01N21/6452Individual samples arranged in a regular 2D-array, e.g. multiwell plates
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64Fluorescence; Phosphorescence
    • G01N21/645Specially adapted constructive features of fluorimeters
    • G01N21/6456Spatial resolved fluorescence measurements; Imaging
    • G01N21/6458Fluorescence microscopy
    • GPHYSICS
    • G02OPTICS
    • G02BOPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00Microscopes
    • G02B21/0004Microscopes specially adapted for specific applications
    • G02B21/002Scanning microscopes
    • G02B21/0024Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders
    • G02B21/0032Optical details of illumination, e.g. light-sources, pinholes, beam splitters, slits, fibers
    • GPHYSICS
    • G02OPTICS
    • G02BOPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B21/00Microscopes
    • G02B21/0004Microscopes specially adapted for specific applications
    • G02B21/002Scanning microscopes
    • G02B21/0024Confocal scanning microscopes (CSOMs) or confocal "macroscopes"; Accessories which are not restricted to use with CSOMs, e.g. sample holders
    • G02B21/0036Scanning details, e.g. scanning stages
    • G02B21/004Scanning details, e.g. scanning stages fixed arrays, e.g. switchable aperture arrays
    • GPHYSICS
    • G01MEASURING; TESTING
    • G01NINVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64Fluorescence; Phosphorescence
    • G01N2021/6417Spectrofluorimetric devices
    • G01N2021/6421Measuring at two or more wavelengths

Abstract

The invention provides imaging apparatus and methods useful for obtaining a high resolution image of a sample at rapid scan rates. A rectangular detector array having a horizontal dimension that is longer than the vertical dimension can be used along with imaging optics positioned to direct a rectangular image of a portion of a sample to the rectangular detector array. A scanning device can be configured to scan the sample in a scan-axis dimension, wherein the vertical dimension for the rectangular detector array and the shorter of the two rectangular dimensions for the image are in the scan-axis dimension, and wherein the vertical dimension for the rectangular detector array is short enough to achieve confocality in a single axis.

Description

CONFOCAL IMAGING METHODS AND APPARATUS
BACKGROUNI) [0001] The present invention relates generally to the field of optical imaging.
Specifically, the present invention relates to imaging systems for use in detecting microarrays.
[0002] Light microscopes provide a powerful tool for investigating samples at submicron resolution. For example, in biology and medicine, appropriate molecular tags, such as fluorescent and immunofluorescent tags, are used to label individual molecules and unique signals from the tags are detected by light microscope to identify their presence. Detection at submicron resolution allows not only determination of the presence of tagged molecules, but also their location in and around cells or tissues.
[0003] Two conflicting goals of light microscopy inspection systems concern providing high speed imaging and high resolution imaging. Typically, 'the resolution of a light microscope is inversely proportional to the imaging speed. Thus, greater resolution is often achieved at the cost of lower inspection rate. One technique to accommodate the aforementioned conflict is to selectively choose the resolution of the system according to specifics of the sample being observed or other conditions of the experiment. Thus, one can use lower resolution to achieve higher speeds while searching for an area of interest in a sample and then once a location of interest is found, imaging can be carried out at higher resolution, albeit at the cost of increasing the time of acquisition for the image.
[0004] Significant advances have been made in the ability of microscopes to investigate samples in three dimensions. The advent of confocal microscopes and improvements gained through related technology, allow a discrete point i,n 3-dimensional space to be detected at high resolution while rejecting unwanted signal from the volume around that point. Scanning confocal microscopy can be carried out to effectively move the point of detection through the sample and collect signal from each point to reconstruct an accurate 3-dimensional image of the sample.
[0005] Technology developed for light microscopy has been applied to other fields of image detection as well. For example, the technology has been used to obtain images of microarrays containing thousands of molecular probes attached to the surface of a substrate. Imaging of the surface of the microarrays after exposure to a biological sample of interest allows thousands of target molecules to be evaluated simultaneously, thereby providing vast amounts of information about the sample.
For example, microarrays can be used to determine the number and types of genes that are expressed under particular conditions, which can in turn provide a holistic view of the biological response to the condition. Furthermore, similarities and differences between the genetic make-up of individuals can be evaluated using microarrays such that the genetic basis for particular traits can be determined.
Information about the gene expression responses and genetic make-up of individuals can be used for diagnostic and prognostic purposes, for example, to determine susceptibility to a certain disease or response to a particular drug.
[0006] Although microarray detection has benefited from advances in light microscopy, there are a number of areas that have not been addressed adequately in regard to microarray imaging. In particular, advances directed to increasing image resolution and collection efficiency in light microscopy have come about by improving 3-dimensional confocal detection and altering magnification levels.
However, typically array detection is carried out in only 2-dimensions and at a fixed magnification level. Furthermore, many of the advances in high resolution light microscopy have favored improvements in resolution over scan speed. These advances are favorable for imaging small samples, on the order of one or a few biological cells; however, the advances have not necessarily benefited high resolution scanning of substantially larger samples such as microarrays.
[0007] Thus, there exists a need for scanning devices and methods that allow imaging of microarrays and other 2-dimensional substrates at high resolution and at high speed. The present invention satisfies this need and provides other advantages as well.

BRIEF DESCRIPTION
[0008] The present invention provides a novel approach to microarray imaging and analysis to respond to such needs. The technique may be used with a wide range of microarray technologies, including arrays made by microbeads, photolithography, printing techniques, electrochemistry, and so forth. The technique relies upon confocal line scanning of the microarray to image individual sites on a substrate.
Scanned lines may comprise more than one wavelength of light, such as a pair of complimentary wavelengths for reading different colors in a retrobeam resulting from excitation by combined wavelengths from lasers, confocally directed towards successive lines of sites on the microarray. The use of confocal line scanning greatly improves the speed of imaging of the inicroarray, while significantly reducing the potential for crosstalk as a result of unwanted excitation of neighboring sites on the array.
[0009] The invention provides an imaging apparatus. The imaging apparatus can include (a) a radiation source positioned to send excitation radiation to at least a portion of a sample region; (b) a rectangular detector array; (c) imaging optics positioned to direct a rectangular image of the portion to the rectangular detector array; and (d) a scanning device configured to scan the sample region in a scan-axis dimension, whereby the portion of the sample region that forms a rectangular image at the rectangular detector array is changed, wherein the shorter of the two rectangular dimensions for the rectangular detector array and the shorter of the two rectangular dimensions for the image are in the scan-axis dimension, and wherein the shorter of the two rectangular dimensions for the rectangular detector array is short enough to achieve confocality in a single axis of the rectangular detector array, wherein the single axis is the shorter of the two rectangular dimensions for the rectangular detector array.
[0010] The invention further provides a method of obtaining an image of a sample.
The method can include the steps of (a) contacting at least a first portion of a sample with excitation radiation under conditions wherein radiation is emanated from the first portion; (b) directing the radiation emanated from the first portion to form a rectangular image of the first portion at a rectangular detector array; and (c) scanning the sample region in a scan-axis dimension, thereby repeating steps (a) and (b) to forrn a rectangular image of a second portion of the sample at the rectangular detector array, wherein the shorter of the two rectangular dimensions for the rectangular detector array and the shorter of the two rectangular dimensions for the images are in the scan-axis dimension, and wherein the shorter of the two rectangular dimensions for the rectangular detector array is short enough to achieve confocality in a single axis of the rectangular detector array, wherein the single axis is the shorter of the two rectangular dimensions for the rectangular detector array.
[0011] The invention also provides a method of configuring a scanner to achieve confocality in a single axis. The method can include the steps of (a) providing an apparatus having (i) a radiation source positioned to send excitation radiation to at least a portion of a sample region; (ii) a rectangular detector array; (iii) imaging optics positioned to direct a rectangular image of the portion to the rectangular detector array; and (iv) a scanning device configured to scan the sample region in a scan-axis dimension, whereby the portion of the sample region that forms a rectangular image at the rectangular detector array is changed, wherein the shorter of the two rectangular dimensions for the rectangular detector array and the shorter of the two rectangular dimensions for the image are in the scan-axis dimension; and (b) positioning the rectangular detector array or the imaging optics to restrict the shorter of the two rectangular dimensions for the rectangular detector array to be short enough to achieve confocality in a single axis of the rectangular detector array, wherein the single axis is the shorter of the two rectangular dimensions for the rectangular detector array.
[0012] The methods can be carried out using the apparatus described in further detail below. However, it will be understood that the method steps exemplified below with regard to particular apparatus can also be carried out using an alternative apparatus.

[00131 In accordance with certain aspects of the invention, a method for imaging a biological microarray includes generating a plurality of radiation beams from respective lasers. The radiation beams are then converted to radiation lines, the lines being greater in width than in height. The radiation lines are then combined into a singular radiation line. A portion of a microarray is then radiated with the single combined radiation line. Radiation from the microarray resulting from irradiation of the portion is confocally returned to a detector, such as a detector array.
I)iscrete sites in the portion of the microarray are then imaged based upon radiation received by the detector. In an alternative embodiment, the two radiation lines can be combined such that two lines are nearly collinear and the portion of the microarray is irradiated with the nearly collinear lines of radiation. The two lines are typically separated by a distance equivalent to the width of each line in order to minimize crosstalk between channels. In particular embodiments, the discrete sites in the portion of the microarray can be imaged based upon radiation received by two detectors, one for each of the two nearly co-linear lines.

[0014] The invention may make use of various optical devices for generating the radiation lines, and for confocally irradiating the microarray, for example, a line generator optic may be used for converting a radiation beam from each laser to a line.
Exemplary line generator optics include, but are not limited to, an aspherical lens, such as a Powell lens, a cylindrical lens or a diffractive element. Optics may then be provided for focusing the line on the portion of the microarray, and for returning radiation caused by fluorescence of sites on the microarray to the detector.

[0015] In an alternative embodiment, radiation lines from individual lasers may be first combined, then the combined beam converted to a single radiation line.
This single 'beam, as before, may then be confocally directed toward a portion of the microarray. As set forth above, the combined beam can be configured to form a single radiation line or it can be configured such that the two lines are nearly collinear and a portion of the microarray is irradiated with the nearly collinear lines of radiation.

[0016] In the various embodiments, the microarray may be advanced slowly in a desired direction to successively irradiate lines of the sites on the microarray for imaging purposes. The lines themselves may be continuous or, in certain embodiments, discontinuous, but simultaneously irradiate multiple sites along the line on the microarray.

DRAWINGS
[0017] These and otlier features, aspects, and advantages of the present invention will become better understood when the following detailed description is read with reference to the accompanying drawings in which like characters represent like parts throughout the drawings, wherein:

[0018] FIG. 1 is a diagrammatical overview of a microarray scanning system for confocal line scanning of a microarray in accordance with aspects of the present technique;

[0019] FIG. 2 is a diagrammatical perspective view of a portion of a microarray illustrating an exemplary"manner in which a radiation line is directed toward regions of the microarray in which sites are located that are to be imaged;

-[0020] FIG. 3 is a more detailed diagrammatical representation of a portion of a microarray that is illuminated by a confocal radiation line to image the sites on the microarray in accordance with the present technique;

[0021] FIG. 4 is a diagrammatical perspective view of a combined radiation line directed toward a surface of a microarray to confocally irradiate sites on the array, and to confocally return radiation to a detector in accordance with aspects of the present technique;

[0022] FIG. 5 is a similar diagrammatical perspective view illustrating a series of confocally directed beams of radiation along a line for similarly irradiating sites of a microarray in accordance with the present technique;

[0023] FIG. 6 is a diagrammatical side view of a technique for converting output of a laser to a radiation line for confocal line scanning of a microarray;

[0024] FIG. 7 is a similar, top view of the conversion of the output of a laser to a radiation line for use in the present confocal line scanning technique;

[0025] FIG. 8 is a graphical representation of an intensity profile for a radiation line produced by the arrangements of FIGS. 6 aid 7;

[0026] FIG. 9 is a diagrammatical representation of a first exemplary configuration for a modular arrangement used in converting output of a laser to a radiation line for confocal line scanning in accordance with the invention;

[0027] FIG. 10 is an alternative arrangement, for conversion of a laser output to a radiation line in accordance with the present invention;

[0028] FIG. 11 is a further alternative arrangement for converting laser output to a radiation line in accordance with the invention;

[0029] FIG. 12 is yet another alternative configuration for converting laser output to align a radiation;

[0030] FIG. 13 is a sectional view of an exemplary line generator module suitable for use in accordance with the invention;

[0031] FIG. 14 is a diagrammatical overview for a scanning system that includes two laser beams, the output of which is combined for confocal line scanning of a microarray;

[0032] FIG. 15 is a diagrammatical overview of an alternative arrangement for multi-wavelength confocal line scanning of a microarray;

[0033] FIG. 16 is an opto-mechanical diagrammatical representation of a presently contemplated implementation for multi-wavelength confocal line scanning of a microarray;

[0034] FIG. 17 is a diagrammatical view of a series of individual sites on a biological microarray, illustrating how the confocal line scanning of the present invention improves accuracy by reducing the potential crosstalk, particularly in certain types of layout of the sites on the micorarray with respect to radiation lines used in imaging;

[0035] FIGS. 18 through 21 are diagrammatical views of exemplary radiation line generators that may be suitable for use in the invention;

[0036] FIGS. 22 and 23 are diagrammatical views of line generators in a fluorescence imaging system, suitable for use in the invention;

[0037] FIGS. 24(a)-(c) are diagrains showing the projection of a laser spot on a line scan camera and binning and TDI implementations in accordance with certain aspects of the invention;

[0038] FIG. 25 is a diagrammatical view of an image scanning system that is configured to conduct multi-spectral fluorescence imaging in accordance with aspects of the invention;

[0039] FIG. 26 is a block diagram of an exemplary line-scan imaging sensor for use with the system shown in FIG. 25;

[0040] FIG. 27 is a diagrammatical view of a further image scanning system that is configured to conduct multi-spectral fluorescence imaging;

[0041] FIG. 28 is a block diagram of an exemplary line-scan imaging sensor for use witll the system shown in FIG. 27;

[0042] FIG. 29 is a block diagram of an exemplary line-scan imaging detector for use with the invention; and [0043] FIGS. 30(a)-(b) are block diagrams of other exemplary line-scan imaging detectors for use with the invention.

DETAILED DESCRIPTION

[0044] The present invention provides an image scanning system and architecture having rapid scan times while maintaining high resolution and image quality.
These and other advantages result from configuring a detector array to achieve confocality in the scanning axis by restricting the scan-axis dimension of the detector array. As set forth in further detail below, an apparatus of the invention can be configured to achieve confocality in a single axis of a detector array such that confocality only occurs in that dimension.

[0045] The detector array can have rectangular dimensions such that the shorter dimension of the detector is in the scan-axis dimension. Imaging optics can be placed to direct a rectangular image of a sample region to the detector array such that the shorter dimension of the image is also in the scan-axis dimension. In this way, the detector array forms a virtual slit. A virtual slit configuration provides several advantages over the use of a typical slit placed in front of a detector. For example, configuring a detector array as a virtual slit reduces the number of unused array elements compared to a configuration in which a detector array, having standard dimensions, is used with a slit. Reducing the number of unused elements increases efficiency of data acquisition and reduces image processing time. Furthermore, using a virtual slit allows both the detector and slit to be at the focal plane of the projection lens eliminating any focus compromise of either position or the requirement for a relay lens between the slit and detector.

[0046] A detector array configured to have a virtual slit is particularly useful when employed in an imaging apparatus that is further configured to direct a radiation line to a sample. The radiation line can have rectangular dimensions in which the shorter dimension is short enough to achieve confocality in a single axis corresponding to the shorter dimension of the detector array. Thus, confocality can be achieved for excitation, detection or both. An instrument can be configured to limit excitation error in the confocal axis such that predominantly all of the excitation radiation is contained within a spot comparable with the resolution of the instrument.

[0047] An apparatus that includes a detector array forming a virtual slit can be configured to obtain an image of the sample at high resolution, for example, in the low micron to submicron range. In particular embodiments, an image can be obtained at a Rayleigh resolution between 0.2 and 10 micrometers. Furthermore, the ratio of the shorter of the two rectangular dimensions for the rectangular detector array and the product of the Rayleigh resolution of the imaging optics multiplied by the magnification of the imaging optics can be used to determine the size and dimensions of the virtual slit for achieving confocality in a single axis. If desired, the ratio of the shorter of two rectangular dimensions for a radiation line to the Rayleigh resolution of the imaging optics can be selected to achieve confocality in a single axis.

[0048] Accordingly, an imaging apparatus of the invention can be configured to have resolution along the length of the line perpendicular to the scan axis that is matched to the system resolution. For example in a CCD device, 4000 CCD
elements can be used along the length of a 2mm radiation line (the horizontal axis) resulting in a 0.5 m pixel resolution at a sample. The number of CCD elements "n" in the direction perpendicular to the radiation line (the vertical axis) can be chosen to collect substantially all of the emitted radiation while reducing the amount of unwanted background radiation collected.

[0049] . An imaging apparatus of the invention can be further configured such that all pixel elements in the vertical axis are collected in a common "bin" and read out as a single value. Advantages of the binning approach compared to a typical Time Delay Integration (TDI) design are that the readout rate can be reduced by a factor of "n", the system has confocality in one axis, and the tolerance of the synchronization timing of the readout with the y-stage movement can be reduced. It will be understood that a TDI design can be configured to have a virtual slit by limiting the number of vertical pixels. An additional advantage over system designs where n=1 are that the collection efficiency of the system can be increased and the sensitivity to small optical alignment drifts can be decreased.

[0050] Turning now to the drawings, and referring first to FIG. 1, an imaging system 10 is illustrated diagrammatically as including a scanner 12 in which a sample or microarray 14 may be inserted for imaging purposes. As described more fully below, the microarray 14 includes a substrate or support on which an array of sites is formed. Each site including an attached molecular fragment, such as a gene or gene fragment, which may have attached thereto a molecule, which may be a complementary molecule in the case of DNA or RNA probes, from a specific sample.
In present embodiments, many thousands of such sites may be provided in rows or a grid pattern in portions or segments on the microarray. The microarray itself may be formed by various technologies, including, as in a present embodiment, microbeads.
Other microarrays which may be imaged in accordance with the present techniques may include inicroarrays formed by photolithography, and other processes known or developed in the art.

[0051] The scanner 12 will include optics described in greater detail below for confocal line scanning of the sites on microarray 14. In the illustrated embodiment, the scanner is a table-top device having a sample tray 16 in which the microarray, or a plurality of microarrays may be positioned. The tray may be configured to advance the microarray 14 into a scanning position, and subsequently slowly move the microarray, as described below, to allow successive lines on the microarray to be irradiated, and return radiation or retrobeams caused by fluorescence of individual sites. The retrobeams are focused on a detector for imaging and analyzing the sites, also described below. In particular embodiments, multiple retrobeams can be focused to multiple different detectors. For example, a retrobeam of a first wavelength can be focused to a first detector and a retrobeam of a second retrobeam can be focused to a second detector, as set forth in further detail below.

[00521 Control signals for operation of the scanner 12 originate from a controller or workstation 18. The workstation 18 also includes software for receiving the imaging signals from the scanner 12. The imaging software of worlcstation 18 will typically be embodied in a general purpose or application-specific computer 20 which also controls and receives signals from interface components 22, which will typically include a monitor 24 and input devices 26. The imaging software operable in workstation 18 will preferably provide an intuitive interface for loading and initializing the scanner, for performing imaging scans on microarrays, and for saving the data. During the scanning process, the system 10 creates individual files for different wavelengths of radiation used to image the microarray, which may be referred to herein as red and green channels. These may be provided in a consolidated file. Data and associated images may then be saved in a convenient format, such as a conventional TIFF format, or any other suitable image data format or protocol.
The workstation 18 may be coupled to other network components, including down-stream processing and application-specific software for higher-level and data analysis, such as via a networlc indicated generally by reference numeral 28 in FIG. 1.

10053] As noted above, the microarray 14 will include a plurality of sites arranged in portions or regions of a substrate, for example, as indicated generally in FIG. 2. As shown in FIG. 2, the microarray 14 may include a support or substrate 30, which may be a glass, a plastic, a semiconductor, or any other convenient support such as those described elsewhere herein. On this support 30, one or more sample areas 32 are provided in which individual sites will be formed, each typically provided with a respective probe molecule used to test a sample. In a present invention, the sample area 32 is scanned for imaging purposes by a radiation line, indicated generally by reference numeral 34 in FIG. 2. The radiation line is formed by excitation radiation which is confocally directed along the line 34 to irradiate a plurality of sites simultaneously, as indicated generally by arrows 36 in FIG. 2. The individual sites at which target molecules (e.g., genetic fragments) will have bound are thereby caused to fluoresce due to the presence of dyes indicative of an interaction of a target with the site, returning radiation as indicated by lines 38 in FIG. 2. As described below, this returned radiation, or retrobeam, will be confocally directed toward an imaging detector where an image will be made of the line for further processing and analysis.
To permit the sites to be successively imaged, then, the entire microarray may be displaced slowly as indicated generally by reference numeral 40. The line 34 along which the sites are irradiated will thereby generally progress along successive parallel locations on the microarray as the microarray is displaced.

[0054] An exemplary portion of a microarray imaged in accordance with such confocal line scanning is illustrated in FIG. 3. Again, reference numeral 14 refers to the microarray, while reference numeral 32 refers to one of the sample areas in which individual sites 42 are disposed. In the illustrated embodiment, the sites are provided in a generally hexagonal pattern. Scanning by line 34 progresses through successive lines 44 of sites 42. As described in greater detail below, while the present confocal line scanning approach may be used witli different layouts or grid patterns of sites on the micorarray, a hexagonal pattern is particularly useful with confocal line scanning insomuch as it provides for a reduced probability of crosstalk due to the placement and spacing between the sites or site edges. The hexagonal packing, designated generally by reference numeral 46 in FIG. 3, is believed to provide an optimal degree of accuracy due to such crosstalk reduction, balanced with a superior packing density of the sites.

[0055] As described below, and as also illustrated in FIG. 3, as the microarray 14 is advanced as indicated by reference numeral 40, the confocal radiation line irradiates a phirality of sites located along the line. The line is wider, in a horizontal direction shown in FIG. 3 than it is high. Thus, the line may irradiate adjacent sites in a line or row of sites without irradiating sites in adjacent lines. In a present embodiment, however, the radiation line 34 is sufficiently thin, at the level of the sites, or of a sufficient vertical height in the arrangement illustrated in FIG. 3 to permit it to illuminate less than the entire area occupied by the sites. In a presently contemplated embodiment, the radiation line 34 is, for example, 2 mm in length (horizontal dimension) and less than 3 mm in height (vertical dimension).
Thus, the software provided for imaging; mentioned above, may employ techniques such as time delay imaging, in which the readout from the detector described below is shifted with movement of the sample to provide more accurate representations of the individual sites in each row or line.

[0056] For purposes of explanation, several aspects of the invention have been exemplified with regard to moving a microarray past a radiation line. It will be understood that embodiments in which the radiation line is moved in addition to or alternatively to moving the microarray can also be used. Thus, line-scanning can be
13 carried out by relative displacement of a radiation line and/or microarray relative to each other. A portion of the sample excited by the radiation line can form a rectangular image on the detector array (described below).

[0057] FIG. 4 is a further diagraminatical representation of the present confocal line scanning approach to imaging the microarray 14. As indicated above, the microarray is radiated along a line 34 as the support 30 is slowly moved as indicated by reference numeral 40. As illustrated in FIG. 4, the line 34 is formed by radiation from a source 48 which is directed towards directing optics 50 and therefrom to focusing optics 52. As described more fully below, the radiation source 48 will be a beam with a linear cross section or a radiation line including a plurality of wavelengths of light used to cause fluorescence at correspondingly different wavelengths from the sample, depending upon the particular dyes used. The focusing optics 52 will then confocally direct the radiation line toward the substrate 30 to irradiate the sites as described above along line 34. It should be noted that the sites may be provided at the surface of the substrate 30 or slightly below the surface (e.g., below a protective film or layer). The confocal irradiation along line 34 will essentially focus the radiation toward the sites themselves at whatever level they are found in the microarray.

[0058] The excitation path 54, in the present embodiment, is coplanar with a retrobeam path 56 for radiation returned from the sample by fluorescence of dyes associated with molecules attached to probes at the individual microarray sites. The returned radiation is again focused by focusing optics 58 such that it impacts a detector 60 to create imaging signals used to reconstruct an image of the microarray, and of individual sites on the microarray. Specific embodiments for creating the radiation beam, directing the beam to the microarray, and for detecting returned radiation are described in greater detail below.

[0059] It should be noted that, as illustrated generally in FIG. 5, the radiation line used to image the sites simultaneously, in accordance with the present invention, may be a continuous or discontinuous line. FIG. 5 represents, diagrammatically, a discontinuous line made up of a plurality of confocally directed beams of light which
14 nevertheless irradiate a plurality of points along a line 34. In the embodiment illustrated in FIG. 5, discontinuous beams 62 are created from separate but adjacent radiation sources 48. These beams, as before, are confocally directed toward the microarray and irradiate adjacent spots 64 along the microarray in a line 34.
As with the continuous confocal line scanning described above, the microarray will typically be advanced slowly as indicated by arrow 40 to irradiate successive lines along the microarray, and thereby successive rows or lines of sites.

[0060] Typically, the invention is used to excite and detect a line simultaneously.
In some embodiinents, line confocal point scanning can be used such that the optical system directs an excitation point or spot across a sample by scanning the excitation beam through an objective lens. The detection system images the emission from the excited point on the detector without "descanning" the retrobeam. This occurs since the retrobeam is collected by the objective lens and is split off the excitation beam optical path before returning back through the scan means. Therefore the retrobeam will appear on the detector at different points depending on the field angle of the original excitation spot in the objective lens. The image of the excitation point, at the detector, will appear in the shape of a line as the excitation point is scanned across the sample. This architecture is useful, for example, if the scan means cannot for some reason accept the retrobeam from the sample. Examples are holographic and acoustic optic scan means that are able to scan a beam at very high speeds but utilize diffraction to create the scan. Therefore the scan properties are a function of wavelength. The retrobeam in fluorescence is at a different wavelength from the excitation beanl.

[0061] FIGS. 6 and 7 illustrate an exemplary linearization of an input laser beam for confocal line scanning of a microarray in accordance with a presently contemplated embodiment. FIG. 6 represents what may be considered an elevational view of the conversion or linearization of the input beam, while FIG. 7 may be considered to illustrate a top plan view, although these orientations are understandably interchangeable, depending upon the orientation of the line and microarray to be scanned, as described below. As shown in FIG. 6, an input beam 66 from a laser (not shown) will typically take the form of a circular Gaussian beam 66. An aspherical lens 68, such as a Powell lens converts the input beam to a line 70 of radiation which is directed toward an objective lens 72. As illustrated in the top view of FIG. 7, the aspherical lens 68 effectively produces a generally flat radiation line which is further converted to a confocally concentrated beain 74 by the objective lens 72.

[0062] As illustrated in FIG. 8, the arrangement shown in FIGS. 6 and 7 produces a linear region of radiation which can be used to simultaneously irradiate a number of sites on the microarray. FIG. 8 is a graphical representation of the siinulated illumination along a radiation line produced by an aspherical lens as described with reference to FIGS. 6 and 7. The relative illumination of the beam is indicated by vertical axis 76, while the image coordinate in millimeters is represented by the horizontal axis 78. In the illustrated embodiment, the illumination intensity rises rapidly near an edge of the aspherical lens, as indicated by reference numeral 80 and drops rapidly near an opposite edge, as indicated by reference numeral 82.
Between the edges a useful segment of radiation 84 has a substantially constant relative illumination level. In a present embodiment, the useful width 86 of the radiation line is used to irradiate lines or rows of sites on the microarray simultaneously.
The simulation illustrated in FIG. 8, for example, provided a useful scanning length 86 of approximately 1.024 millimeters, although a number of factors, including the optics involved may provide for other useful radiation line lengths.

[0063] As will be appreciated by those skilled in the art, for imaging at a plurality of wavelengths, a confocal line scanning fluorescence imaging system in accordance with the present technique will provide for lines of multiple wavelengths with the diffraction-limited width and uniform distribution along a length to irradiate sample sites and thereby to excite multiple fluorescent dyes. The line generator approach illustrated in FIGS. 6, 7 and 8 provide an exeinplary mechanism for such linearization of irradiating, multiple wavelength light. The provision of multiple wavelengths in the radiation line will be described in greater detail below. Effectively, the arrangement illustrated in FIG. 6, 7 and 8 fan a collimated input beain in one dimension and maintairi the beam collimated in a perpendicular dimension. The beam is then focused by,the objective lens 72 to a diffraction-limited line on a focal plane of the lens.

[0064] Based upon the sag of the aspherical lens, a collimated pure Gaussian input beam with adefined beam diameter is preferred to generate a line of uniform distribution. A presently contemplated technique for obtaining a beam with an almost pure Gaussian distribution is the use of a single mode fiber or fiber cable to provide input to the aspherical lens.

[0065] Several arrangements may be foreseen for use of such a single mode fiber or fiber cable. FIG. 9 illustrates a first exemplary embodiment in which a linear radiation source 88 includes a laser 90 coupled to a single mode fiber pigtail 92 and therethrough to a line generator module 94. The objective lens downstream of the' aspherical lens is omitted from the illustration in FIG. 9. The generated line profile is not only sensitive to the input beam profile but also sensitive to input beam diameter, collimation characteristics and centering Qf the beam to the aspherical lens.
That is, the aspherical lens may be designed for a defined input beam diameter, and the assembly, particularly the components of the line generator module 94, is aligned to achieve the design performance.

[0066] In the illustrated einbodiment, the line generator 94 includes several optical components which are pre-aligned in a modularized assembly to facilitate both their quality control and packaging in the scanner. In particular, line generator modular 94 may include a collimator 96 that collimates the input beam from the single mode fiber 92 and directs the collimated beam to an aspherical lens 100. A laser line filter 98 may also be einployed, particularly for applications of fluorescence imaging, to reduce background noise. The illustration of FIG. 9 may provide for pre-assembling or terminating the single mode fiber 92 on both ends, that is, at the laser 90 and at the line generator module 94.

[0067] Alternatively, the linear radiation source 88 may provide for splicing a pair of fiber pigtails as illustrated generally in FIG. 10. In the embodiment of FIG. 10, the fiber pigtail 92 is pre-coupled to the laser 90, while a second fiber pigtail 102 is pre-coupled to the line generator module 94. The two fibers may then be connected or spliced at an intermediate point as indicated generally by reference numeral 104.

[0068] In a further alternative configuration, illustrated in FIG. 11, a single fiber pigtail 102 may again be used, which may be pre-assembled with the line generator module 94. In this embodiment, however, the laser 90 provides input to the fiber pigtail 92 by active coupling, as indicated by reference numeral 106.

[0069] In a further alternative configuration, illustrated generally in FIG.
12, a fiber pigtail 102 may be pre-assembled with laser 90. Rather than providing a collimator in the line generator module 94 as described above, a variable beam expander 108 may be employed for providing input to a modified module 110 which includes an aspherical lens, as before. The embodiment of FIG. 12 may require that the input beam diameter match the desired diameter by virtue of the variable beam expander 108.

[0070] An exemplary line generator module 94 is illustrated generally in FIG.
13.
As indicated above, and as shown in the physical implementation of FIG. 13, the module 94 may receive an input beam, designated generally by reference numeral 112, via a single mode fiber 92. An output radiation line 114 is emitted by the module. In the illustrated embodiment, a fiber optic connector 116 serves to join the single mode fiber 92 to the input side of the module 94. Therefrom, the beam propagates through collimator 96, laser line filter 98 (where provided), and aspherical lens 100. Again, the modularization of the optical components used to convert the output of the laser to a radiation line is favored insomuch as it facilitates assembly of the overall system, alignment of the optics, and later servicing and replacement of the optical components, if needed.

[0071] As indicated above, in certain contemplated embodiments, the radiation source is a laser. Other useful radiation sources might include, for example, a lamp such as an are lamp, quartz halogen lamp and light emitting diodes. Any of a variety of other radiation sources can be used as desired for exciting a sample at a particular wavelength. As desired for a particular application, the radiation source can generate radiation at various wavelengtlis including, for example, a wavelength in the UV, VIS
or IR range. For example, an apparatus of the invention can include a laser that generates light at 405 nm, 488 nm, 532 nm or 633 nm.

[0072] Moreover as noted below, the systein can include more than one radiation source. The multiple radiation sources can be lasers each capable of generating radiation at different wavelengths. The use of multiple radiation sources that generate radiation at different wavelengths can be useful, for example, in applications wherein a sample includes one or more fluorophores that produce different emission signals when excited at different wavelengths. Different emission signals can be collected simultaneously, for example, using multiple detection arms as set forth below in further detail. Alternatively or additionally, different emission signals can be collected sequentially following sequential excitation at different wavelengths.

[0073] As noted above, certain embodiments of the invention may further include an expander positioned to receive excitation radiation from a radiation source and to send an expanded beam of the radiation to a line generator. In particular embodiments, the diameter of the excitation beam generated by the radiation source is approximately 1 mm in diameter. A first expander is capable of expanding the diameter of the beam. For example, according to one embodiment, the expander expands the excitation beam to a diameter of 4 nnn. Other useful beam expanders can bring the diameter of a radiation beam to at least about 0.5 mm, 1 mm, 2 mm, 5 mm, mm, 15 mm, 20 mm or more.

[0074] As also discussed above a line generator useful in the invention can include a diffractive element configured to generate a diffraction-limited line with uniform intensity distribution. For example a cylindrical micro-lens array and a condenser can be used. The cylindrical micro-lens array can be configured to focus excitation radiation onto the front focal plane of the condenser to generate a diffraction-limited line with uniform intensity distribution. A further example of a line generator is a one-dimensional diffuser having an angular uniformity and a condenser, wherein the one-dimensional diffuser is placed at the front focal plane of the condenser to generate a diffraction-limited line with uniform intensity distribution. If desired, the line generator can further include an aspheric refractive lens to generate a diffraction-limited line with uniform intensity distribution. An exemplary aspheric refractive lens is a Powell lens.

[0075] In a particular embodiment, the line generator can be configured to receive an input excitation beam having a diameter of 4 mm to obtain a fan angle of 6 degrees. Other useful configurations include, but are not limited to, those that receive an input excitation beam having a diameter of at most about 0.1 to 50 mm. A
line generator can obtain a fan angle of at least about 0.1 to at most about 80 , full width.
The beam diameter and fan angle can be selected to achieve a desired shape for a radiation line. Generally, the width of the radiation line depends upon beam diameter such that a larger beam diameter provides a wider radiation line in the vertical dimension and the length of the radiation line depends on the fan angle such that a larger fan angle provides a longer radiation line in the horizontal dimension.
Typically, the line should appear to originate at the pupil of the objective, however this is not a requirement.

[0076] As set forth above, any of a variety of optical elements capable of generating a line can be placed in the optical path between a radiation source and a sample region to be irradiated. For example, an arc lamp focused on a slit and then collimated can be used to generate a line. A further example is an edge emitting diode laser having an anamorphic beam which generates a line when focused. It will be understood that a radiation source used to irradiate a sample region can itself be capable of generating a line. Thus, a radiation source useful in the invention can include a line generator.

[0077] Any of a variety of methods and apparatus including, but not limited to those exemplified above, can be used to direct a radiation line to a sample region.
The dimensions of the radiation line can be selected to achieve confocality in a single axis of a rectangular detector array. More specifically, the vertical dimension of the radiation line can be short enough to achieve confocality in the vertical dimension of the rectangular detector array.

[0078] A line generator of the invention is typically configured to produce a radiation line having a shape at a sample region that is rectangular,.or oblong.
Exemplary shapes include, but are not limited to, a rectangular, elliptical, or oval shape. A line generator can be configured to produce a radiation line having one or more of the properties set forth below.

[0079] A radiation line that contacts a sample region can have a ratio of the 1/e2 width of the vertical dimension for the radiation line to the quotient of the vertical dimension for the rectangular detector array divided by the magnification of the imaging optics that results in confocality. in one dimension. For example, the ratio can be at least about 0.5, 1, 1.5, 2, 3 or higher. An apparatus of the invention can be configured to have an upper end for the ratio that is at most about 2, 1.5, 1, 0.5 or lower. The ratio can be outside or inside the above ranges as desired including, for example, being in the range of 0.5 to 3.

[0080] A radiation line that contacts a sample region can have a ratio of the vertical dimension for the radiation line to the quotient of the vertical dimension for the rectangular detector array divided by the magnification of the imaging optics that results in confocality in one dimension. For example, the ratio can be at least about 0.1, 0.5, 1, 5, 10 or higher. The upper end of the ratio can be at most about 10, 5, 1, 0.5, 0.1 or lower. The ratio can be outside or inside the above ranges as desired including, for example, being in the range of 0.1 to 10.

[0081] Furthermore, the ratio of the vertical dimension for the radiation line to the Rayleigh resolution of the imaging optics can be at least about 0.1, 0.5 1, 5, 10 or higher. The upper end of the ratio can be at most about 10, 5, 1, 0.5, 0.1 or lower.
The ratio can be outside or inside the above ranges as desired including, for example, being in the range of 0.1 to 10.

[00821 Although the invention is exemplified herein with regard to embodiments in which a sample region is contacted with a radiation line, it will be understood that the radiation that contacts a sample region can have other shapes including, for example, a square or circle.

[0083] As described below, an apparatus of the invention can include an objective positioned to receive radiation therethrough to illuminate a sample region.
The objective can be further positioned to collect radiation emanating from a sample region and direct it to a detector array. Optionally, the apparatus can include a second expander positioned to receive the excitation radiation from the line generator and send an expanded beam of the radiation to the objective. The second expander can be further configured to decrease the field angle of the radiation line. For example, after the excitation beam passes through the line generator and/or a second expander, it may be directed to an objective by a beam splitter. In particular embodiments, the objective has an external pupil positioned to receive the radiation line therethrough to illuminate the sample region. Preferably, the beanl splitter may be located near the entrance pupil of the objective lens. The beam splitter can be placed at an axial or lateral position relative to the objective. If desired, an objective can have a property of color correction, high numerical aperture, telecentricity, afocality at the backplane or a combination of such properties.

[0084] The beam splitter directs the radiation line to an objective. The objective can be a microscope objective. The objective may have a focal length of 20 mm.
Accordingly, the objective may possess a numerical aperture of 0.366. Further, the objective may have a field angle of +/- 3 degrees and an entrance pupil having a 16 mm dianieter. Preferably, the objective is telecentric. Exemplary telecentric objective lenses useful in the invention include those that are described in U.S.
5,847,400, which is incorporated herein by reference.

[0085] FIG. 14 illustrates an overall optical layout fQr the various components described above in a multiple wavelength scanner 118. The scanner 118 may include a plurality of laser light sources, with two such sources being illustrated in the embodiment of FIG. 14. These include a first laser 120 and a second laser 122.
The first laser 120, in presently contemplated embodiments may be a 658 nm laser, a 750 nm laser, or a 635 nm laser, depending upon the desired application. The second laser 122 may be, for example, a 488 nm laser, a 594 nm laser, or a 532 nm laser.
Other wavelength lasers may, of course, be used. In the present embodiment, the first laser 120 is a 635 nm laser when the second laser 122 is a 488 nm laser, or the first laser 120 is a 750 nm laser when the second laser 122 is a 594 nm laser, or the first laser 120 is a 658 nm laser when the second laser 122 is a 532 nm laser. The selection of the wavelength for each laser will depend, of course, upon the fluorescence properties of the dyes used in the microarray, although the wavelengths of the lasers used in unison for any particular imaging sequence will be distinct from one another to permit differentiation of the dyes at the various sites of the microarray.

[0086] Each laser 120 and 122 is coupled to a single mode fiber 124 and 126, respectively, as described above. Moreover, each fiber 124 and 126 feeds a line generator module 94 of the type described above. Downstream of each module 94, a filter wheel 128 and 130 may be provided. The filter wheels serve to block, pass or attenuate the light depending upon the desired function.

[0087] Output from each of the lasers 120 and 122 will be converted to a near pure Gaussian distribution by the respective single mode fibers 124 and 126, and the resulting beams will be converted to beams with linear cross-sections, also referred to as radiation lines, by the line generator modules 94. Downstream of the filter wheels 128 and 130, the two radiation lines will be combined by a beanz combiner 132.
The coinbined radiation line 134 will, then, comprise liglit at two different wavelengths for irradiating the microarray. The combined radiation line 134 is then directed to a dichroic beam splitter 136 which, directs the beam toward focusing optics 138.
The focusing optics 138 constitute a microscope objective that confocally directs and concentrates the radiation line along the line to the microarray 14 as described above.
Although the invention is exemplified herein with regard to a combined radiation line that forms a single radiation line it will be understood that the two radiation lines can be combined such that two lines are nearly collinear. Thus, a portion of the microarray that is irradiated with the combined radiation line will be irradiated with the nearly collinear lines of radiation. The two lines are typically separated by a distance equivalent to the width of each line in order to minimize crosstalk between channels.

[0088] As illustrated . diagrainmatically in FIG. 14, the microarray 14 may be supported on a stage that allows for proper focusing and movement of the microarray before and during iinaging. The stage can be configured to move the sample, thereby changing the relative positions of the rectangular image and the rectangular detector array in the scan-axis (vertical) dimension. Movement of the translation stage can be in one or more dimensions including, for example, one or both of the dimensions that are orthogonal to the direction of propagation for the radiation line and typically denoted as the x and y dimensions. In particular embodiments, the translation stage can be configured to move in the direction perpendicular to the scan axis for a detector array.. A stage useful in the invention can be fiutlier configured for movement in the dimension along which the radiation line propagates, typically denoted as the Z dimension. Movement in the Z dimension can be useful for focusing the apparatus. In the configuration of FIG. 14, the stage component include tilt actuatqrs 140, typically used for focusing the radiation line, Y-direction actuators and eject components 142 for placing the microarray in a position for scanning, and for gross movements of the microarray between scans, and an X-direction actuators for fine movements of the microarray during scanning.

(0089] Sites on the microarray 14 may fluoresce at wavelengths corresponding to those of the excitation beam and return radiation for imaging. As will be appreciated by those skilled in the art, the wavelength at which the dyes of the sample are excited and the wavelength at which they fluoresce will depend upon the absorption and emission spectra of the specific dyes. Such returned radiation will propagate through beam splitter 136 as indicated generally by retrobeam 146 in FIG. 14. This retrobeam will generally be directed toward one or more detectors for imaging purposes.
In the illustrated embodiment, for example, the beam is directed toward a mirror 148 and therefrom to a second dichroic beam splitter 150. A portion of the beam, as indicated by reference numeral 154, is then directed by mirrors 152 to a bandpass filter wheel 158 that filters the beam to obtain the desired output wavelength corresponding to one of the fluorescent dyes of the sites in the microarray. In particular embodiments, the portions of the beam that are directed to different mirrors can be the respective lines of a combined beam that forms two nearly co-linear lines. A projection lens 160 then directs the filtered beam to a charge coupled device (CCD) sensor 164 which produces output signals corresponding to locations of the radiation in the received beam. Similarly, a second portion 156 of the beam from beam splitter 150 is directed.
to another mirror through a different bandpass filter wheel 158 and projection lens 160. The second beam 156 may also be directed through an optional chromatic aberration coinpensation device 162, which may be motorized. The chromatic aberration compensation device 162 serves to bring both wavelength channels into co-focus. Finally; beam 156, filtered and focused by filter wheel 158 and lens 160 is directed to a second CCD sensor 166. The receipt and processing of signals from the sensors 154 and 166 may be managed by a control board 168.

[0090] A rectangular detector array of the invention can be configured to form a virtual slit as set forth previously herein. In particular embodiments, the size and dimensions of the virtual slit can be determined from the ratio of the vertical dimension for the rectangular detector array and the product of the Rayleigh resolution of the imaging optics multiplied by the magnification of the imaging optics.
For example, the ratio of the vertical dimension for the rectangular detector array and the product of the Rayleigh resolution of the imaging optics multiplied by the magnification of the imaging optics can be in the range of 0.1 to 10 or in the range of 0.5 to 3. An apparatus of the invention can be configured to obtain an image of a sample at a desired pr optimal Rayleigh resolution including, for example, a Rayleigh resqlution between 0.2 and 10 micrometers.

[0091] In particular embodiments, the aspect ratio of the number of detection elements in a first dimension to the number of detection elements in the scan-axis dimension for a rectangular detector array can be greater than 2, 10, 20, 50, 100, 1000 or higher. For example, a line scan CCD camera can be configured to capture, four thousand (4,000) pixels in the first dimension and n pixels in the scan-axis (vertical) dimension. The CCD line scan camera can be designed such that the resolution along the length of the line is matched to the system resolution. In this case, the horizontal axis includes approximately 4,00Q CCD elements along the length of a 2 mm radiation line, resulting in a 0.51Am pixel resolution at the object. The number of CCD
elements "n" in the direction perpendicular to the horizontal axis, also referred to as the vertical axis, can be chosen to collect substantially all of the emitted radiation while reducing the amount of background radiation collected. According to one embodiment of the invention, the CCD has 4096 pixels, each 12 m in size. To image a 2 mm line to this size CCD requires a magnification of 25X. Accordingly, n can be in the range of six to eight pixels. The design architecture limits the excitation error in the confocal axis such that predominantly 100% of the excitation radiation is contained within a spot comparable with the resolution of the system. In this case, the spot size would be roughly 1.0 m.

[0092] Although the apparatus has been exemplified above with regard to a CCD
line scan camera, it will be understood that any of a variety of other detectors can be used including, but not limited to a detector array configured for TDI
operation, a CMOS detector, APD detector, Geiger-mode photon counter or other detector set forth elsewhere herein.

[0093] In general, the operation of the various components illustrated in FIG.

may be coordinated by system controller M. In a practical application, the system controller will include hardware, firmware and software designed to control operation of the lasers, movement and focusing of the objective 138 and microarray support, and the acquisition and processing of signals from the sensors 164 and 166.
The system controller may thus store processed data, and further process the data for generating a reconstructed image of the irradiated sites that fluoresce on the microarray.

[0094] FIG. 15 illustrates an alternative arrangement for the multiple wavelength scanner, designated generally by reference numeral 172. In this alternative arrangement, beams from separate lasers are combined and the cross section of the combined beam then converted to a linear shape by an aspherical lens. Thus, as in the previous embodiment summarized with reference to FIG. 14, input lasers 120 and provide wavelengths of light corresponding to dyes used at various sites on a microarray 14. In the embodiment 172, however, a first laser 120 outputs its beam to a single mode fiber 124, followed by a collimator 174 that collimates this output. The collimated output,may then be directed to a filter wheel 130, and the resulting beam 176 is directed, by mirrors 152 to a variable beam expander 180 of the type described above with reference to FIG. 12.

[0095] Similarly, output from the second laser 122 is directed through a second filter wheel 130 and the resulting beam 178 is directed, such as via mirrors 152 to a second variable beam expander 182. Output from the variable beam expanders, then, is joined by a beam combiner 132. The combined beam 182, which will include light at the desired wavelengths for radiation of the microarray is converted to a line by an aspherical lens 100. As before, then, a combined radiation line 134 including light at the desired wavelengths will be produced and directed to the microarray 14 by a beam splitter 136. The remaining components of the system may be essentially identical to those described above with respect to FIG. 14.

[0096] FIG. 16 provides a somewhat more detailed opto-mechanical diagrammatical representation of a multiple wave-length scanner in accordance with aspects of a presently contemplated embodiment. The scanner 184 may include a first laser assembly 186 which, itself, includes multiple lasers. In the illustrated embodiment, for example, laser assembly 186 includes a first laser 188 which may be a 488 nm laser, and a-second laser 190 which may be a 658 nm laser. The system may further include a second laser assembly 192, which may include, for example, a 594 nm laser 194 and a 750 nm laser 196. As will be appreciated by those skilled in the art, the inclusion of multiple laser assemblies 190 and 192 may allow for different types of scanning operations to be performed with a single scanner, such as decoding functions, analytical functions, and so forth. For example, lasers 188 and 190 may be used in conjunction with one another for certain types of decoding operations, while lasers 194 and 196 may be used in conjunction with one another for other types of decoding. The assemblies may include other lasers which may alternatively be used, or other assemblies may be provided, such as an assembly employing a 635 nm laser and a 532 nm laser, such as for certain analytical operations.

[0097] The laser assemblies 190 and 192 are coupled to single mode fibers 122 and 124 that, as described above, convert the output of the lasers to near pure Gaussian distributions. The light transmitted via the fibers 122 and 124 is input to line generator modules 94 to produce radiation lines. The beams of radiation are then directed to excitation filters 128, and combined by combiner 132 to form a combined radiation line 134. A filter wheel 13p may filter this combined radiation line, such as to block, pass or attenuate the beam as desired.

[0098] As in the embodiments described above, the filtered combined radiation line is then directed to a beam splitter 136 and therefrom to an objective 138. In the embodiment illustrated in FIG. 16, the objective is provided with an autofocus system 198 that may include one or more actuators, such as a voice coil, a linear motor stage, a piezo motor stage, or a piezo flexure stage. Sensors 200 provide for sensing the distance or focus of the system on the microarray 14, and serve to provide feedback for dynamic focusing of the confocally-directed radiation line on the appropriate depth along the microarray 14.

[0099] FIG. 16 also provides somewhat more detail regarding a presently contemplated arrangement for moving the microarray 14 prior and during scanning.
For example, a sample handling tray 202 is provided along with a motor 204 for moving the tray in and out of an imaging position. An adapter plate 206 allows for positioning of the microarray in a docking station 208. Actuators 210 provide for appropriate positioning of the microarray in the docking station. A coarse stage 212, controlled by a stepper motor 214 allows for coarse control of the position of the microarray with respect to the combined radiation line confocally directed toward the microarray. The coarse stage 212 may, for example, be used to appropriately position a portion of the microarray on which the sites are located that are to be imaged. A
precision stage 216, which may include a linear motor 218 and a linear encoder serve to provide for fine positioning and moveinent of the microarray prior to and during scanning.

[00100] As before, radiation resulting from fluorescence of individual sites on the microarray is returned through the beam splitter 136 to mirrors or other optical devices used to direct the retrobeam through bandpass filters 158, projection lenses 160 and ultimately to CCI) sensors 164 and 166.

[00101] The foregoing arrangements provide for extremely rapid and accurate imaging of multiple sites on a microarray by use of a radiation line that excites the sites simultaneously. It has been found that the confocal line scanning technique of the present invention is particularly useful in applications where sites on the microarray are spaced from one another such as to, in combination with the linear scanning described above, reduce the potential for crosstalk between returned radiation from the individual sites. FIG. 17 illustrates a presently conteinplated arrangement of sites in a hexagonal grid array to take advantage of this aspect of the confocal line scanning tecluiique of the invention.

[00102] As illustrated in FIG. 17, an array section 222 will include a plurality of sites 42 provided in a predetermined pattern. A presently contemplated embodiment provides a hexagonal packing pattern as illustrated. The pattern includes what may be termed adjacent rows or lines of sites designated by reference numerals 224 and 226 in FIG. 17. As will be appreciated by those skilled in the art, the orientation of the lines may generally be thought of with reference to the direction of scanning by the confocally directed radiation line described above. As radiation is directed along lines parallel to the site lines 224 and 226, then, a portion of the lines of sites will be illuminated by the radiation, and return a retrobeam which will be bright in those areas that fluoresce. Adjacent sites 228 and 230 in each row or line of sites will be spaced from one another, and both of these sites will be spaced from a nearest adjacent site, such as site 232 of an adjacent row or line 226. The distance between successive or adjacent lines of sites may be designated generally by reference numeral 234, such as by reference to the center of the sites in each line. It will be noted that with the hexagonal packing pattern of FIG. 17, the distance between the centers of adjacent sites in the same line, however, is greater than the distance between the adjacent lines of sites. Moreover, in the orientation of FIG. 17, the distance between centers of adjacent sites in the same line is greater than the nearest distance 236 between sites in the adjacent lines. In particular, for a hexagonal packing pattern of the type illustrated in FIG. 17, distance 234 will be approximately 0.866 (the cosine of 60 degrees) of the distance 236.

[00103] Moreover, if the sites 228, 2,30 and 232 are considered to have edges 238, these edges will be spaced from one another by a distance greater than would result if the sites were disposed in a rectilinear pattern. That is, the projection of the distance between the edges 238 of sites 228 and 232 along the axis of scanning may be denoted by reference numeral 240. The actual distance, however, between the edges will be greater, as indicated by reference numeral 242. Again, for the hexagonal pattenl illustrated in FIG. 17, the distance 242 will be approximately 15%
greater than the distance 240.

[00104] As will be appreciated by those skilled in the art, as the density of the sites on microarrays is increased, and spacing between the sites is consequently decreased, increasing demands are made on the ability to carefully focus the irradiating light beam on the sites, and to properly focus the retrobeam for imaging purposes.
The present technique provides excellent results in the ability to confocally irradiate a line of sites, where the confocality exists in the axis parallel to the width of the radiation line and not along the length of the radiation line. However, crosstalk between the sites may be considered as a relative inability to distinguish between the sites, as the images produced from high intensity sites spills over in the nonconfocal axis to neighboring sites. This can be problematic, for example, when high intensity sites are located immediately adjacent to very low intensity sites. The combination of confocal line scanning with non-rectilinearly paclced sites, in particularly in combination with hexagonally packed sites is believed to provide far superior distinction between irradiated and imaged sites, due to the reditction in crosstalk and blurring between the imaged sites.

[00105] The combination of a hexagonal arrangement of sites and the radiation line orientation set forth above is one example of an embodiment of the invention wherein the distances between nearest neighbor sites that are irradiated simultaneously by a radiation line at a first scan position is greater than the distance between nearest neighbor sites that are irradiated at different times by the scanning radiation line. It will be understood that other combinations of site packing and line orientation can also be used to achieve similar advantages. For example, although circular sites in a rectilinear grid are not packed as closely as in a hexagonal grid, the orientation for a radiation line a.nd its direction of scan can be selected for a desired reduction in cross-talk. More specifically, the radiation line can be oriented diagonally with respect to the rows and columns of sites in the rectilinear grid and the radiation line can be scanned across the grid in the diagqnal direction to achieve less cross talk between the sites than if the radiation line was oriented orthogonally with respect to the rows and columns of sites in the rectilinear grid and scanned in the orthogonal direction. An advantage being that the line is oriented such that the greatest spacing between adjacent sites occurs in the nonconfocal axis, parallel to the radiation line.

[00106] The packing arrangements described above are particularly useful when used with a radiation line that is substantially narrower than the width of the sites being irradiated. In particular embodiments, the width of the radiation line (i.e. the shorter dimension of the line) will be at most 75%, 66%, 50%, 30%, 25% or 10%
of the width of the sites being irradiated. Generally, sites having a regular shape are preferred, for example, sites having reflectional symmetry or rotational symmetry.
However, irregular shaped sites can be used if desired for a particular application.
Whether a site is regular or irregular in shape the width for the site will typically be measured at the widest dimension, for example, width is measured as the diameter of a site having a circular cross-section.

[00107] As illustrated in Figs. 18-23, a diffraction-limited line with uniform intensity distribution can be generated using a number of architectures. In one such embodiment, shown in Fig. 18, the line generator 244 can be formed with a cylindrical micro-lens array 246 and a condenser 248. A cylindrical micro-lens array 246 is used to focus the excitation beam 250 to the front focal plane of a condenser 248 in one dimension while leaving a second dimension unaffected. A
diffraction-limited line 252 with uniform intensity distribution will be generated on the back focal plane of the condenser 248. The uniformity of the line is related to the number of cylindrical micro-lenses 246 that cover the entrance pupil of the condenser 248.
The greater the number of cylindrical micro-lens arrays 246, the more uniform the line intensity distribution will be.

[00108] According to another embodiment and as shown in Fig. 19, the line generator 244 can be formed with a one-dimensional diffuser 254 and a condenser 248. A one-dimensional diffuser 254 having an angular uniformity is placed at the front focal plane of a condenser 248. The diffuser 254 fans the input collimated beam 250 in one dimension and leaves another dimension unaffected. A diffraction-limited line 252 with uniform intensity distribution will be generated on the back focal plane of the condenser 248. Since the diffuser 254 has angular uniformity, the generated line will be uniform.

[00109] In still another embodiment of the invention, an objective 256. is used as a condenser 248. Preferably, the objective lens 256 is a telecentric lens with an external pupil size of 15.75 mm. Preferably, this size is configured to match the diameter of the collimated input excitation beam 250. In addition, the input field angle of the lens is +/- 3 degrees, which corresponds to a field view of 2 mm.

[00110] Fig. 20 shows a one-dimensional diffuser 254 in use with the objective described above. As shown in Fig. 20, a one-dimensional diffuser 254 is placed at the pupil stop of the objective 256. The objective 256 diffuses the collimated input beam 250 to different angles in a certain range in one dimension and leaves another dimension unaffected. The diffuser 254 has angular uniformity, i.e. the intensities of beams diffused to different angles are the same. The lens 256 focuses the beam at each particular angle to a point in the line: The uniformity of the line is determined by the angular sensitivity of the diffuser, 254. In addition, the length of radiation line 268 is determined by the fan angle of the diffuser 254. The larger the fan angle is, the longer the generated radiation line 268 will be. If the fan angle of the diffuser 254 is +/- 3 , the generated line length will be 2 mm. Altliough the length of the radiation line 268 can be longer than 2 mm, a desired uniformity can be obtained by a line 2 mm in length.

[001111 According to another embodiment, Fig. 21 shows a cylindrical micro-lens array 246 in use with the above-described objective 256. Each cylindrical micro-lens 246 samples a portion of the collimated input beam 250, focuses it to the pupil stop of the objective 256 in one dimension, and leaves the second dimension unaffected. The cylindrical micro-lens array 246 fans the beam 250 to different angles in a certain range in one dimension. The fan angle is deterinined by the f-number of the cylindrical micro-lenses 246. The objective lens 256 focuses the beam 250 at each angle to a point in the line. Since each point in the focused line gets contribution from all the cylindrical micro-lenses 246, the uniformity of the line is related to the number of cylindrical micro-lenses 246 that covers the entrance pupil of the objective lens 256. For example, according to one embodiment of the invention, 158 micro-lenses are used to cover the pupil stop in order to generate a uniform line excitation 268.
[00112] Figs. 22 and 23 show additional embodiments of relay telescopes, configured for fluorescent imaging. A relay telescope 258 is positioned between the one-dimensional diffuser 254 (see Fig. 22) or cylindrical micro-lens array 246 (see Fig. 23) and a dichroic beam splitter 260. The dichroic beam splitter 260 is configured to separate the fluorescence imaging path (retro-beam) 262 from the excitation path 250.

[00113] A CCD camera or other detector array used in the invention can be configured for binning. Binning increases the detector array's sensitivity by summing the charges from multiple pixels in the array into one pixel. Exemplary types of binning that can be used include horizontal binning, vertical binning, or full binning.
With horizontal binning, pairs of adjacent pixels in each line of a detector array are summed. With vertical binning, pairs of adjacent pixels from two lines in the array are summed. Full binning is a combination of horizontal and vertical binning in which four adjacent pixels are summed.

[00114] Binning in the invention can be carried out with larger sets of sensor elements. As illustrated in Fig. 24(a), the line scan CCD camera and corresponding control electronics can be configured such that all pixel elements in the vertical axis are collected in a common bin and read out as a single value. Thus, binning need not be limited to adjacent pairs or adjacent groups of array elements.
Accordingly, a set of more than 2.sensor elements, such as pixels of a CCD camera, can be binned even if the set includes non-adjacent sensor elements. Non-adjacent sensor elements occur, for example, in a lineax= arrangement of 3 sensor elements where the first and third elements are separated from eacll other by the intervening second sensor element.

[00115] As shown in Fig. 24(b), in binning, all of the pixels in a row are shifted out at once after a single integration time. The advantage of this approach, wllen used in an apparatus of the invention, is that compared to a common TDI design the readout rate is less sensitive to jitter. Furthermore, the apparatus would have confocality in one axis, and the tolerance of the synchronization timing of the readout with the Y-stage movement would be reduced. Fig. 24(b) shows the projection of a 1 m laser spot on a line scan CCD camera. The projection is symmetrical in both the X
and Y-axis. Limiting the number of CCD pixels to 6 in the vertical axis creates a virtual slit in that axis. The same effect can be achieved with a TDI camera, the main requirement is that the number of pixels in the vertical axis be optimized to pass a signal while also rejecting background noise. To achieve this, the laser spot size is set to match the resolution of the system in conjunction with limiting the nuinber of vertical pixels.

[001161 An alternate embodiment of the invention uses a TDI design which limits the number of vertical pixels such that the virtual slit is still created. As shown in Fig.
24(c), in TDI, pixels are shifted in sync with the encoder output of the y-stage.
Additionally, the advantage over system designs where n=1 are that the collection efficiency of the system would be increased and the sensitivity to small optical alignment drifts would be decreased. Exemplary TDI designs and methods that can be used in the invention are described in U.S. 5,754,291, which is incorporated herein by reference.

[001171 According to another embodiment of the invention, the present scanning system architecture is configured to use parallel multi-spectral fluorescence imaging using line-scan imaging sensors. As shown in Fig. 25, radiation line 134 is used to excite fluorescent molecules in a full spectral range and a chromatic dispersion element 264 is used to spread the line fluorescence image 262 across multiple line-scan imaging sensors 266. The system can be implemented using side illumination or collinear illumination. According to this embodiment of the invention, a multi-band filter set 268 is used to excite and detect multiple fluorescent molecules. As represented in Fig. 26, each of the plurality of sensors 266 is mapped to a narrow band spectral range. The sensors 266 can be imaging sensors such as a linear line-scan CCD or a TDI line-scan CCD. Sensors are also referred to as detectors herein.

[00118J As shown in Fig. 27, according to still another embodiment of the invention, the scanning system architecture can be configured to use a multi-line illumination technique. The system can be implemented using side illumination or collinear illumination. Here, each line 268 excites a sample region at a different wavelength, for example, to excite different fluorescent molecules. The resulting multi-line fluorescence image is collected by a detector 266 with multiple line-scan imaging sensors 266. Each sensor 266 generates the corresponded fluorescent image.
Because the fluorescence with different spectral ranges is already spatially separated, no chromatic dispersion element 264 is required. A multi-notch filter 270 is used to effectively block residual Rayleigh and Raman scattered radiation.

[00119] Further, if a chromatic dispersion element is used in the system of Fig. 27, images with higher spectral resolution can be collected. As illustrated in Fig. 28, each sensor group 266 in the figure can also worlc in TDI mode to generate a single integrated image, which provides images with hierarchical spectral resolution.

[00120] The scanning system architecture can be designed to excite fluorescence of multiple dyes in different spectral ranges simultaneously. Exemplary architectures include a single line with multi-colors used in the system of Fig. 25 or spaced multi-lines with multi-colors used in the system of Fig. 27. The radiation source can be a white liglit lamp with a multi-band excitation filter or a combination of multiple lasers. The excitation filter of the multi-band filter set 268 in the system of Fig. 25 is not required, for example, if the combination of multiple lasers is used as the radiation source. In addition, the illumination can be collinear illumination (illumination shares the same objective lens 138 as the collection) as shown in Fig. 24 or slide illumination (dark field) as shown in Fig. 28. A multi-band dichroic beam splitter 136 (shown in Fig. 25) can be used for the collinear illumination and omitted for the side illumination embodiment. Also as shown in Fig. 25, a multi-band emission filter 272 of the multi band filter set 82 can be used to selectively block excitation radiation while passing fluorescence bands. For illumination with multiple lasers, a multi-notch filter 270 can also be used to selectively block excitation radiation while passing fluorescence bands, which provides even more efficient florescence detection.
[00121] According to particular embodiments of the invention, emission filters can be integrated with the image sensor 266. An exemplary orientation is shown in Fig. 29. A different orientation for blocking multi-band illumination and multiple laser illumination is shown in Figs. 30(a) and 30(b) respectively.

[00122] An apparatus or metllod of the invention is particularly useful for obtaining an image of a 2-dimensional area of a sample. Thus, if desired, detection can be substantially restricted to obtaining an image in 2 of the 3 possible dimensions for a sample. Accordingly, an image of a surface for a sample of interest can be detected or imaged. A particularly relevant sample is a microarray. Using the invention the surface of a microarray can be detected or imaged to determine one or more property of the microarray. Exemplary properties of a microarray that can be detected include, but are not limited to, the presence or absence of a label, the location of a label at a particular location such as a location where a particular probe resides, or a specific characteristic of a label such as emission of radiation at a particular wavelength or wavelength range.

[00123] Detection of such properties for a microarray can be used to determine the presence or absence of a particular target molecule in a sample contacted with the microarray. This can be determined, for example, based on binding of a labeled target analyte to a particular probe of the microarray or due to a target-dependent modification of a particular probe to incorporate, remove or alter a label at the probe location. Any one of several assays can be used to identify or characterize targets using a microarray as described, for exainple, in U.S. Pat. App. Pub. Nos.
2003/0108867, 2003/0108900, 2003/0170684, 2003/0207295, or 2005/0181394, each of which is hereby incorporated by reference.

[00124] Exemplary labels that can be detected in accordance with the invention, for example, when present on a microarray include, but are not limited to, a chromophore; luminophore; fluorophore; optically encoded nanoparticles;
particles encoded with a diffraction-grating; electrochemiluminescent label such as Ru(bpy)268+; or moiety that can be detected based on an optical characteristic.
Fluorophores that are useful in the invention include, for example, fluorescent lanthanide complexes, including those of Europium and Terbium, fluorescein, rhodamine, tetramethylrhodamine, eosin, erythrosin, coumarin, methyl-coumarins, pyrene, Malacite green, Cy3, Cy5, stilbene, Lucifer Yellow, Cascade B1ueTM, Texas Red, alexa dyes, phycoerythin, bodipy, and others known in the art such as those described in Haugland, Molecular Probes Handbook, (Eugene, OR) 6th Edition;
The Synthegen catalog (Houston, TX.), Lakowicz, Principles of Fluorescence Spectroscopy, 2nd Ed., Plenum Press New York (1999), or WO 98/59066, each of which is hereby incorporated by reference.

[00125] Any of a variety of microarrays known in the art, including, for example, those set forth elsewllere herein, can used as a sample in the invention. A
typical microarray contains sites, sometimes referred to as features, each having a population of probes. The population of probes at each site typically is homogenous, having a single species of probe but in some embodiments the populations can each be heterogeneous. Sites or features of an array are typically discrete, being separated with spaces between each other. The size of the probe sites and/or spacing between the sites can vary such that arrays can be high density, medium density or lower density. High density arrays are characterized as having sites separated by less than about 15 m. Medium density arrays have sites separated by about 15 to 30 gm, while low density arrays have sites separated by greater than 30 pm. An array useful in the invention can have sites that are separated by less than 100 m, 50 m, 10 m, m, 1 m or 0.5 m. An apparatus or method of the invention can be used to image an array at a resolution sufficient to distinguish sites at the above densities or density ranges.

[00126] Altllough the invention has been exemplified above with regard to the use of a microarray as a sample, it will be understood that other samples having features or sites at the above densities can be imaged at the resolutions set forth above. Other exemplary samples include, but are not limited to, biological specimens such as cells or tissues, electronic chips such as those used in computer processors, or the like. A
inicroarray or other sample can be placed in a sample region of an apparatus of the invention by being placed on a sample stage such as those described elsewhere herein.
[00127] An apparatus of the invention can further include a processor, operably coupled to a rectangular detector array or otherwise configured to obtain data from the rectangular detector array, wherein the processor is configured to perform a plurality of functions on the image. The processor can include a conventional or general purpose computer system that is programmed with, or otherwise has access to, one or more program modules involved in the analysis of imaging data. Exemplary computer systems that are useful in the invention include, but are not limited to personal computer systems, such as those based on Intel , IBM , or Motorola microprocessors; or work stations such as a SPARC workstation or UNIX
workstation. Useful systems include those using the Microsoft Windows , UNIX
or LINUXO operating system. The systems and methods described herein can also be implemented to run on client-server systems or wide-area networks such as the Internet.

[00128] The processor can be included in a computer system, configured to operate as either a client or server. The processor can execute instructions included in one or more program modules. Results from one or more program modules sucli as an image of a sample or sample region, or analysis of the sample or sample region can be reported to a user via a graphical user interface. For example, results can be reported via a monitor or printing device operably connected to the processor. Thus, an image of an array or other sample can be provided to a user via a graphical user interface.
[00129] According to certain aspects of the invention, several advantages are realized. The system of the present invention scans samples faster than other technologies and provides improved data qijality at lower cost. Specifically, the readout rate of the present invention is increased by a factor of n as compared to conventional TDI systems. Confocality can be achieved in one or more axis. In addition, the present invention is less sensitive to optical alignment drifts.

[00130] Further, the present invention combines the advantages of simultaneous excitation/detection of multiple fluorescent molecules using inulti-band filters and parallel readout of multiple line-scan imaging sensors on the same sample. The present invention can simultaneously generate multi-spectral fluorescence images in a fast speed. In particular embodiments, an apparatus of method of the invention can scan a sample at a rate of at least about 0.01 mm2/sec. Depending upon the particular application of the invention faster scan rates can also be used including, for exa:mple, in terms of the area scanned, a rate of at least about 0.02 mm2/sec, 0.05 mm2/sec, 0.1 mm2/sec, 1 mm2/sec, 1.5 mm2/sec, 5 mm2/sec, 10 mma/sec, 50 rrun2/sec orlOO
mm2/sec or faster. If desired, for example, to reduce noise, scan rate can have an upper limit of about 0.05 mm2/sec, 0.1 mm2/sec, 1 mm2/sec, 1.5 mm2/sec, 5 mm2/sec, mm2/sec, 50 mm2/sec orlOO mm2/sec. Scan rate can also be measured in terms of the rate of relative movement for an image and detector in the scan-axis (vertical) dimension and, can be, for example, at least about 0.1 mm/sec, 0.5 mm/sec, 1 mm/sec, 10 mm/sec, or 100 mm/sec. Again, to reduce noise, scan rate can have an upper limit of about 0.5 mm/see, 1 min/sec, 10 mm/sec, or 100 mm/sec. In sum, the present invention can be used to build multi-spectral fluorescence imagers, which are more efficient and cost-effective than other imaging systems.

[00131] The following are terms that are used in thepresent discussion, and which are intended to have the meanings ascribed below.

[00132] As used herein, the term "radiation source" is intended to mean an origin or generator of propagated electromagnetic energy. The term can include an illumination source producing electromagnetic radiation in the ultra violet (UV) range (about 200 to 390 nm), visible (VIS) range (about 390 to 770 nm), or infrared (IR) range (about 0.77 to 25 microns), or other range of the electromagnetic spectrum. A
radiation source can include, for example, a lamp such as an arc lamp or quartz halogen lamp, or a laser such as a solid state laser or a gas laser or an LED such as an LED/single mode fiber system.

[00133] As used herein, the term "excitation radiation" is intended to mean electromagnetic energy propagated toward a sample or sample region. Excitation radiation can be in a form to induce any of a variety of responses from a sample including, but not limited to, absorption of energy, reflection, fluorescence emission or luminescence.

[00134] As used herein, the term "sample region" is intended to mean a location that is to be detected. The location can be, for example, in, on or proximal to a support device that is configured to support or contain an object to be detected. A
sample can occupy a sample region permanently or temporarily such that the sample can be removed from the sample region. For example a sample region can be a location on or near a translation stage, the location being occupied by a microarray when placed on the translation stage.

[00135] As used herein, the term "detector array" is intended to mean a device or apparatus having several elements that convert the energy of contacted photons into an electrical response. An exemplary detector array is a charge coupled device (CCD), wherein the elements are photosensitive charge collection sites that accumulate charge in response to impinging photons. Further examples of detector arrays include, without limitation, a conlplementary metal oxide semiconductor (CMOS) detector array, avalanche photodiode (APD) detector array, or a Geiger-mode plioton counter detector array. The elements of a detector array can have any of a variety of arrangements. For example, a rectangular detector array has elements in a 2-dimensional, orthogonal arrangement in which a first dimension, referred to as the "horizontal" dimension is longer than a second dimension referred to as the "vertical"
dimension. A square detector array has elements in a 2-dimensional, orthogonal arrangement in which the first and second dimensions of the arrangement are the same length.

[00136] As used herein, the term "rectangular image" is intended to mean an optically formed representation of a sample, or portion of the sample, that occurs within a 2-dimensional, orthogonal region having a horizontal dimension that is longer than the vertical dimension. The rectangular image can represent the entirety of an image emanating from a sample region or; alternatively, can be a rectangular portion of a larger image, the larger image having any of a variety of shapes.

[00137] As used herein, the term "scanning device" is intended to mean a device capable of sequentially detecting different portions of a sample. A scanning device can operate, by changing the positiori of one or more component of a detection apparatus including, for example, a sample, radiation source, optical device that directs excitation radiation to a sample, optical device that directs radiation emanating from a sample, or detector array. Exemplary scanning devices include, but are not limited to a galvanQmeter configured to move a beam or line of radiation across a sample or a translation stage configured to move a sample across a beam or line of radiation.
[00138] As used herein, the term "Rayleigh resolution" is RR in the following equation RR - ((1.22)Q,)(f))/D

wherein k is wavelengtli, f is focal length and D is distance between two objects that are detected. The term is intended to be consistent with its use in the art of optics, for example, as set forth in Hecht, Optics, 4th ed., Addison Wesley, Boston MA
(2001), which is hereby incorporated by reference.

[00139] As used herein, the term "magnification" is intended to mean the ratio of the size of an object to the size of an image of the object. For example, magnification can be determined from the ratio of the size of sample region (i.e. the object) to the size of an image of the sample region at a detector array. In systems including an objective and projection lens, magnification can be determined from the ratio of focal length of the objective to back focal length of the projection lens.

[00140] As used herein, the term "radiation line" is intended to mean a collection of electromagnetic waves or particles propagated in a uniform direction, wherein the 2-dimensional cross section orthogonal to the direction of propagation is rectangular or oblong. Exemplary 2-dimensional cross sections of a radiation line include, but are not limited to, a. rectangular, elliptical, or oval shape. The cross sectional width of a radiation line can have one or both dimensions in a range of, for example, about 0.05 m to about 10 m. For example, a dimension of the radiation line can be at least about 0.05 m, 0.1 m, 0.5 m, 1 gm, 5 m or 10 gm. Furthermore, a dimension of a radiation line can be, for example, at most about 0.1 m, Q.5 m, 1 m, 5 m or 10 m. It will be understood that these dimensions are merely exemplary and radiation lines having other dimensions can be used if desired.

[00141] As used herein, the term "line generator" is intended to mean an optical element that is configured to generate a diffraction-limited or near diffraction-limited radiation line in the plane perpendicular to the optical axis of propagation with a substantially uniform intensity distribution along the horizontal axis of the line.
Exemplary line generators include, but are not limited to, a one dimensional diffuser having angular uniformity, cylindrical microlens array, diffractive element or aspheric refractive lens such as a Powell lens. The one dimensional diffuser having angular uniformity or cylindrical microlens array can be placed to direct radiation to a condenser.

[00142] As used herein, the term "beam splitter" is intended to mean an optical element that passes a first portion of a radiation beam and reflects a second portion of the beam. For example a beam splitter can be configured to selectively pass radiation in a first wavelength range and reflect radiation in a second, different radiation range.
When used for fluorescence detection the beam splitter will typically reflect the shorter wavelength excitation radiation and transmit the longer wavelength emission radiation.

[00143] As used herein, the term " external pupil" is used in reference to an objective, where the entrance pupil to the back aperture of the objective is behind the physical dimensions of the objective in the excitation beam path.

[00144] As used herein, the term "expander" is intended to mean one or more optical elements configured to adjust the diameter and collimation of a radiation beam. For exainple, an expander can be configured to increase the diameter of a radiation beani by a desired amount such as at least 2 fold, 5 fold, 10 fold or more.
Optical elements of an expander can include, for example, one or more mirrors or lenses.

[00145] As used herein, the term "projection lens" is intended to mean an optical element configured to transfer the image of an object to a detector. For example, a lens can be placed to transfer an image emanating from an objective lens to a detector array.

[00146] As used herein, the term "optical filter" is intended to mean a device for selectively passing or rejecting passage of radiation in a wavelength, polarization or frequency dependent manner. The term can include an interference filter in which multiple layers of dielectric materials pass or reflect radiation according to constructive or destructive interference between reflections from the various layers.
Interference filters are also referred to in the art as dichroic filters, or dielectric filters.
The term can include an absorptive filter which prevents passage of radiation having a selective wavelength or wavelength range by absorption. Absorptive filters include, for example, colored glass or liquid.

[00147] A filter used in the invention can have one or more particular filter transmission characteristics including, for example, bandpass, short pass and long pass. A band pass filter selectively passes radiation in a wavelength range defined by a center wavelength of maximum radiation transmission (Tmax) and a bandwidth and blocks passage of radiation outside of this range. Tmax defines the percentage of radiation transmitted at the center wavelength. The bandwidth is typically described as the full width at half maximum (FWHM) which is the range of wavelengths passed by the filter at a transmission value that is half of Tmax. A band pass filter useful in the invention can have a FWHM of 10 nanometers (nm), 20 nm, 30 nm, 40 nm or 5p nm. A long pass filter selectively passes higher wavelength radiation as defined by a Tmax and a cut on wavelength. The cut on wavelength is the wavelength at which radiation transmission is half of Tmax; as wavelength increases above the cut on wavelength, transmission percentage increases and as wavelength decreases below the cut on wavelength transmission percentage decreases. A short pass filter selectively passes lower vuavelength radiation as defined by a Tmax and a cut off wavelength.
The cut off wavelength is the wavelength at which radiation transmission is half of Tmax; as wavelength increases above the cut off wavelength, transmission percentage decreases and as wavelength decreases below the cut off wavelength transmission percentage increases. A filter of the invention can have a Tmax of 50-100%, 60-90%
or 70-80%. -[00148] As used herein, the tenn "microarray" refers to a population of different probe molecules that are attached to one or more substrates such that the different probe molecules can be differentiated from each other according to relative location.
An array can- include different probe molecules, or populations of the probe molecules, that are each located at a different addressable location on a substrate.
Alternatively, a microarray can include separate substrates each bearing a different probe molecule, or population of the probe molecules, that can be identified according to the locations of the substrates on a surface to which the substrates are attached or according to the locations of the substrates in a liquid. Exemplary arrays in which separate substrates are located on a surface include, without limitation, a Sentrix Array or Sentrix BeadChip Array available from Illumina , Inc. (San Diego, CA) or others including beads in wells such as those described in U.S. Patent Nos.
6,266,459, 6,355,431, 6,770,441, and 6,859,570; and PCT Publication No. WO
00/63437, each of which is hereby incorporated by reference. Otller arrays having particles on a surface include those set forth in US 2005/0227252; WO
05/033681;
and WO 04/024328.

[00149] Furtller examples of commercially available microarrays that can be used in the invention include, for example, an Affymetrix GeneChip microarray or other microarray synthesized in accordance with techniques sometimes referred to as VLSIPSTM (Very Large Scale Immobilized Polymer Synthesis) technologies as described, for example, in U.S. Pat. Nos. 5,324,633; 5,744,305; 5,451,683;
5,482,867;
5,491,074; 5,624,711; 5,795,716; 5,831,070; 5,856,101; 5,858,659; 5,874,219;
5,968,740; 5,974,164; , 5,981,185; 5,981,956; 6,025,601; 6,033,860; 6,090,555;
6,136,269; 6,022,963; 6,083,697; 6,291,183; 6,309,831; 6,416,949; 6,428,752 and 6,482,591, eacli of which is hereby incorporated by reference. A spotted microarray can also be used in a method of the invention. An exemplary spotted microarray is a CodeLinlcTM Array available from Amersham Biosciences. Another microarray that is useful in the invention is one that is manufactured using inkjet printing methods such as SurePrintTM Technology available from Agilent Technologies. Other microarrays that can be used in the invention include, without limitation, those described in Butte, Nature Reviews Drug Discov. 1:951-60 (2002) or U.S. Pat Nos. 5,429,807;
5,436,327; 5,561,071; 5,583,211; 5,658,734; 5,837,858; 5,919,523; 6,287,768;

6,287,776; 6,288,220; 6,297,006; 6,291,193; and 6,514,751; and WO 93/17126; WO
95/35505, each of which is hereby incorporated by reference.

[00150] As used herein, the term "time delay integration" or "TDI" is intended to mean sequential detection of different portions of a sample by different subsets of elements of a detector array, wherein transfer of charge between the subsets of elements proceeds at a rate synchronized with and in the same direction as the apparent motion of the sample being imaged. For example, TDI can be carried out by scanning a sample such that a frame transfer device produces a continuous video image of the sample by means of a stack of linear arrays aligned with and synchronized to the apparent movement of the sample, whereby as the image moves from one line to the next, the stored charge moves along with it. Accumulation of charge can integrate during the entire time required for the row of charge to move from one end of the detector to the serial register (or to the storage area of the device, in the case of a frame transfer CCD).

[00151] As used herein, the term "collection arm" is intended to mean an optical component or set of optical components positioned to direct radiation from a sample region to a detector.

[00152] While only certain features of the invention have been illustrated and described herein, many modifications and changes will occur to those skilled in the art. It is, tlierefore, to be understood that the appended claims are intended to cover all such modifications and changes as fall within the true spirit of the invention.

Claims (88)

CLAIMS:
1. An imaging apparatus comprising:
(a) a radiation source positioned to send excitation radiation to at least a portion of a sample region;
(b) a rectangular detector array;
(c) imaging optics positioned to direct a rectangular image of said portion to said rectangular detector array; and (d) a scanning device configured to scan said sample region in a scan-axis dimension, whereby the portion of said sample region that forms a rectangular image at said rectangular detector array is changed, wherein the shorter of the two rectangular dimensions for said rectangular detector array and the shorter of the two rectangular dimensions for said image are in said scan-axis dimension, and wherein said shorter of the two rectangular dimensions for said rectangular detector array is short enough to achieve confocality in a single axis of said rectangular detector array, wherein said single axis is said shorter of the two rectangular dimensions for said rectangular detector array.
2. The apparatus of claim 1, wherein the ratio of said shorter of the two rectangular dimensions for said rectangular detector array and the product of the Rayleigh resolution of the imaging optics multiplied by the magnification of the imaging optics is in the range of 0.1 to 10.
3. The apparatus of claim 1, further comprising a line generator positioned to receive excitation radiation from said radiation source and to send a radiation line to said sample region.
4. The apparatus of claim 3, further comprising an objective positioned to receive said radiation line therethrough to illuminate said sample region.
5. The apparatus of claim 4, wherein said imaging optics comprise said objective, wherein said objective is further positioned to collect radiation emanating from said sample region, wherein said radiation emanating from said sample region forms said rectangular image that is directed to said rectangular detector array.
6. The apparatus of claim 5, further comprising a beam splitter positioned to separate said radiation line from said radiation emanating from said sample region and to direct said radiation emanating from said sample region to the rectangular detector array.
7. The apparatus of claim 4, wherein said objective has an external pupil positioned to receive said radiation line therethrough to illuminate said sample region.
8. The apparatus of claim 4, further comprising a first expander positioned to receive excitation radiation from said radiation source and to send an expanded beam of said radiation to said line generator.
9. The apparatus of claim 8, further comprising a second expander positioned to receive said excitation radiation from said line generator and send an expanded beam of said radiation to said objective, wherein said second expander is further configured to decrease the field angle of said radiation line.
10. The apparatus of claim 4, wherein said objective has a property selected from the group consisting of color correction, high numerical aperture, telecentricity, and afocality at the backplane.
11. The apparatus of claim 3, wherein said line generator has a full fan angle of six degrees and is configured to receive an input beam having a diameter of at most 4 mm.
12. The apparatus of claim 3, wherein said line generator further comprises a cylindrical micro-lens array, one-dimensional diffuser having an angular uniformity, aspheric refractive lens, diffractive element or Powell lens.
13. The apparatus of claim 3, wherein said line generator further comprises a diffractive element to generate a diffraction-limited line with uniform intensity distribution.
14. The apparatus of claim 3, wherein the shorter of two rectangular dimensions for said radiation line is short enough to achieve confocality in a single axis of said rectangular detector array, wherein said single axis is said shorter of the two rectangular dimensions for said rectangular detector array.
15. The apparatus of Claim 3, wherein the ratio of the shorter of two rectangular dimensions for said radiation line to the quotient of said shorter of the two rectangular dimensions divided by the magnification of the imaging optics is in the range of 0.1 to 10.
16. The apparatus of Claim 3, wherein the ratio of the shorter of two rectangular dimensions for said radiation line to the Rayleigh resolution of the imaging optics is in the range of 0.1 to 10.
17. The apparatus of claims 3, wherein the ratio of the 1/e~2 width of the shorter of two rectangular dimensions for said radiation line to the quotient of said shorter of the two rectangular dimensions for said rectangular detector array divided by the magnification of the imaging optics is in the range of 0.5 to 2.
18. The apparatus of claim 1, further comprising a projection lens positioned to collect radiation emanating from said sample region, wherein said radiation emanating from said sample region forms said rectangular image that is directed to said rectangular detector array.
19. The apparatus of claim 1, further comprising a band pass filter positioned to collect radiation emanating from said sample region, wherein said radiation emanating from said sample region forms said rectangular image that is directed to said rectangular detector array.
20. The apparatus of claim 1, further comprising an emission filter positioned to collect radiation emanating from said sample region, wherein said radiation emanating from said sample region forms said rectangular image that is directed to said rectangular detector array.
21. The apparatus of claim 1, further comprising a translation stage positioned to provide a sample to said sample region.
22. The apparatus of claim 21, wherein said translation stage is configured to move said sample in said scan-axis dimension.
23. The apparatus of claim 21, further comprising a microarray supported by said translation stage, whereby said array is provided to said sample region.
24. The apparatus of claim 1, wherein said rectangular detector array is configured for TDI (Time Delay Integration) operation.
25. The apparatus of claim 1, wherein said rectangular detector array comprises a line scan CCD camera, CMOS detector array, avalanche photodiode (APD) array, or Geiger-mode photon counter array.
26. The apparatus of claim 1, wherein the aspect ratio of said rectangular detector is greater than 20.
27. The apparatus of claim 1, wherein said radiation source comprises at least one laser.
28. The apparatus of claim 1, comprising multiple collection arms positioned to collect radiation emanating from said sample region, wherein said radiation emanating from said sample region forms multiple rectangular images that are directed to multiple rectangular detector arrays.
29. The apparatus of claim 1, wherein said apparatus is configured to obtain an image of said sample comprising a Rayleigh resolution between 0.2 and 10 micrometers.
30. A method of obtaining an image of a sample, comprising (a) contacting at least a first portion of a sample with excitation radiation under conditions wherein radiation is emanated from said first portion;
(b) directing said radiation emanated from said first portion to form a rectangular image of said first portion at a rectangular detector array; and (c) scanning said sample region in a scan-axis dimension, thereby repeating steps (a) and (b) to form a rectangular image of a second portion of said sample at said rectangular detector array, wherein the shorter of the two rectangular dimensions for said rectangular detector array and the shorter of the two rectangular dimensions for said images are in said scan-axis dimension, and wherein said shorter of the two rectangular dimensions for said rectangular detector array is short enough to achieve confocality in a single axis of said rectangular detector array, wherein said single axis is said shorter of the two rectangular dimensions for said rectangular detector array.
31. The method of claim 30, wherein the ratio of said shorter of the two rectangular dimensions for said rectangular detector array and the product of the Rayleigh resolution of said rectangular image multiplied by the magnification of the rectangular image is in the range of 0.1 to 10.
32. The method of claim 30, wherein said excitation radiation that contacts at least a portion of said sample comprises a radiation line.
33. The method of claim 32, wherein the shorter of two rectangular dimensions for said radiation line is short, enough to achieve confocality in a single axis of said rectangular detector array, wherein said single axis is said shorter of the two rectangular dimensions for said rectangular detector array.
34. The method of claim 32, wherein the ratio of the shorter of two rectangular dimensions for said radiation line to the quotient of said shorter of the two rectangular dimensions for said rectangular detector array divided by the magnification is in the range of 0.1 to 10.
35. The method of claim 30, wherein the ratio of the 1/e~2 width the shorter of two rectangular dimensions for said radiation line to the quotient of said shorter of the two rectangular dimensions for said rectangular detector array divided by the magnification is in the range of 0.5 to 2.
36. The method of claim 30, wherein said scanning said sample comprises moving said sample, thereby changing the relative positions of said rectangular image and said rectangular detector array in said scan-axis dimension.
37. The method of claim 30, wherein said scanning comprises TDI (Time Delay Integration).
38. The method of claim 30, wherein all pixel elements in the shorter of the two rectangular dimensions for said rectangular detector array are collected in a common bin and read out as a single value.
39. The method of claim 30, wherein said excitation radiation comprises radiation in a range selected from the group consisting of UV radiation, VIS
radiation and IR radiation.
40. The method of claim 30, further comprising storing a data representation of said image of said sample in a computer readable memory.
41. The method of claim 40, further comprising displaying a graphical representation of said image of said sample on a monitor operably connected to said computer readable memory.
42. The method of claim 30, wherein said sample comprises a microarray having a plurality of individual sites.
43. The method of claim 42, wherein said individual sites are separated by a distance in the range of 0.1 to 50 micrometers.
44. The method of claim 43, further comprising distinguishing said individual sites.
45. The method of claim 30, wherein said image of said sample comprises a Rayleigh resolution between 0.2 and 10 micrometers.
46. A method of configuring a scanner to achieve confocality in a single axis, comprising, (a) providing an apparatus comprising (i) a radiation source positioned to send excitation radiation to at least a portion of a sample region;
(ii) a rectangular detector array;
(iii) imaging optics positioned to direct a rectangular image of said portion to said rectangular detector array; and (iv) a scanning device configured to scan said sample region in a scan-axis dimension, whereby the portion of said sample region that forms a rectangular image at said rectangular detector array is changed, wherein the shorter of the two rectangular dimensions for said rectangular detector array and the shorter of the two rectangular dimensions for said image are in said scan-axis dimension; and (b) positioning said rectangular detector array or said imaging optics to restrict said shorter of the two rectangular dimensions for said rectangular detector array to be short enough to achieve confocality in a single axis of said rectangular detector array, wherein said single axis is said shorter of the two rectangular dimensions for said rectangular detector array.
47. A system for imaging a microarray comprising:
a laser light source;
a single mode fiber optic cable coupled to the laser light source for transmitting laser light in single mode transmission;
a line illuminator for converting laser light from the source to a line of radiation, the line illuminator including a collimator arranged to receive the laser light from the source and an aspherical lens for converting collimated light from the collimator to the line of radiation; and a focusing device for directing the line of radiation onto a plane at the surface of a microarray.
48. The system of claim 47, wherein the line illuminator is configured to convert laser light from the source to a line of radiation of substantially uniform intensity over a desired line length.
49. The system of claim 47, wherein the collimator and aspherical lens are prealigned within a module to permit installation of the module in the system without further alignment within the module during installation.
50. The system of claim 47, wherein the focusing device is configured to create a uniform line of radiation that is diffraction limited in the narrow dimension of the line.
51. The system of claim 47, further comprising a laser line filter disposed in the module intermediate the collimator and the aspherical lens.
52. The system of claim 47, wherein the aspherical lens is a Powell lens.
53. The system of claim 47, wherein the aspherical lens is a cylindrical lens.
54. The system of claim 47, wherein individual sites on the microarray are separated by a distance in the range of about 0.1 to 50 micrometers.
55. The system of claim 54, wherein the system is configured to distinguish the individual sites.
56. The system of claim 47, wherein the system is configured to obtain an image of the microarray at a Rayleigh resolution between about 0.2 and 10 micrometers.
57. The system of claim 47, wherein the line illuminator includes an integral connector, and the single mode fiber optic cable is terminated with a mating connector for coupling to the line illuminator.
58. The system of claim 47, wherein one end of the single mode fiber optic cable is integrally coupled to the line illuminator.
59. The system of claim 47, wherein one end of the single mode fiber optic cable is removably coupled to the line illuminator.
60. The system of claim 47, comprising a second fiber optic cable coupled to the laser light source at one end thereof and coupled to the single mode fiber optic cable at an opposite end thereof.
61. The system of claim 60, wherein the second fiber optic cable is a single mode fiber optic cable.
62. The system of claim 61, wherein the fiber optic cables are coupled to one another via an optical connector.
63. The system of claim 61, wherein the fiber optic cables are spliced to one another.
64. The system of claim 47, wherein the single mode fiber optic cable is configured for single mode transmission of laser light with a wavelength of approximately 405 nm, 488 nm, 532 nm or 633 nm.
65. A system for imaging a microarray comprising:
a laser light source;
a fiber optic cable coupled to the laser light source;
a line illuminator for converting laser light from the source to a line of radiation, the line illuminator including a collimator arranged to receive the laser light from the source and an aspherical lens for converting collimated light from the collimator to the line of radiation;
a second fiber optic cable coupled to the first single mode fiber optic cable at one end thereof and to the line illuminator at another end thereof, at least one of the fiber optic cables being a single mode fiber optic cable; and a focusing device for directing the line of radiation onto a plane at the surface of a microarray.
66. A system for imaging a microarray comprising:
first and second laser light sources, each source configured to output laser light in a different predetermined frequency band;
first and second single mode fiber optic cables coupled to the first and second laser light sources, respectively, for transmitting laser light in single mode transmission; and first and second line illuminators coupled to the first and second single mode fiber optic cables, respectively, for converting laser light from the respective source to a line of radiation, the line illuminators each including a collimator arranged to receive the laser light from the source and an aspherical lens for converting collimated light from the collimator to the line of radiation; and a focusing device for directing the line of radiation onto a plane at the surface of a microarray.
67. The system of claim 66, further comprising:
a combiner for combining the lines of radiation from the first and second illuminators;
means for confocally irradiating the microarray with the combined lines of radiation and for returning radiation from the microarray; and a detector for receiving the returned radiation and for generating signals for use in analysis of the microarray.
68. A method for making a system for imaging a microarray comprising:
placing a laser light source, a line illuminator, a focusing device and a stage in a configuration wherein:
the laser light source is coupled to the line illuminator via a single mode fiber optic cable, the fiber optic cable configured for transmitting laser light from the source in single mode transmission, and the line illuminator configured for converting laser light from the source to a line of radiation, the line illuminator includes a collimator arranged to receive the laser light from the source and an aspherical lens for converting collimated light from the collimator to the line of radiation;
the focusing device directs the line of radiation toward a plane; and the stage is configured to place a microarray surface at the plane.
69. A method for imaging a microarray comprising:
generating laser light;

transmitting the laser light to a line illuminator via a single mode fiber optic cable, the fiber optic cable configured for transmitting laser light from the source in single mode transmission, and the line illuminator configured for converting laser light from the source to a line of radiation, the line illuminator including a collimator arranged to receive the laser light from the source and an aspherical lens for converting collimated light from the collimator to the line of radiation; and directing the line of radiation with a focusing device, wherein the line of radiation is directed onto a plane at the surface of a microarray.
7Q. A method for imaging a microarray comprising:
generating laser light at first and second wavelengths;
transmitting the first and second wavelength laser light to respective line illuminators via respective first and second single mode fiber optic cables, each fiber optic cable configured for transmitting laser light from the respective source in single mode transmission, and each line illuminator configured for converting laser light from the respective source to a line of radiation, each line illuminator including a collimator arranged to receive the laser light from the source and an aspherical lens for converting collimated light from the collimator to the line of radiation;
combining the lines of radiation from the line illuminators;
directing the combined lines of radiation with a focusing device, wherein the line of radiation is directed onto a plane at the surface of a microarray, thereby confocally irradiating the microarray with the combined lines of radiation;
and directing radiation returned from the microarray to a detector configured to generate signals for analysis of the microarray.
71. A method for analyzing an array of discrete sites, comprising:
simultaneously detecting a first plurality of sites in a first line of the array;
and simultaneously detecting a second plurality of sites in a second line of the array;
wherein the second line is generally parallel to the first line, wherein a distance D separates the first line and the second line when passing through nearest edges of the first and second plurality of sites, and wlierein the distance between the nearest edges of adjacent sites within each of the plurality of sites is greater than D.
72. The method of claim 71, wherein the array comprises a biological microarray.
73. The method of claim 71, wherein the array comprises discrete sites on a surface.
74. The method of claim 71, wherein the first plurality of sites is simultaneously irradiated with a radiation line.
75. The method of claim 74, wherein the radiation line is moved across the array to sequentially irradiate the first plurality of sites and the second plurality of sites.
76. The method of claim 74, wherein the first plurality of sites is sequentially irradiated with radiation, and wherein return radiation from the sites forms a line at a detector.
77. The method of claim 74, wherein the radiation line has a width less than D.
78. The method of claim 74, wherein the radiation line is confocally directed to the sites along the first line and the second line.
79. The method of claim 74, wherein the radiation line is substantially continuous along a desired length extending over the plurality of sites.
80. The method of claim 71, wherein the sites have a generally symmetric shape.
81. The method of claim 71, wherein the sites are arranged in a generally hexagonal grid, and wherein the first and second lines are parallel to lines of the grid.
82. The method of claim 71 further comprising returning radiation from the sites to a detector that generates signals for analysis of the sites.
83. A method for analyzing an array of discrete sites comprising:
sequentially irradiating a series of lines of the sites, each line being irradiated with a radiation line;
wherein the distance between the nearest edges of adjacent sites in each of the adjacent lines of sites is greater than the distance between parallel lines passing through the nearest edges of the sites in the adjacent lines of sites.
84. A method for analyzing an array having discrete sites comprising:
(a) irradiating a line of the sites with radiation;
(b) repeating step (a) for a plurality of lines of sites; and (c) returning radiation from each of the lines of sites to a detector that generates signals for analysis of the sites;
wherein the sites are disposed in a non-rectilinear grid on the array surface, whereby the distance between the nearest edges of adjacent sites in each line of sites is greater than the distance between parallel lines passing through the nearest edges of the sites in adjacent lines of the plurality of lines of sites, and wherein an image detected by the detector is confocal in the axis orthogonal to the axes along the parallel lines.
85. A system for analyzing an array of discrete sites, the system configured to:
simultaneously detect a first plurality of sites in a first line of the array;
and simultaneously detect a second plurality of sites in a second line of the array, wherein the second line is generally parallel to the first line, wherein a distance D separates the first line and the second line when passing through the nearest edges of the first and second plurality of sites, and wherein the distance between the nearest edges of adjacent sites within each of the plurality of sites is greater than D.
86. A system for analyzing an array of discrete sites, the system configured to:
sequentially irradiate a series of lines of the sites, each line being irradiated with a radiation line;
wherein the distance between the nearest edges of adjacent sites in each of the adjacent lines of sites is greater than the distance between parallel lines passing through the nearest edges of the sites in the adjacent lines of sites.
87. A system for analyzing an array of discrete sites, the system configured to:
(a) simultaneously irradiate a plurality of sites with a line of radiation;
(b) repeate step (a) for a plurality of lines of sites; and (c) return radiation from each of the pluralities of sites to a detector that generates signals for analysis of the sites;
wherein the sites are disposed in a non-rectilinear grid on the array surface, wherein the distance between the nearest edges of adjacent sites in each line of sites is greater than the distance between parallel lines passing through the nearest edges of the sites in adjacent lines of the plurality of lines of sites, and wherein an image detected by the detector is confocal in the axis orthogonal to the axes along the parallel lines.
88. A system for analyzing an array of discrete sites, the system configured to:
(a) sequentially irradiate a plurality of sites with radiation, wherein said sites are disposed along a line, wherein the return radiation from the plurality of sites forms a line at a detector;
(b) repeat step (a) for a plurality of lines of the sites; and (c) return radiation from each of the pluralities of sites to a detector that generates signals for analysis of the sites;
wherein the sites are disposed in a non-rectilinear grid on the array surface, wherein the distance between the nearest edges of adjacent sites in each line of sites is greater than the distance between parallel lines passing through the nearest edges of the sites in adjacent lines of the plurality of lines of sites, and wherein an image detected by the detector is confocal in the axis orthogonal to the axes along the parallel lines.
CA2632221A 2005-11-23 2006-11-21 Confocal imaging methods and apparatus Active CA2632221C (en)

Applications Claiming Priority (3)

Application Number Priority Date Filing Date Title
US11/286,309 US7329860B2 (en) 2005-11-23 2005-11-23 Confocal imaging methods and apparatus
US11/286,309 2005-11-23
PCT/US2006/045058 WO2007062039A2 (en) 2005-11-23 2006-11-21 Confocal imaging methods and apparatus

Publications (2)

Publication Number Publication Date
CA2632221A1 true CA2632221A1 (en) 2007-05-31
CA2632221C CA2632221C (en) 2013-08-27

Family

ID=37808340

Family Applications (1)

Application Number Title Priority Date Filing Date
CA2632221A Active CA2632221C (en) 2005-11-23 2006-11-21 Confocal imaging methods and apparatus

Country Status (9)

Country Link
US (6) US7329860B2 (en)
EP (2) EP1955102B1 (en)
JP (1) JP5055292B2 (en)
CN (1) CN101361015B (en)
CA (1) CA2632221C (en)
DK (2) DK1955102T3 (en)
ES (2) ES2635094T3 (en)
PL (1) PL2594981T3 (en)
WO (1) WO2007062039A2 (en)

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN108519329A (en) * 2018-03-26 2018-09-11 华中科技大学 A kind of line co-focusing imaging device of multi-channel scanning and detection
US10823612B2 (en) 2015-01-30 2020-11-03 Japan Science And Technology Agency Multifocal spectrometric measurement device, and optical system for multifocal spectrometric measurement device

Families Citing this family (264)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20040082863A1 (en) * 2002-03-15 2004-04-29 Mcgreevy James Device and method for the photodynamic diagnosis of tumor tissue
EP2703871A3 (en) 2005-05-25 2014-09-03 Massachusetts Institute Of Technology Multifocal scanning microscopy systems and methods
US7329860B2 (en) 2005-11-23 2008-02-12 Illumina, Inc. Confocal imaging methods and apparatus
US7995202B2 (en) * 2006-02-13 2011-08-09 Pacific Biosciences Of California, Inc. Methods and systems for simultaneous real-time monitoring of optical signals from multiple sources
US7715001B2 (en) * 2006-02-13 2010-05-11 Pacific Biosciences Of California, Inc. Methods and systems for simultaneous real-time monitoring of optical signals from multiple sources
US7813013B2 (en) * 2006-11-21 2010-10-12 Illumina, Inc. Hexagonal site line scanning method and system
US8315817B2 (en) * 2007-01-26 2012-11-20 Illumina, Inc. Independently removable nucleic acid sequencing system and method
EP2126766A2 (en) 2007-01-26 2009-12-02 Illumina Inc. Image data efficient genetic sequencing method and system
US7687776B2 (en) * 2007-04-11 2010-03-30 General Monitors, Inc. Gas and/or flame imaging system with explosion proof housing
EP2201352B2 (en) 2007-09-28 2018-08-29 Illumina, Inc. Fluorescence excitation and detection system and method
JP5072688B2 (en) * 2008-04-02 2012-11-14 キヤノン株式会社 Scanning imaging device
CN102084003A (en) * 2008-04-04 2011-06-01 生命科技公司 Scanning system and method for imaging and sequencing
US8039817B2 (en) 2008-05-05 2011-10-18 Illumina, Inc. Compensator for multiple surface imaging
US8198028B2 (en) 2008-07-02 2012-06-12 Illumina Cambridge Limited Using populations of beads for the fabrication of arrays on surfaces
US20100087325A1 (en) * 2008-10-07 2010-04-08 Illumina, Inc. Biological sample temperature control system and method
US7675045B1 (en) * 2008-10-09 2010-03-09 Los Alamos National Security, Llc 3-dimensional imaging at nanometer resolutions
EP2335111A4 (en) * 2008-10-09 2013-12-25 Ge Healthcare Bio Sciences A system and method for adjusting a beam expander in an imaging system
US8541207B2 (en) 2008-10-22 2013-09-24 Illumina, Inc. Preservation of information related to genomic DNA methylation
KR101061004B1 (en) * 2008-12-10 2011-09-01 한국전기연구원 Device for photodynamic therapy and light detection
US20100157086A1 (en) * 2008-12-15 2010-06-24 Illumina, Inc Dynamic autofocus method and system for assay imager
CA2760439A1 (en) 2009-04-30 2010-11-04 Good Start Genetics, Inc. Methods and compositions for evaluating genetic markers
US8222589B2 (en) * 2009-05-29 2012-07-17 General Electric Company Solid-state photomultiplier module with improved signal-to-noise ratio
JP5568912B2 (en) * 2009-07-22 2014-08-13 富士ゼロックス株式会社 Light-emitting element head characteristic measuring apparatus and light-emitting element head light quantity correction method
US20110017915A1 (en) * 2009-07-23 2011-01-27 Palo Alto Research Center Incorporated Drift scanner for rare cell detection
WO2011011175A2 (en) 2009-07-24 2011-01-27 Illumina, Inc. Method for sequencing a polynucleotide template
EP2293032A1 (en) * 2009-09-04 2011-03-09 Radisens Diagnostic Limited An Integrated Cytometric Sensor System and Method
KR101075155B1 (en) 2009-10-07 2011-10-19 한국과학기술원 Apparatus and method for confocal laser scanning microscopy system using line scanning way
FI20096067A0 (en) * 2009-10-15 2009-10-15 Valtion Teknillinen Measurement of Raman radiation
DE102009057304A1 (en) * 2009-12-07 2011-06-09 Leica Microsystems Cms Gmbh Apparatus for examining a sample comprising a microscope
DE102009058295A1 (en) * 2009-12-10 2011-06-16 Biostep Gmbh Trans-illuminator for e.g. rear illumination of fluorescent material sample for qualitative and quantitative evaluation of sample in research laboratory, has adjustment component for adjustment of specific wave length
US8470733B2 (en) * 2009-12-22 2013-06-25 Zih Corp. Direct thermal media and registration sensor system and method for use in a color thermal printer
US8422031B2 (en) * 2010-02-01 2013-04-16 Illumina, Inc. Focusing methods and optical systems and assemblies using the same
DE202011003570U1 (en) 2010-03-06 2012-01-30 Illumina, Inc. Systems and apparatus for detecting optical signals from a sample
US8532398B2 (en) 2010-03-26 2013-09-10 General Electric Company Methods and apparatus for optical segmentation of biological samples
ITRM20100286A1 (en) 2010-05-28 2011-11-29 Consiglio Nazionale Ricerche SPECTRAL CONFOCAL MICROSCOPE IN REFLECTION WITH A WIDE BAND.
US9029103B2 (en) 2010-08-27 2015-05-12 Illumina Cambridge Limited Methods for sequencing polynucleotides
US8575071B2 (en) 2010-11-03 2013-11-05 Illumina, Inc. Reducing adapter dimer formation
US8773526B2 (en) 2010-12-17 2014-07-08 Mitutoyo Corporation Edge detection using structured illumination
US9163281B2 (en) 2010-12-23 2015-10-20 Good Start Genetics, Inc. Methods for maintaining the integrity and identification of a nucleic acid template in a multiplex sequencing reaction
US8951781B2 (en) 2011-01-10 2015-02-10 Illumina, Inc. Systems, methods, and apparatuses to image a sample for biological or chemical analysis
US10908403B2 (en) * 2011-02-14 2021-02-02 European Molecular Biology Laboratory (Embl) Light-pad microscope for high-resolution 3D fluorescence imaging and 2D fluctuation spectroscopy
US9291802B2 (en) * 2011-04-29 2016-03-22 Corning Incorporated Compact label free imaging system
US20120306998A1 (en) * 2011-06-01 2012-12-06 Merrill Ii Dennis E Macro Area Camera for an Infrared (IR) Microscope
US10378051B2 (en) 2011-09-29 2019-08-13 Illumina Cambridge Limited Continuous extension and deblocking in reactions for nucleic acids synthesis and sequencing
WO2013058907A1 (en) 2011-10-17 2013-04-25 Good Start Genetics, Inc. Analysis methods
EP2771103B1 (en) 2011-10-28 2017-08-16 Illumina, Inc. Microarray fabrication system and method
KR101302162B1 (en) * 2011-11-04 2013-09-10 광주과학기술원 3-dimensional confocal electroluminescence spectral-microscope apparatus
WO2013070627A2 (en) 2011-11-07 2013-05-16 Illumina, Inc. Integrated sequencing apparatuses and methods of use
EP2753967B1 (en) * 2011-11-14 2023-04-12 Koninklijke Philips N.V. Optical microscopy probe for scanning microscopy of an associated object
US9200274B2 (en) 2011-12-09 2015-12-01 Illumina, Inc. Expanded radix for polymeric tags
WO2013117595A2 (en) 2012-02-07 2013-08-15 Illumina Cambridge Limited Targeted enrichment and amplification of nucleic acids on a support
EP2816388A4 (en) * 2012-02-15 2015-07-22 Olympus Corp Laser scanning-type viewing device
US20130250088A1 (en) * 2012-03-22 2013-09-26 Molecular Devices, Llc Multi-color confocal microscope and imaging methods
CN102638659B (en) * 2012-03-28 2014-05-14 西安电子科技大学 High-resolution imaging system and method based on CMOS-TDI (Complementary Metal Oxide Semiconductor-Time Delay and Integration) mode
CA3138752C (en) 2012-04-03 2024-02-06 Illumina, Inc. Integrated optoelectronic read head and fluidic cartridge useful for nucleic acid sequencing
US8209130B1 (en) 2012-04-04 2012-06-26 Good Start Genetics, Inc. Sequence assembly
US8812422B2 (en) 2012-04-09 2014-08-19 Good Start Genetics, Inc. Variant database
US20130274148A1 (en) 2012-04-11 2013-10-17 Illumina, Inc. Portable genetic detection and analysis system and method
US10227635B2 (en) 2012-04-16 2019-03-12 Molecular Loop Biosolutions, Llc Capture reactions
RU2616653C2 (en) * 2012-06-05 2017-04-18 Хайпермед Имэджинг, Инк. Methods and device for coaxial image forming with multiple wavelengths
NL2017959B1 (en) 2016-12-08 2018-06-19 Illumina Inc Cartridge assembly
CA2881823C (en) 2012-08-20 2019-06-11 Illumina, Inc. Method and system for fluorescence lifetime based sequencing
US9116139B2 (en) 2012-11-05 2015-08-25 Illumina, Inc. Sequence scheduling and sample distribution techniques
WO2014074611A1 (en) 2012-11-07 2014-05-15 Good Start Genetics, Inc. Methods and systems for identifying contamination in samples
JP5388078B1 (en) 2012-11-14 2014-01-15 レーザーテック株式会社 Analysis apparatus and analysis method
US9426400B2 (en) * 2012-12-10 2016-08-23 Kla-Tencor Corporation Method and apparatus for high speed acquisition of moving images using pulsed illumination
US20140168402A1 (en) * 2012-12-13 2014-06-19 Vala Sciences, Inc. Continuous-Scanning Image Acquisition in Automated Microscopy Using Reflective Autofocus
CN103018911B (en) * 2012-12-14 2015-06-24 清华大学 Optical fiber laser synthesizer based on wavelength division multiplexing
US9805407B2 (en) 2013-01-25 2017-10-31 Illumina, Inc. Methods and systems for using a cloud computing environment to configure and sell a biological sample preparation cartridge and share related data
US8842273B2 (en) 2013-02-14 2014-09-23 United Sciences, Llc Optical measurement of drilled holes
EP2971159B1 (en) 2013-03-14 2019-05-08 Molecular Loop Biosolutions, LLC Methods for analyzing nucleic acids
US9193998B2 (en) 2013-03-15 2015-11-24 Illumina, Inc. Super resolution imaging
US20140274747A1 (en) 2013-03-15 2014-09-18 Illumina, Inc. Super resolution imaging
CN103257438B (en) * 2013-05-29 2015-02-18 哈尔滨工业大学 Plane two-dimension rectangular scanning device based on automatic-control electric translation stage and scanning method thereof
WO2014197377A2 (en) 2013-06-03 2014-12-11 Good Start Genetics, Inc. Methods and systems for storing sequence read data
CN105358715B (en) 2013-07-03 2018-09-18 伊鲁米那股份有限公司 Orthogonal synthesis is sequenced
US9324900B2 (en) * 2013-08-01 2016-04-26 Teledyne Scientific & Imaging, Llc Method of fabricating a superlattice structure
US9116866B2 (en) 2013-08-21 2015-08-25 Seven Bridges Genomics Inc. Methods and systems for detecting sequence variants
US9898575B2 (en) 2013-08-21 2018-02-20 Seven Bridges Genomics Inc. Methods and systems for aligning sequences
JP3206040U (en) 2013-08-28 2016-09-01 イルミナ インコーポレイテッド Optical alignment tool
US9188775B2 (en) 2013-08-28 2015-11-17 United Sciences, Llc Optical scanning and measurement
EP3038834B1 (en) 2013-08-30 2018-12-12 Illumina, Inc. Manipulation of droplets on hydrophilic or variegated-hydrophilic surfaces
US9352315B2 (en) 2013-09-27 2016-05-31 Taiwan Semiconductor Manufacturing Company, Ltd. Method to produce chemical pattern in micro-fluidic structure
CA2927637A1 (en) 2013-10-18 2015-04-23 Seven Bridges Genomics, Inc. Methods and systems for identifying disease-induced mutations
WO2015058120A1 (en) 2013-10-18 2015-04-23 Seven Bridges Genomics Inc. Methods and systems for aligning sequences in the presence of repeating elements
US10832797B2 (en) 2013-10-18 2020-11-10 Seven Bridges Genomics Inc. Method and system for quantifying sequence alignment
AU2014337089B2 (en) 2013-10-18 2019-08-08 Seven Bridges Genomics Inc. Methods and systems for genotyping genetic samples
US10851414B2 (en) 2013-10-18 2020-12-01 Good Start Genetics, Inc. Methods for determining carrier status
EP3058096A1 (en) 2013-10-18 2016-08-24 Good Start Genetics, Inc. Methods for assessing a genomic region of a subject
US9092402B2 (en) 2013-10-21 2015-07-28 Seven Bridges Genomics Inc. Systems and methods for using paired-end data in directed acyclic structure
US10540783B2 (en) 2013-11-01 2020-01-21 Illumina, Inc. Image analysis useful for patterned objects
WO2015100421A1 (en) * 2013-12-24 2015-07-02 Tissuevision, Inc. Multi-foci multiphoton imaging systems and methods
WO2015103225A1 (en) 2013-12-31 2015-07-09 Illumina, Inc. Addressable flow cell using patterned electrodes
AU2015204819B2 (en) 2014-01-10 2021-05-06 Seven Bridges Genomics Inc. Systems and methods for use of known alleles in read mapping
US9817944B2 (en) 2014-02-11 2017-11-14 Seven Bridges Genomics Inc. Systems and methods for analyzing sequence data
WO2015138648A1 (en) 2014-03-11 2015-09-17 Illumina, Inc. Disposable, integrated microfluidic cartridge and methods of making and using same
KR101476820B1 (en) * 2014-04-07 2014-12-29 주식회사 썸텍 3D video microscope
FR3019901B1 (en) 2014-04-09 2020-10-30 Bio Rad Innovations CONTROL MARKER FOR THE IMPLEMENTATION OF ANALYSIS METHODS ON SPOTS
WO2015175530A1 (en) 2014-05-12 2015-11-19 Gore Athurva Methods for detecting aneuploidy
AU2015259024B2 (en) 2014-05-16 2021-07-01 Illumina, Inc. Nucleic acid synthesis techniques
CA2949984C (en) 2014-05-27 2021-10-19 Illumina, Inc. Systems and methods for biochemical analysis including a base instrument and a removable cartridge
GB201409777D0 (en) 2014-06-02 2014-07-16 Illumina Cambridge Ltd Methods of reducing density-dependent GC bias in isothermal amplification
JP6408796B2 (en) * 2014-06-11 2018-10-17 オリンパス株式会社 Laser microscope equipment
US9439568B2 (en) 2014-07-03 2016-09-13 Align Technology, Inc. Apparatus and method for measuring surface topography optically
US9261358B2 (en) 2014-07-03 2016-02-16 Align Technology, Inc. Chromatic confocal system
US11408024B2 (en) 2014-09-10 2022-08-09 Molecular Loop Biosciences, Inc. Methods for selectively suppressing non-target sequences
EP3191606B1 (en) 2014-09-12 2020-05-27 Illumina, Inc. Methods for detecting the presence of polymer subunits using chemiluminescence
JP2017536087A (en) 2014-09-24 2017-12-07 グッド スタート ジェネティクス, インコーポレイテッド Process control to increase the robustness of genetic assays
US10118173B2 (en) 2014-10-09 2018-11-06 Illumina, Inc. Method and device for separating immiscible liquids to effectively isolate at least one of the liquids
CN107408043A (en) 2014-10-14 2017-11-28 七桥基因公司 System and method for the intelligence tool in sequence streamline
US9897791B2 (en) 2014-10-16 2018-02-20 Illumina, Inc. Optical scanning systems for in situ genetic analysis
CA3002133A1 (en) 2014-10-17 2016-04-21 Good Start Genetics, Inc. Pre-implantation genetic screening and aneuploidy detection
US10000799B2 (en) 2014-11-04 2018-06-19 Boreal Genomics, Inc. Methods of sequencing with linked fragments
TWI569912B (en) * 2014-12-08 2017-02-11 國立高雄應用科技大學 Laser focusing optical module and laser focusing method
CA3010579A1 (en) 2015-01-06 2016-07-14 Good Start Genetics, Inc. Screening for structural variants
CN104568710B (en) * 2015-01-22 2017-03-08 福建师范大学 A kind of high time-space resolution optical detection and micro imaging method and device
JP6375239B2 (en) 2015-02-05 2018-08-15 オリンパス株式会社 Laser microscope equipment
US10192026B2 (en) 2015-03-05 2019-01-29 Seven Bridges Genomics Inc. Systems and methods for genomic pattern analysis
WO2016145366A1 (en) 2015-03-11 2016-09-15 Timothy Ragan System and methods for serial staining and imaging
US10576471B2 (en) 2015-03-20 2020-03-03 Illumina, Inc. Fluidics cartridge for use in the vertical or substantially vertical position
CA3077811C (en) 2015-03-24 2024-02-27 Illumina, Inc. Methods, carrier assemblies, and systems for imaging samples for biological or chemical analysis
US10275567B2 (en) 2015-05-22 2019-04-30 Seven Bridges Genomics Inc. Systems and methods for haplotyping
WO2016209731A1 (en) 2015-06-22 2016-12-29 Fluxergy, Llc Test card for assay and method of manufacturing same
US11371091B2 (en) 2015-06-22 2022-06-28 Fluxergy, Inc. Device for analyzing a fluid sample and use of test card with same
US10519493B2 (en) 2015-06-22 2019-12-31 Fluxergy, Llc Apparatus and method for image analysis of a fluid sample undergoing a polymerase chain reaction (PCR)
EP3310899B1 (en) * 2015-06-22 2022-05-25 FluxErgy, LLC Camera imaging system for a fluid sample assay and method of using same
WO2017007757A1 (en) 2015-07-06 2017-01-12 Illumina, Inc. Balanced ac modulation for driving droplet operations electrodes
EP3323070A4 (en) 2015-07-14 2019-03-13 Personal Genome Diagnostics Inc. Neoantigen analysis
ES2745556T3 (en) 2015-07-29 2020-03-02 Progenity Inc Nucleic acids and methods to detect chromosomal abnormalities
IL255445B (en) 2015-07-30 2022-07-01 Illumina Inc Orthogonal deblocking of nucleotides
US10793895B2 (en) 2015-08-24 2020-10-06 Seven Bridges Genomics Inc. Systems and methods for epigenetic analysis
US10584380B2 (en) 2015-09-01 2020-03-10 Seven Bridges Genomics Inc. Systems and methods for mitochondrial analysis
US10724110B2 (en) 2015-09-01 2020-07-28 Seven Bridges Genomics Inc. Systems and methods for analyzing viral nucleic acids
CN106541132B (en) * 2015-09-18 2018-09-25 广东汉邦激光科技有限公司 Laser 3D printing machine and its focusing system and method
DE102015116598A1 (en) 2015-09-30 2017-03-30 Carl Zeiss Microscopy Gmbh Method and microscope for high-resolution imaging by means of SIM
US10577643B2 (en) 2015-10-07 2020-03-03 Illumina, Inc. Off-target capture reduction in sequencing techniques
EP3693459A1 (en) 2015-10-10 2020-08-12 Guardant Health, Inc. Methods and applications of gene fusion detection in cell-free dna analysis
US11347704B2 (en) 2015-10-16 2022-05-31 Seven Bridges Genomics Inc. Biological graph or sequence serialization
CN108779487A (en) * 2015-11-16 2018-11-09 普罗格尼迪公司 Nucleic acid for detecting methylation state and method
WO2017095845A1 (en) 2015-12-01 2017-06-08 Illumina, Inc. Liquid storage and delivery mechanisms and methods
EP3384046B1 (en) 2015-12-01 2021-04-28 Illumina, Inc. Digital microfluidic system for single-cell isolation and characterization of analytes
EP3390668A4 (en) 2015-12-17 2020-04-01 Guardant Health, Inc. Methods to determine tumor gene copy number by analysis of cell-free dna
US20170199960A1 (en) 2016-01-07 2017-07-13 Seven Bridges Genomics Inc. Systems and methods for adaptive local alignment for graph genomes
US10364468B2 (en) 2016-01-13 2019-07-30 Seven Bridges Genomics Inc. Systems and methods for analyzing circulating tumor DNA
US10460829B2 (en) 2016-01-26 2019-10-29 Seven Bridges Genomics Inc. Systems and methods for encoding genetic variation for a population
US10262102B2 (en) 2016-02-24 2019-04-16 Seven Bridges Genomics Inc. Systems and methods for genotyping with graph reference
US10961573B2 (en) 2016-03-28 2021-03-30 Boreal Genomics, Inc. Linked duplex target capture
EP3377226B1 (en) 2016-03-28 2021-02-17 Illumina, Inc. Multi-plane microarrays
EP3436607B1 (en) 2016-03-28 2023-06-14 Ncan Genomics, Inc. Linked duplex target capture
ES2786974T3 (en) 2016-04-07 2020-10-14 Illumina Inc Methods and systems for the construction of standard nucleic acid libraries
CN108885260B (en) * 2016-04-08 2022-06-03 苹果公司 Time-of-flight detector with single axis scanning
EP3974815A1 (en) * 2016-05-27 2022-03-30 Verily Life Sciences LLC Systems and methods for 4-d hyperspectrial imaging
EP3465318B1 (en) * 2016-05-30 2022-03-23 The Trustees of Columbia University in the City of New York Three-dimensional imaging using swept, confocally aligned planar excitation
US11250931B2 (en) 2016-09-01 2022-02-15 Seven Bridges Genomics Inc. Systems and methods for detecting recombination
KR102416441B1 (en) 2016-09-22 2022-07-04 일루미나, 인코포레이티드 Detection of somatic copy number mutations
WO2018064116A1 (en) 2016-09-28 2018-04-05 Illumina, Inc. Methods and systems for data compression
US10955652B2 (en) 2016-09-30 2021-03-23 The Trustees Of Columbia University In The City Of New York Three-dimensional imaging using swept, confocally aligned planar excitation with a Powell lens and/or deliberate misalignment
DE102016119268B3 (en) * 2016-10-10 2017-12-21 Leica Microsystems Cms Gmbh Wrong plane microscope
TWI781669B (en) 2016-10-14 2022-10-21 美商伊路米納有限公司 Cartridge assembly
CN110023509A (en) 2016-11-15 2019-07-16 私人基因诊断公司 Non- unique bar code in genotyping measurement
KR101936120B1 (en) * 2016-11-30 2019-01-08 부경대학교 산학협력단 Probe for Photoacoustic Tomography and Real-time Photoacoustic Tomography Apparatus having the same
WO2018104908A2 (en) 2016-12-09 2018-06-14 Boreal Genomics, Inc. Linked ligation
GB201704771D0 (en) 2017-01-05 2017-05-10 Illumina Inc Modular optical analytic systems and methods
TWI773721B (en) * 2017-01-20 2022-08-11 日商東京威力科創股份有限公司 Foreign body detection device, foreign body detection method, and memory medium
WO2018140661A1 (en) * 2017-01-26 2018-08-02 Azure Biosystems, Inc. Devices and methods for imaging biomolecules
CN109414673B (en) 2017-02-01 2021-09-07 伊鲁米那股份有限公司 System and method having a reference responsive to multiple excitation frequencies
GB201701686D0 (en) 2017-02-01 2017-03-15 Illunina Inc System & method with fiducials having offset layouts
GB201701689D0 (en) 2017-02-01 2017-03-15 Illumia Inc System and method with fiducials of non-closed shapes
GB201701688D0 (en) 2017-02-01 2017-03-15 Illumia Inc System and method with fiducials in non-recliner layouts
GB201701691D0 (en) 2017-02-01 2017-03-15 Illumina Inc System and method with reflective fiducials
CN117116360A (en) 2017-03-30 2023-11-24 Illumina公司 Genome data analysis system and method
WO2018187013A1 (en) 2017-04-04 2018-10-11 Omniome, Inc. Fluidic apparatus and methods useful for chemical and biological reactions
US10161003B2 (en) 2017-04-25 2018-12-25 Omniome, Inc. Methods and apparatus that increase sequencing-by-binding efficiency
KR20200075814A (en) 2017-08-15 2020-06-26 옴니옴 인코포레이티드 Scanning devices and methods useful for the detection of chemical and biological analytes
CA3067421C (en) 2017-11-06 2023-08-15 Illumina, Inc. Nucleic acid indexing techniques
WO2019099306A1 (en) 2017-11-14 2019-05-23 Illumina, Inc. Droplet dispensing
KR102402002B1 (en) 2017-11-16 2022-05-25 일루미나, 인코포레이티드 Systems and Methods for Determining Microsatellite Instability
CN110870016A (en) 2017-11-30 2020-03-06 伊鲁米那股份有限公司 Verification method and system for sequence variant callouts
CA3097583A1 (en) 2018-04-19 2019-10-24 Omniome, Inc. Improving accuracy of base calls in nucleic acid sequencing methods
EP4234718A3 (en) 2018-04-26 2023-11-29 Pacific Biosciences Of California, Inc. Methods and compositions for stabilizing nucleic acid-nucleotide-polymerase complexes
WO2019231568A1 (en) 2018-05-31 2019-12-05 Omniome, Inc. Increased signal to noise in nucleic acid sequencing
US10753734B2 (en) 2018-06-08 2020-08-25 Dentsply Sirona Inc. Device, method and system for generating dynamic projection patterns in a confocal camera
AU2019312152A1 (en) 2018-07-24 2021-02-18 Pacific Biosciences Of California, Inc. Serial formation of ternary complex species
WO2020072030A1 (en) * 2018-10-01 2020-04-09 Hewlett-Packard Development Company, L.P. Microscopy systems
US10710076B2 (en) 2018-12-04 2020-07-14 Omniome, Inc. Mixed-phase fluids for nucleic acid sequencing and other analytical assays
AU2019400090A1 (en) 2018-12-14 2021-01-07 Illumina Cambridge Limited Decreasing phasing with unlabeled nucleotides during sequencing
CA3103744A1 (en) 2018-12-17 2020-06-25 Pietro GATTI-LAFRANCONI Compositions for use in polynucleotide sequencing
CA3103750A1 (en) 2018-12-17 2020-06-25 Illumina Cambridge Limited Primer oligonucleotide for sequencing
US11041199B2 (en) 2018-12-20 2021-06-22 Omniome, Inc. Temperature control for analysis of nucleic acids and other analytes
US11473136B2 (en) 2019-01-03 2022-10-18 Ncan Genomics, Inc. Linked target capture
WO2020146741A1 (en) 2019-01-10 2020-07-16 Selim Olcum Calibration of a functional biomarker instrument
EP3924513B1 (en) 2019-02-14 2023-04-12 Pacific Biosciences of California, Inc. Mitigating adverse impacts of detection systems on nucleic acids and other biological analytes
WO2020172444A1 (en) 2019-02-20 2020-08-27 Omniome, Inc. Scanning apparatus and methods for detecting chemical and biological analytes
NL2023312B1 (en) 2019-03-21 2020-09-28 Illumina Inc Artificial intelligence-based base calling
WO2020191390A2 (en) 2019-03-21 2020-09-24 Illumina, Inc. Artificial intelligence-based quality scoring
US11210554B2 (en) 2019-03-21 2021-12-28 Illumina, Inc. Artificial intelligence-based generation of sequencing metadata
NL2023310B1 (en) 2019-03-21 2020-09-28 Illumina Inc Training data generation for artificial intelligence-based sequencing
NL2023311B9 (en) 2019-03-21 2021-03-12 Illumina Inc Artificial intelligence-based generation of sequencing metadata
NL2023314B1 (en) 2019-03-21 2020-09-28 Illumina Inc Artificial intelligence-based quality scoring
NL2023316B1 (en) 2019-03-21 2020-09-28 Illumina Inc Artificial intelligence-based sequencing
US20220205917A1 (en) * 2019-05-02 2022-06-30 Duke University Devices and methods for imaging microarray chips
US11644406B2 (en) 2019-06-11 2023-05-09 Pacific Biosciences Of California, Inc. Calibrated focus sensing
EP3987274A1 (en) 2019-06-19 2022-04-27 Life Technologies Holdings Pte Limited Biological analysis devices and systems
DE102019004870B4 (en) * 2019-07-11 2023-03-09 Particle Metrix Gmbh Device and method for reducing the intensity reduction of the fluorescent dye by laser light when determining the fluorescence and the number of antibodies on exosomes.
US10656368B1 (en) 2019-07-24 2020-05-19 Omniome, Inc. Method and system for biological imaging using a wide field objective lens
CN114728996B (en) 2019-09-10 2022-11-29 加利福尼亚太平洋生物科学股份有限公司 Reversible modification of nucleotides
EP4045683A1 (en) 2019-10-18 2022-08-24 Omniome, Inc. Methods and compositions for capping nucleic acids
JP2023510438A (en) * 2019-12-31 2023-03-14 イルミナ インコーポレイテッド Autofocus function for optical sample analysis
CN115698282A (en) 2020-01-13 2023-02-03 福路伦特生物科学公司 Single cell sequencing
US20230054204A1 (en) 2020-02-04 2023-02-23 Pacific Biosciences Of California, Inc. Flow cells and methods for their manufacture and use
US20210265018A1 (en) 2020-02-20 2021-08-26 Illumina, Inc. Knowledge Distillation and Gradient Pruning-Based Compression of Artificial Intelligence-Based Base Caller
CN115516104A (en) 2020-03-03 2022-12-23 加利福尼亚太平洋生物科学股份有限公司 Methods and compositions for sequencing double-stranded nucleic acids
WO2021188500A1 (en) 2020-03-16 2021-09-23 Fluent Biosciences Inc. Multi-omic analysis in monodisperse droplets
WO2021224677A1 (en) 2020-05-05 2021-11-11 Akershus Universitetssykehus Hf Compositions and methods for characterizing bowel cancer
CN115836135A (en) 2020-05-05 2023-03-21 加利福尼亚太平洋生物科学股份有限公司 Compositions and methods for modifying polymerase-nucleic acid complexes
US11188778B1 (en) 2020-05-05 2021-11-30 Illumina, Inc. Equalization-based image processing and spatial crosstalk attenuator
AU2021269069A1 (en) 2020-05-08 2022-10-20 Illumina, Inc. Genome sequencing and detection techniques
EP4165549A1 (en) 2020-06-11 2023-04-19 Nautilus Biotechnology, Inc. Methods and systems for computational decoding of biological, chemical, and physical entities
WO2022029484A1 (en) 2020-08-06 2022-02-10 Agendia NV Methods of assessing breast cancer using circulating hormone receptor transcripts
US20220042106A1 (en) 2020-08-06 2022-02-10 Agendia NV Systems and methods of using cell-free nucleic acids to tailor cancer treatment
US20220042107A1 (en) 2020-08-06 2022-02-10 Agendia NV Systems and methods of scoring risk and residual disease from passenger mutations
WO2022029488A1 (en) 2020-08-06 2022-02-10 Agenda Nv Systems and methods of assessing breast cancer
EP4211508A1 (en) 2020-09-14 2023-07-19 Singular Genomics Systems, Inc. Methods and systems for multidimensional imaging
WO2022103887A1 (en) 2020-11-11 2022-05-19 Nautilus Biotechnology, Inc. Affinity reagents having enhanced binding and detection characteristics
JP2023552015A (en) 2020-12-02 2023-12-14 イルミナ ソフトウェア, インコーポレイテッド Systems and methods for detecting genetic mutations
WO2022120595A1 (en) * 2020-12-08 2022-06-16 深圳华大智造科技股份有限公司 Super-resolution measurement system and super-resolution measurement method
EP4153964A4 (en) * 2020-12-21 2023-11-29 Singular Genomics Systems, Inc. Systems and methods for multicolor imaging
WO2022159663A1 (en) 2021-01-21 2022-07-28 Nautilus Biotechnology, Inc. Systems and methods for biomolecule preparation
AU2022232933A1 (en) 2021-03-11 2023-09-07 Nautilus Subsidiary, Inc. Systems and methods for biomolecule retention
CA3214206A1 (en) 2021-03-31 2022-10-06 Carla SANMARTIN Nucleic acid library sequencing techniques with adapter dimer detection
WO2022232425A2 (en) 2021-04-29 2022-11-03 Illumina, Inc. Amplification techniques for nucleic acid characterization
WO2022240766A1 (en) 2021-05-10 2022-11-17 Pacific Biosciences Of California, Inc. Dna amplification buffer replenishment during rolling circle amplification
EP4337786A1 (en) 2021-05-10 2024-03-20 Pacific Biosciences of California, Inc. Single-molecule seeding and amplification on a surface
US20220403450A1 (en) 2021-06-03 2022-12-22 Illumina Software, Inc. Systems and methods for sequencing nucleotides using two optical channels
US20220414853A1 (en) 2021-06-25 2022-12-29 Illumina, Inc. Fiducials for use in registration of a patterned surface
KR20240025515A (en) 2021-06-25 2024-02-27 일루미나, 인코포레이티드 Linear Fourier origin
US11455487B1 (en) 2021-10-26 2022-09-27 Illumina Software, Inc. Intensity extraction and crosstalk attenuation using interpolation and adaptation for base calling
WO2023003757A1 (en) 2021-07-19 2023-01-26 Illumina Software, Inc. Intensity extraction with interpolation and adaptation for base calling
CN113780521B (en) * 2021-08-24 2022-03-04 中国人民解放军93114部队 Radiation source individual identification method based on deep learning
WO2023034079A1 (en) 2021-09-01 2023-03-09 Illumina Software, Inc. Amplitude modulation for accelerated base calling
US20230070896A1 (en) 2021-09-09 2023-03-09 Nautilus Biotechnology, Inc. Characterization and localization of protein modifications
US20230088338A1 (en) 2021-09-10 2023-03-23 Illumina, Inc. Sequencer focus quality metrics and focus tracking for periodically patterned surfaces
WO2023049215A1 (en) 2021-09-22 2023-03-30 Illumina, Inc. Compressed state-based base calling
AU2022352593A1 (en) 2021-09-22 2024-02-15 Nautilus Subsidiary, Inc. Methods and systems for determining polypeptide interactions
US20230096386A1 (en) 2021-09-30 2023-03-30 Illumina Cambridge Limited Polynucleotide sequencing
WO2023064181A1 (en) 2021-10-11 2023-04-20 Nautilus Biotechnology, Inc. Highly multiplexable analysis of proteins and proteomes
CN114035199A (en) * 2021-10-15 2022-02-11 中国人民解放军91977部队 Photoelectric search tracking device based on avalanche diode imaging device
WO2023081728A1 (en) 2021-11-03 2023-05-11 Nautilus Biotechnology, Inc. Systems and methods for surface structuring
WO2023081485A1 (en) 2021-11-08 2023-05-11 Pacific Biosciences Of California, Inc. Stepwise sequencing of a polynucleotide with a homogenous reaction mixture
US20230183799A1 (en) 2021-12-10 2023-06-15 Illumina, Inc. Parallel sample and index sequencing
US20230215515A1 (en) 2021-12-23 2023-07-06 Illumina Software, Inc. Facilitating secure execution of external workflows for genomic sequencing diagnostics
WO2023122363A1 (en) 2021-12-23 2023-06-29 Illumina Software, Inc. Dynamic graphical status summaries for nucelotide sequencing
WO2023129764A1 (en) 2021-12-29 2023-07-06 Illumina Software, Inc. Automatically switching variant analysis model versions for genomic analysis applications
US20230296516A1 (en) 2022-02-17 2023-09-21 Illumina, Inc. Ai-driven signal enhancement of sequencing images
WO2023158809A1 (en) 2022-02-17 2023-08-24 Illumina, Inc. Ai-driven enhancement of motion blurred sequencing images
US11639499B1 (en) 2022-03-08 2023-05-02 Watchmaker Genomics, Inc. Reverse transcriptase variants
WO2023175021A1 (en) 2022-03-15 2023-09-21 Illumina, Inc. Methods of preparing loop fork libraries
WO2023183937A1 (en) 2022-03-25 2023-09-28 Illumina, Inc. Sequence-to-sequence base calling
US20230314324A1 (en) 2022-03-29 2023-10-05 Nautilus Subsidiary, Inc. Integrated arrays for single-analyte processes
CA3224264A1 (en) 2022-04-08 2023-10-12 Illumina, Inc. Aptamer dynamic range compression and detection techniques
WO2023212490A1 (en) 2022-04-25 2023-11-02 Nautilus Subsidiary, Inc. Systems and methods for assessing and improving the quality of multiplex molecular assays
US20230407386A1 (en) 2022-06-09 2023-12-21 Illumina, Inc. Dependence of base calling on flow cell tilt
WO2023250364A1 (en) 2022-06-21 2023-12-28 Nautilus Subsidiary, Inc. Method for detecting analytes at sites of optically non-resolvable distances
US20240033738A1 (en) 2022-07-27 2024-02-01 Illumina, Inc. Flow cell based motion system calibration and control methods

Family Cites Families (118)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US4204230A (en) 1978-10-25 1980-05-20 Xerox Corporation High resolution input scanner using a two dimensional detector array
US4382267A (en) 1981-09-24 1983-05-03 Rca Corporation Digital control of number of effective rows of two-dimensional charge-transfer imager array
US4700298A (en) 1984-09-14 1987-10-13 Branko Palcic Dynamic microscope image processing scanner
US4826299A (en) 1987-01-30 1989-05-02 Canadian Patents And Development Limited Linear deiverging lens
US4845552A (en) 1987-08-20 1989-07-04 Bruno Jaggi Quantitative light microscope using a solid state detector in the primary image plane
US4877326A (en) 1988-02-19 1989-10-31 Kla Instruments Corporation Method and apparatus for optical inspection of substrates
GB8822228D0 (en) 1988-09-21 1988-10-26 Southern E M Support-bound oligonucleotides
US5143854A (en) 1989-06-07 1992-09-01 Affymax Technologies N.V. Large scale photolithographic solid phase synthesis of polypeptides and receptor binding screening thereof
US5744101A (en) 1989-06-07 1998-04-28 Affymax Technologies N.V. Photolabile nucleoside protecting groups
DE3924454A1 (en) 1989-07-24 1991-02-07 Cornelis P Prof Dr Hollenberg THE APPLICATION OF DNA AND DNA TECHNOLOGY FOR THE CONSTRUCTION OF NETWORKS FOR USE IN CHIP CONSTRUCTION AND CHIP PRODUCTION (DNA CHIPS)
US5252743A (en) 1989-11-13 1993-10-12 Affymax Technologies N.V. Spatially-addressable immobilization of anti-ligands on surfaces
US5159199A (en) 1991-08-12 1992-10-27 The United States Of America As Represented By The Administrator Of The National Aeronautics And Space Administration Integrated filter and detector array for spectral imaging
ATE148889T1 (en) 1991-09-18 1997-02-15 Affymax Tech Nv METHOD FOR SYNTHESIS OF VARIOUS COLLECTIONS OF OLIGOMERS
WO1993009668A1 (en) 1991-11-22 1993-05-27 Affymax Technology N.V. Combinatorial strategies for polymer synthesis
US5324633A (en) 1991-11-22 1994-06-28 Affymax Technologies N.V. Method and apparatus for measuring binding affinity
US5173748A (en) 1991-12-05 1992-12-22 Eastman Kodak Company Scanning multichannel spectrometry using a charge-coupled device (CCD) in time-delay integration (TDI) mode
AU3728093A (en) 1992-02-19 1993-09-13 Public Health Research Institute Of The City Of New York, Inc., The Novel oligonucleotide arrays and their use for sorting, isolating, sequencing, and manipulating nucleic acids
US5583211A (en) 1992-10-29 1996-12-10 Beckman Instruments, Inc. Surface activated organic polymers useful for location - specific attachment of nucleic acids, peptides, proteins and oligosaccharides
US5491074A (en) 1993-04-01 1996-02-13 Affymax Technologies Nv Association peptides
US5858659A (en) 1995-11-29 1999-01-12 Affymetrix, Inc. Polymorphism detection
US5472672A (en) 1993-10-22 1995-12-05 The Board Of Trustees Of The Leland Stanford Junior University Apparatus and method for polymer synthesis using arrays
US5429807A (en) 1993-10-28 1995-07-04 Beckman Instruments, Inc. Method and apparatus for creating biopolymer arrays on a solid support surface
US6309601B1 (en) 1993-11-01 2001-10-30 Nanogen, Inc. Scanning optical detection system
US6090555A (en) 1997-12-11 2000-07-18 Affymetrix, Inc. Scanned image alignment systems and methods
US5578832A (en) 1994-09-02 1996-11-26 Affymetrix, Inc. Method and apparatus for imaging a sample on a device
US6741344B1 (en) 1994-02-10 2004-05-25 Affymetrix, Inc. Method and apparatus for detection of fluorescently labeled materials
DE69503126T2 (en) 1994-05-05 1998-11-12 Beckman Instruments Inc REPETITIVE OLIGONUCLEOTIDE MATRIX
US5782770A (en) 1994-05-12 1998-07-21 Science Applications International Corporation Hyperspectral imaging methods and apparatus for non-invasive diagnosis of tissue for cancer
US5571639A (en) 1994-05-24 1996-11-05 Affymax Technologies N.V. Computer-aided engineering system for design of sequence arrays and lithographic masks
US5807522A (en) 1994-06-17 1998-09-15 The Board Of Trustees Of The Leland Stanford Junior University Methods for fabricating microarrays of biological samples
US5795716A (en) 1994-10-21 1998-08-18 Chee; Mark S. Computer-aided visualization and analysis system for sequence evaluation
US5556752A (en) 1994-10-24 1996-09-17 Affymetrix, Inc. Surface-bound, unimolecular, double-stranded DNA
WO1996018205A1 (en) 1994-12-08 1996-06-13 Molecular Dynamics, Inc. Fluorescence imaging system employing a macro scanning objective
US5599695A (en) 1995-02-27 1997-02-04 Affymetrix, Inc. Printing molecular library arrays using deprotection agents solely in the vapor phase
US5624711A (en) 1995-04-27 1997-04-29 Affymax Technologies, N.V. Derivatization of solid supports and methods for oligomer synthesis
US5578818A (en) 1995-05-10 1996-11-26 Molecular Dynamics LED point scanning system
US5545531A (en) 1995-06-07 1996-08-13 Affymax Technologies N.V. Methods for making a device for concurrently processing multiple biological chip assays
US5528050A (en) 1995-07-24 1996-06-18 Molecular Dynamics, Inc. Compact scan head with multiple scanning modalities
US5968740A (en) 1995-07-24 1999-10-19 Affymetrix, Inc. Method of Identifying a Base in a Nucleic Acid
US5585639A (en) 1995-07-27 1996-12-17 Hewlett-Packard Company Optical scanning apparatus
US5658734A (en) 1995-10-17 1997-08-19 International Business Machines Corporation Process for synthesizing chemical compounds
US6022963A (en) 1995-12-15 2000-02-08 Affymetrix, Inc. Synthesis of oligonucleotide arrays using photocleavable protecting groups
US5629808A (en) 1995-12-15 1997-05-13 National Research Council Of Canada D-shape laser beam projector
GB9526183D0 (en) * 1995-12-21 1996-02-21 Stc Submarine Systems Ltd Dispersion slope equalisaion for wdm systems wih branches
US5847400A (en) 1996-02-01 1998-12-08 Molecular Dynamics, Inc. Fluorescence imaging system having reduced background fluorescence
US7244622B2 (en) * 1996-04-03 2007-07-17 Applera Corporation Device and method for multiple analyte detection
WO1997043611A1 (en) 1996-05-16 1997-11-20 Affymetrix, Inc. Systems and methods for detection of labeled materials
US5754291A (en) 1996-09-19 1998-05-19 Molecular Dynamics, Inc. Micro-imaging system
ATE234674T1 (en) 1996-11-14 2003-04-15 Affymetrix Inc CHEMICAL AMPLIFICATION FOR SYNTHESIS OF PATTERN ORDERS
US6297006B1 (en) 1997-01-16 2001-10-02 Hyseq, Inc. Methods for sequencing repetitive sequences and for determining the order of sequence subfragments
JP3438855B2 (en) 1997-01-23 2003-08-18 横河電機株式会社 Confocal device
US5837475A (en) 1997-01-30 1998-11-17 Hewlett-Packard Co. Apparatus and method for scanning a chemical array
DE19707227A1 (en) 1997-02-24 1998-08-27 Bodenseewerk Perkin Elmer Co Light scanner
US6327410B1 (en) 1997-03-14 2001-12-04 The Trustees Of Tufts College Target analyte sensors utilizing Microspheres
US6023540A (en) 1997-03-14 2000-02-08 Trustees Of Tufts College Fiber optic sensor with encoded microspheres
US6008892A (en) 1997-05-23 1999-12-28 Molecular Dynamics, Inc. Optical substrate for enhanced detectability of fluorescence
CA2294053A1 (en) 1997-06-25 1998-12-30 Orchid Biocomputer, Inc. Methods for the detection of multiple single nucleotide polymorphisms in a single reaction
US6043506A (en) 1997-08-13 2000-03-28 Bio-Rad Laboratories, Inc. Multi parameter scanner
US6043880A (en) 1997-09-15 2000-03-28 Becton Dickinson And Company Automated optical reader for nucleic acid assays
US6033860A (en) 1997-10-31 2000-03-07 Affymetrix, Inc. Expression profiles in adult and fetal organs
US5998796A (en) 1997-12-22 1999-12-07 Spectrumedix Corporation Detector having a transmission grating beam splitter for multi-wavelength sample analysis
US6087102A (en) 1998-01-07 2000-07-11 Clontech Laboratories, Inc. Polymeric arrays and methods for their use in binding assays
US6428752B1 (en) 1998-05-14 2002-08-06 Affymetrix, Inc. Cleaning deposit devices that form microarrays and the like
US6287776B1 (en) 1998-02-02 2001-09-11 Signature Bioscience, Inc. Method for detecting and classifying nucleic acid hybridization
US6309831B1 (en) 1998-02-06 2001-10-30 Affymetrix, Inc. Method of manufacturing biological chips
JP3944996B2 (en) 1998-03-05 2007-07-18 株式会社日立製作所 DNA probe array
US6388788B1 (en) 1998-03-16 2002-05-14 Praelux, Inc. Method and apparatus for screening chemical compounds
KR100618502B1 (en) * 1998-03-16 2006-09-01 지이 헬스케어 바이오-사이언시즈 코프. System and method of focusing electromagnetic radiation for use in a confocal microscopy imaging system
WO1999047041A1 (en) 1998-03-19 1999-09-23 Board Of Regents, The University Of Texas System Fiber-optic confocal imaging apparatus and methods of use
US6031078A (en) 1998-06-16 2000-02-29 Millennium Pharmaceuticals, Inc. MTbx protein and nucleic acid molecules and uses therefor
US6160618A (en) 1998-06-19 2000-12-12 Board Of Regents, The University Of Texas System Hyperspectral slide reader
US6678048B1 (en) 1998-07-20 2004-01-13 Sandia Corporation Information-efficient spectral imaging sensor with TDI
US6245507B1 (en) 1998-08-18 2001-06-12 Orchid Biosciences, Inc. In-line complete hyperspectral fluorescent imaging of nucleic acid molecules
KR20010090718A (en) * 1998-08-21 2001-10-19 써로메드, 인크. Novel optical architectures for microvolume laser-scanning cytometers
US6277628B1 (en) 1998-10-02 2001-08-21 Incyte Genomics, Inc. Linear microarrays
AT410718B (en) 1998-10-28 2003-07-25 Schindler Hansgeorg Dr DEVICE FOR VISUALIZING MOLECULES
US6545264B1 (en) 1998-10-30 2003-04-08 Affymetrix, Inc. Systems and methods for high performance scanning
US6134002A (en) 1999-01-14 2000-10-17 Duke University Apparatus and method for the rapid spectral resolution of confocal images
US6355934B1 (en) 1999-02-26 2002-03-12 Packard Biochip Technologies Imaging system for an optical scanner
US6355431B1 (en) 1999-04-20 2002-03-12 Illumina, Inc. Detection of nucleic acid amplification reactions using bead arrays
DK1923471T3 (en) 1999-04-20 2013-04-02 Illumina Inc Detection of nucleic acid reactions on bead arrays
US20030207295A1 (en) 1999-04-20 2003-11-06 Kevin Gunderson Detection of nucleic acid reactions on bead arrays
US20030108867A1 (en) 1999-04-20 2003-06-12 Chee Mark S Nucleic acid sequencing using microsphere arrays
US6371370B2 (en) 1999-05-24 2002-04-16 Agilent Technologies, Inc. Apparatus and method for scanning a surface
US6222664B1 (en) 1999-07-22 2001-04-24 Agilent Technologies Inc. Background reduction apparatus and method for confocal fluorescence detection systems
WO2001037025A1 (en) 1999-11-16 2001-05-25 Agilent Technologies, Inc. Confocal imaging
US7582420B2 (en) 2001-07-12 2009-09-01 Illumina, Inc. Multiplex nucleic acid reactions
US7611869B2 (en) 2000-02-07 2009-11-03 Illumina, Inc. Multiplexed methylation detection methods
US6770441B2 (en) 2000-02-10 2004-08-03 Illumina, Inc. Array compositions and methods of making same
US6687000B1 (en) 2000-06-26 2004-02-03 Wisconsin Alumni Research Foundation Photon-sorting spectroscopic microscope system
US6818907B2 (en) * 2000-10-17 2004-11-16 The President And Fellows Of Harvard College Surface plasmon enhanced illumination system
JP3590765B2 (en) * 2000-12-21 2004-11-17 Smc株式会社 solenoid valve
TW555954B (en) * 2001-02-28 2003-10-01 Olympus Optical Co Confocal microscope, optical height-measurement method, automatic focusing method
GB0106342D0 (en) 2001-03-15 2001-05-02 Renishaw Plc Spectroscopy apparatus and method
US6650411B2 (en) 2001-04-26 2003-11-18 Affymetrix, Inc. System, method, and product for pixel clocking in scanning of biological materials
US7248716B2 (en) * 2001-07-06 2007-07-24 Palantyr Research, Llc Imaging system, methodology, and applications employing reciprocal space optical design
US7105795B2 (en) * 2001-07-06 2006-09-12 Palantyr Research, Llc Imaging system, methodology, and applications employing reciprocal space optical design
JP2003028798A (en) * 2001-07-11 2003-01-29 Olympus Optical Co Ltd Fluorescence acquisition device
US20080099667A1 (en) * 2001-08-14 2008-05-01 President And Fellows Of Harvard College Methods and apparatus for sensing a physical substance
JP3616999B2 (en) 2001-12-04 2005-02-02 レーザーテック株式会社 Confocal microscope
CN1435684A (en) * 2002-01-30 2003-08-13 无锡市朗珈生物医学工程有限公司 Bio-chip fluorescent testing scanning device
US6825930B2 (en) 2002-06-04 2004-11-30 Cambridge Research And Instrumentation, Inc. Multispectral imaging system
US7399643B2 (en) 2002-09-12 2008-07-15 Cyvera Corporation Method and apparatus for aligning microbeads in order to interrogate the same
US20050227252A1 (en) 2002-08-20 2005-10-13 Moon John A Diffraction grating-based encoded articles for multiplexed experiments
US6813018B2 (en) 2002-11-07 2004-11-02 The Boeing Company Hyperspectral imager
US20040140417A1 (en) * 2002-12-27 2004-07-22 Accretech (Israel) Ltd. Image sensor for confocal microscopy
US20050181394A1 (en) 2003-06-20 2005-08-18 Illumina, Inc. Methods and compositions for whole genome amplification and genotyping
JP4235046B2 (en) * 2003-07-08 2009-03-04 新日本製鐵株式会社 Steel plate surface inspection method, system, image processing apparatus, and computer program
US7218446B2 (en) 2003-08-27 2007-05-15 Biomedical Photometrics Inc. Imaging system having a fine focus
CA2559801A1 (en) 2003-10-01 2005-04-14 Cyvera Corporation Optical reader for diffraction grating-based encoded optical identification elements
US7532323B2 (en) * 2003-10-28 2009-05-12 Cha-Min Tang Spatial light modulator apparatus and method
US7227113B2 (en) * 2003-11-21 2007-06-05 Olympus Corporation Confocal laser scanning microscope
JP4589101B2 (en) * 2003-12-25 2010-12-01 昭和電工株式会社 Surface inspection method and apparatus
JP4635145B2 (en) 2004-03-17 2011-02-16 レーザーテック株式会社 Confocal microscope and film thickness measuring device
US20050225849A1 (en) * 2004-04-05 2005-10-13 Fujifilm Electronic Imaging Ltd. Slit confocal microscope and method
JP5244386B2 (en) 2004-06-30 2013-07-24 アノト アクティエボラーク Data management with electronic pen
US7329860B2 (en) * 2005-11-23 2008-02-12 Illumina, Inc. Confocal imaging methods and apparatus
US7791013B2 (en) * 2006-11-21 2010-09-07 Illumina, Inc. Biological microarray line scanning method and system

Cited By (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US10823612B2 (en) 2015-01-30 2020-11-03 Japan Science And Technology Agency Multifocal spectrometric measurement device, and optical system for multifocal spectrometric measurement device
CN108519329A (en) * 2018-03-26 2018-09-11 华中科技大学 A kind of line co-focusing imaging device of multi-channel scanning and detection
CN108519329B (en) * 2018-03-26 2021-01-15 华中科技大学 Multi-channel scanning and detecting line confocal imaging device

Also Published As

Publication number Publication date
CA2632221C (en) 2013-08-27
EP2594981B1 (en) 2017-05-10
CN101361015A (en) 2009-02-04
US20120168644A1 (en) 2012-07-05
US20080290263A1 (en) 2008-11-27
EP2594981A3 (en) 2013-06-19
CN101361015B (en) 2012-11-21
US9816929B2 (en) 2017-11-14
EP1955102B1 (en) 2013-04-24
ES2635094T3 (en) 2017-10-02
WO2007062039A3 (en) 2007-07-19
US20150069267A1 (en) 2015-03-12
US20070114362A1 (en) 2007-05-24
ES2407968T3 (en) 2013-06-17
US7589315B2 (en) 2009-09-15
WO2007062039A2 (en) 2007-05-31
PL2594981T3 (en) 2017-10-31
EP1955102A2 (en) 2008-08-13
US7960685B2 (en) 2011-06-14
JP5055292B2 (en) 2012-10-24
JP2009517662A (en) 2009-04-30
US20110204212A1 (en) 2011-08-25
US20100012825A1 (en) 2010-01-21
DK1955102T3 (en) 2013-05-13
DK2594981T3 (en) 2017-08-21
US8884211B2 (en) 2014-11-11
EP2594981A2 (en) 2013-05-22
US8158926B2 (en) 2012-04-17
US7329860B2 (en) 2008-02-12

Similar Documents

Publication Publication Date Title
EP1955102B1 (en) Confocal imaging methods and apparatus
US7813013B2 (en) Hexagonal site line scanning method and system
US7791013B2 (en) Biological microarray line scanning method and system
US7567346B2 (en) System and method for multimode imaging
US7830575B2 (en) Optical scanner with improved scan time
US20100277580A1 (en) Multi-modal spot generator and multi-modal multi-spot scanning microscope
WO2015100421A1 (en) Multi-foci multiphoton imaging systems and methods
JP2004170977A (en) Method and arrangement for optically grasping sample with depth of resolution
JP2012237647A (en) Multifocal confocal raman spectroscopic microscope
EP2533033A1 (en) Device for analyzing luminescent bio-microchips
EP2828700A1 (en) Multi-color confocal microscope and imaging methods
EP2215503A2 (en) Multi-focal spot generator and multi-focal multi-spot scanning microscope
US20230221178A1 (en) Apparatus and a method for fluorescence imaging

Legal Events

Date Code Title Description
EEER Examination request