WO1994008352A1 - Illuminator for fluorescence-analyzer such as a microtitration fluorimeter used in cell biology - Google Patents

Illuminator for fluorescence-analyzer such as a microtitration fluorimeter used in cell biology Download PDF

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Publication number
WO1994008352A1
WO1994008352A1 PCT/US1993/009249 US9309249W WO9408352A1 WO 1994008352 A1 WO1994008352 A1 WO 1994008352A1 US 9309249 W US9309249 W US 9309249W WO 9408352 A1 WO9408352 A1 WO 9408352A1
Authority
WO
WIPO (PCT)
Prior art keywords
lamp
illuminator
analyzer
microtitration
product
Prior art date
Application number
PCT/US1993/009249
Other languages
French (fr)
Inventor
Patrice Rat
Original Assignee
Dynatech Laboratories, Inc.
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
Application filed by Dynatech Laboratories, Inc. filed Critical Dynatech Laboratories, Inc.
Priority to JP6509273A priority Critical patent/JPH08505697A/en
Publication of WO1994008352A1 publication Critical patent/WO1994008352A1/en

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Classifications

    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/645—Specially adapted constructive features of fluorimeters
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/645—Specially adapted constructive features of fluorimeters
    • G01N2021/6463—Optics
    • G01N2021/6473—In-line geometry
    • G01N2021/6476—Front end, i.e. backscatter, geometry

Definitions

  • the present invention concerns an illuminator for a fluorescence analyzer, illustratively a microtitration fluorimeter used in particular in biology.
  • Biochemical fluorimeter ⁇ such as described in US patent 4,501,970 are presently commercially available.
  • This apparatus serves to measure the fluorescence of samples placed in microtitration wells; it comprises an optical system serving on one hand to point an energizing light beam downward into the opening of a well to excite the sample therein by fluorescence and on the other hand to detect the light beam emitted by the sample and passing through the opening cf this well toward an analyzer.
  • the lamp is arranged in such manner that its axis is perpendicular to the emitted light beam entering the analyzer. The light spectrum of this beam ranges from ultraviolet to visible.
  • the sensitivity cf such apparatus is 10 "f to 10 "s g/ml of detected sample; in biochemistry this is adequate because the sample in the well is in solution.
  • the sample in a well is present as a cell culture, that is in a heterogeneous medium. Also the cells settling, the light beam must be able to continue as far as the bottom of the well, and this entrails substantial energy.
  • the sensitivity must be about 10 "10 to 10 "12 g/ l of detected fluorochrome: this is impossible using the apparatus of the prior art known to the expert .
  • one object of the present invention is to create an illuminator for a fluorescence analyzer allowing to achieve a minimum sensitivity of 10 "10 to 10 "11 g/ml at least across the entire visible and ultraviolet spectrum and able to operate across this entire spectrum. Accordingly the light beam must evince a spectrum covering 290 - 300 nm and 700 - 750 nm.
  • the power emitted by the lamp must be higher than that from the lamps presently in use because the cells are at the bottom of the well .
  • the only high-power lamps known are those using xenon, which however achieve sensitivities of the order of 10 "5 g/ml of detected sample. Even if the sensitivity could be improved, such lamps are impractical, not only because of the long time required to reach their operational temperatures, but also because they dissipate much heat, thereby requiring large ventilating means as otherwise the optics would rapidly degrade or even be destroyed.
  • these known lamps for fluorescence analysis on account of the blowers and devices supplying sufficient electric power generate background noise lowering the sensitivity of the analytical apparatus .
  • an illuminator for a fluorescence analyzer comprising an optics, at least one support for the products to be analyzed and an analytical device
  • said, illuminator is characterized in the present invention by comprising on one hand a lamp of which the emission axis is collinear with the axis of the light flux entering the optics, said lamp emitting a flux of at least 2,000 lumens and evincing a mean luminous efficiency of at least 30 lumens/watt and a flux density of at least 1,200 x 10 6 lumens/m 3 , and on the other hand a heat dissipator.
  • the lamp shall be argon-iodide, xenon-flash or rare-gas/halogen, preferably xenon, for instance a halogen-krypton-xenon lamp.
  • the heat dissipator shall be an infrared trap illustratively consisting of cups arranged in a semi-cylindrical envelope enclosing the rear part of the lamp in relation to the direction of the emitted flux.
  • the illuminator also includes an interference excitation filter located in front of the lamp as seen in the direction of the emitted light beam.
  • the support for the product being analyzed consists of at least one microtitration well or of two (microscope) slides between which said product is placed.
  • the fluorescence analyzer is a microtitration fluorimeter especially as regards cell biology.
  • Fig. 1 schematically shows a fluorimeter of the prior art
  • FIG. 2 schematically shows the same fluorimeter now fitted with an illuminator of the invention.
  • the illuminator is described in relation to the fluorimeter which is the object of the US patent 4,501,970.
  • This fluorimeter comprises a lamp 1 mounted just in front of a diaphragm 2 constituting the entry into a tunnel receiving an optics such as previously described.
  • the light beam 3 entering the tunnel then is directed toward a microtitration well 4 holding the solution which must be analyzed.
  • the reflected beam 5 is directed toward an analyzer 6.
  • Lamp 1 is a mercury-vapor lamp emitting in the ultraviolet and with an axis perpendicular to that of the light beam 3.
  • the lamp's light flux crossing the diaphragm 2 is about 1,300 lumens, its mean luminous efficiency being about 26 lumen/watt.
  • the lamp 11 is a conventional halogen-krypton-xenon lamp called krypton- xenon by the specialists; it might also be of the argon- iodide, xenon-flash or halogen/rare-gase ⁇ type, one of these preferably being xenon.
  • the axis of this lamp 11 is collinear with the axis of the diaphragm 12, that is, it is collinear with the direction of the mitted flux.
  • the light flux 13 emitted by such a lamp 11 is at least 2,000 lumen, for instance about 2,300 lumen.
  • This lamp 11 is characterized by a mean luminous efficiency of at least 30 lumen/watt and by a luminous flux density of at least 1,200 x 10 6 /ro 3 - Accordingly this is a small bulb requiring no electric-power regulation nor any heat-up time.
  • These lamps emit a light beam 13 of which the spectrum ranges from ultraviolet to visible, whereas the prior art lamp emits in the ultraviolet or in the visible.
  • this lamp comprises an infrared trap 14 allowing heat dissipation and thereby prevents overheating the components of the optics. Accordingly not only the maximum light flux from the lamp 11 enters the tunnel through the diaphragm 12, but furthermore only the photon power. This is highly important because the heat power emitted by this lamp unless trapped would degrade and possibly destroy at least some electronic or optical .components.
  • the infrared trap 14 consists of cups arranged within a semi-cylindrical envelope enclosing the rear part of the lamp relative to the diaphragm 12.
  • such a lamp 11 fitted with its infrared trap 14 must be located in front of the diaphragm 12 at such a distance that the light beam crossing this diaphragm may be considered cold.
  • the illuminator may include an interference excitation filter 15 in front of the diaphragm 12 : only those photons useful in the measurement shall enter the optics.
  • an interference excitation filter 15 in front of the diaphragm 12 : only those photons useful in the measurement shall enter the optics.

Abstract

The invention concerns an illuminator for a fluorescence analyzer comprising an optics, at least one support for the product to be analyzed and an analyzing device. The illuminator furthermore includes a lamp (11) of which the emission axis is collinear with the axis of the emitted light flux entering the optics and amounting to at least 2,000 lumens and evincing a mean luminous efficiency of at least 30 lumen/watt and a mean light flux density of at least 1,200 x 106 lumen/m3, and furthermore a heat dissipator (14).

Description

ILLUMINATOR FOR FLUORESCENCΞ-ANALYZER SUCH AS A MICROTITRATION FLUORIMETER USED IN CELL BIOLOGY.
Background Art
The present invention concerns an illuminator for a fluorescence analyzer, illustratively a microtitration fluorimeter used in particular in biology. Biochemical fluorimeterε such as described in US patent 4,501,970 are presently commercially available. This apparatus serves to measure the fluorescence of samples placed in microtitration wells; it comprises an optical system serving on one hand to point an energizing light beam downward into the opening of a well to excite the sample therein by fluorescence and on the other hand to detect the light beam emitted by the sample and passing through the opening cf this well toward an analyzer. In such a fluorimeter, the lamp is arranged in such manner that its axis is perpendicular to the emitted light beam entering the analyzer. The light spectrum of this beam ranges from ultraviolet to visible.
Presently one apparatus made in accordance with this US patent is known: it comprises a mercury vapor lamp with which only ultraviolet measurements can be taken. Another type of fluorimeter comprises a tungsten lamp emitting mostly in the visible: no reliable measurement is possible in ultraviolet.
Moreover the sensitivity cf such apparatus is 10"f to 10"s g/ml of detected sample; in biochemistry this is adequate because the sample in the well is in solution. In microtitration cell-biology the sample in a well is present as a cell culture, that is in a heterogeneous medium. Also the cells settling, the light beam must be able to continue as far as the bottom of the well, and this entrails substantial energy.
Furthermore the sensitivity must be about 10"10 to 10"12 g/ l of detected fluorochrome: this is impossible using the apparatus of the prior art known to the expert . The presently used lamps in biochemical apparatus -- regardless of being tungsten or mercury or xenon vapor -- preclude such sensitivities.
Accordingly one object of the present invention is to create an illuminator for a fluorescence analyzer allowing to achieve a minimum sensitivity of 10"10 to 10"11 g/ml at least across the entire visible and ultraviolet spectrum and able to operate across this entire spectrum. Accordingly the light beam must evince a spectrum covering 290 - 300 nm and 700 - 750 nm.
To solve such a problem, one might use two lamps in parallel, one for the ultraviolet spectrum and another for the visible spectrum. However such an arrangement is impractical because requiring the operator not only to change lamps but furthermore to alter, with each lamp change, the optical configuration of the apparatus in relation to the excitation spectrum.
As already mentioned above, the power emitted by the lamp must be higher than that from the lamps presently in use because the cells are at the bottom of the well . The only high-power lamps known are those using xenon, which however achieve sensitivities of the order of 10"5 g/ml of detected sample. Even if the sensitivity could be improved, such lamps are impractical, not only because of the long time required to reach their operational temperatures, but also because they dissipate much heat, thereby requiring large ventilating means as otherwise the optics would rapidly degrade or even be destroyed. Moreover, it must be borne in mind that these known lamps for fluorescence analysis on account of the blowers and devices supplying sufficient electric power generate background noise lowering the sensitivity of the analytical apparatus .
Accordingly the expert must fail to resolve the above problem. Summary
The above object of the invention as well as further ones are achieved by an illuminator for a fluorescence analyzer comprising an optics, at least one support for the products to be analyzed and an analytical device, where said, illuminator is characterized in the present invention by comprising on one hand a lamp of which the emission axis is collinear with the axis of the light flux entering the optics, said lamp emitting a flux of at least 2,000 lumens and evincing a mean luminous efficiency of at least 30 lumens/watt and a flux density of at least 1,200 x 106 lumens/m3, and on the other hand a heat dissipator.
Advantageously the lamp shall be argon-iodide, xenon-flash or rare-gas/halogen, preferably xenon, for instance a halogen-krypton-xenon lamp.
Advantageously the heat dissipator shall be an infrared trap illustratively consisting of cups arranged in a semi-cylindrical envelope enclosing the rear part of the lamp in relation to the direction of the emitted flux.
Preferably the illuminator also includes an interference excitation filter located in front of the lamp as seen in the direction of the emitted light beam.
Advantageously the support for the product being analyzed consists of at least one microtitration well or of two (microscope) slides between which said product is placed. In a preferred embodiment mode of the invention, the fluorescence analyzer is a microtitration fluorimeter especially as regards cell biology.
BRIEF DESCRIPTION OF THE DRAWINGS The following description implies no limitation and must be considered in relation to the attached Figures : Fig. 1 schematically shows a fluorimeter of the prior art
Fig. 2 schematically shows the same fluorimeter now fitted with an illuminator of the invention. Detailed Description of the Invention
In order to more clearly explain the advantages of the present invention to the expert, the illuminator is described in relation to the fluorimeter which is the object of the US patent 4,501,970. This fluorimeter comprises a lamp 1 mounted just in front of a diaphragm 2 constituting the entry into a tunnel receiving an optics such as previously described. The light beam 3 entering the tunnel then is directed toward a microtitration well 4 holding the solution which must be analyzed. The reflected beam 5 is directed toward an analyzer 6.
Lamp 1 is a mercury-vapor lamp emitting in the ultraviolet and with an axis perpendicular to that of the light beam 3. The lamp's light flux crossing the diaphragm 2 is about 1,300 lumens, its mean luminous efficiency being about 26 lumen/watt.
In the present invention the lamp 11 is a conventional halogen-krypton-xenon lamp called krypton- xenon by the specialists; it might also be of the argon- iodide, xenon-flash or halogen/rare-gaseε type, one of these preferably being xenon. The axis of this lamp 11 is collinear with the axis of the diaphragm 12, that is, it is collinear with the direction of the mitted flux. The light flux 13 emitted by such a lamp 11 is at least 2,000 lumen, for instance about 2,300 lumen.
This lamp 11 is characterized by a mean luminous efficiency of at least 30 lumen/watt and by a luminous flux density of at least 1,200 x 106/ro3- Accordingly this is a small bulb requiring no electric-power regulation nor any heat-up time.
As is clear to the expert, such a lamp is unidirectional: it might be called a photon gun. Therefore a maximum energy shall enter the analyzer.
These lamps emit a light beam 13 of which the spectrum ranges from ultraviolet to visible, whereas the prior art lamp emits in the ultraviolet or in the visible. Moreover this lamp comprises an infrared trap 14 allowing heat dissipation and thereby prevents overheating the components of the optics. Accordingly not only the maximum light flux from the lamp 11 enters the tunnel through the diaphragm 12, but furthermore only the photon power. This is highly important because the heat power emitted by this lamp unless trapped would degrade and possibly destroy at least some electronic or optical .components.
Illustratively the infrared trap 14 consists of cups arranged within a semi-cylindrical envelope enclosing the rear part of the lamp relative to the diaphragm 12.
As understood by the expert, such a lamp 11 fitted with its infrared trap 14 must be located in front of the diaphragm 12 at such a distance that the light beam crossing this diaphragm may be considered cold.
Moreover, in order to improve the quality of the light beam 13 from the lamp 11, the illuminator may include an interference excitation filter 15 in front of the diaphragm 12 : only those photons useful in the measurement shall enter the optics. Such a fluorimeter allows achieving sensitivities of 10"10 to 10"15 g/ml of detected sample, for instance in a cell culture.
Even though the present invention was described in relation to a fluorescence analyzer which is the object of the US patent 4,501,970, it is clear per se that the illuminator of the present invention can be matched to any other kind of fluorimeter and even to apparatus using fluoresgence for analysis, for instance a fluorescence microscope of which the support for the product to be analyzed consists of two slides between which the product is held.

Claims

1. An illuminator for a fluorescence analyzer comprising an optics, at least one support for the product to be analyzed and an analyzer, characterized in that it comprises on one hand a lamp (11) of which the emission axis is collinear with the axis of the emitted light flux entering the optics, said light flux being at least 2,000 lumens and the mean luminous efficiency being at least 30 lumen/watt, the mean flux density being at least 1,200 x 106 lutnen/m3, and on the other hand a heat dissipator (14) .
2. Illuminator defined in that the lamp (11) is an argon-iodide, xenon flash or rare-gas/halogen lamp, preferably a xenon lamp.
3. Illuminator defined in claim 1, characterized in that the lamp (11) is a halogen-krypton-xenon lamp.
4. Illuminator defined in claim 1, characterized in that the heat dissipator is an infrared trap (14) .
5. Illuminator defined in claim 4, characterized in that the infrared trap (14) consists of cups mounted within a semi-cylindrical envelope enclosing the rear of the lamp (11) in relation to the direction of the emitted light beam.
6. Illuminator defined in any of claims 1 through
5, characterized in that it also includes an interference excitation filter (15) in front of the lamp as seen in the direction of the emitted light flux.
7. Illuminator defined in any of claims 1 through
6, characterized in that the support for the product to be analyzed consists of at least one microtitration well.
8. Illuminator defined in any of claims 1 through 6, characterized in that the support for the product to be analyzed consists of two slides between which the said product is being held.
9. Microtitration fluorimeter, to be used in particular for cell biology, characterized in that it comprises an illuminator defined in any of claims 1 through 6.
PCT/US1993/009249 1992-09-30 1993-09-30 Illuminator for fluorescence-analyzer such as a microtitration fluorimeter used in cell biology WO1994008352A1 (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
JP6509273A JPH08505697A (en) 1992-09-30 1993-09-30 Lighting equipment for fluorescence analyzers such as microtiter fluorometers used in cell biology

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
FR9211648A FR2696235B1 (en) 1992-09-30 1992-09-30 Illumination device for fluorescence analysis system, for example fluorimeter for microtitration, especially in cell biology.
FR92/11648 1992-09-30

Publications (1)

Publication Number Publication Date
WO1994008352A1 true WO1994008352A1 (en) 1994-04-14

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EP (1) EP0591034B1 (en)
JP (1) JPH08505697A (en)
DE (1) DE69322825T2 (en)
FR (1) FR2696235B1 (en)
WO (1) WO1994008352A1 (en)

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6246898B1 (en) 1995-03-28 2001-06-12 Sonometrics Corporation Method for carrying out a medical procedure using a three-dimensional tracking and imaging system

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US4495516A (en) * 1982-09-29 1985-01-22 Eastman Kodak Company Film video player having flash illuminated area image sensor and single frame CCD image sensor for use therewith
US4501970A (en) * 1982-10-12 1985-02-26 Dynatech Laboratories Incorporated Fluorometer
JPS60260830A (en) * 1984-06-07 1985-12-24 Fujisawa Pharmaceut Co Ltd Light source device for irradiating cell in automatic analysis instrument for cell
US4626684A (en) * 1983-07-13 1986-12-02 Landa Isaac J Rapid and automatic fluorescence immunoassay analyzer for multiple micro-samples
US4656790A (en) * 1984-12-04 1987-04-14 Fuji Photo Film Co., Ltd. Burnishing method and apparatus for magnetic disk
US4868103A (en) * 1986-02-19 1989-09-19 Enzo Biochem, Inc. Analyte detection by means of energy transfer
US4900933A (en) * 1986-09-08 1990-02-13 C. R. Bard, Inc. Excitation and detection apparatus for remote sensor connected by optical fiber

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US3854050A (en) * 1973-09-11 1974-12-10 Department Of Health Education High precision fluorometer for measuring enzymatic substrates in tissue
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JPS5681813A (en) * 1979-12-08 1981-07-04 Nippon Telegr & Teleph Corp <Ntt> Mask lighting optical system
JPS6070401A (en) * 1983-09-28 1985-04-22 Toshiba Electric Equip Corp Device for irradiating uv light
JPS60202404A (en) * 1984-03-28 1985-10-12 Toshiba Electric Equip Corp Irradiating device of ultraviolet rays
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Publication number Priority date Publication date Assignee Title
US3947124A (en) * 1973-10-26 1976-03-30 National Research Development Corporation Analytical spectroelectrochemistry
US4495516A (en) * 1982-09-29 1985-01-22 Eastman Kodak Company Film video player having flash illuminated area image sensor and single frame CCD image sensor for use therewith
US4501970A (en) * 1982-10-12 1985-02-26 Dynatech Laboratories Incorporated Fluorometer
US4626684A (en) * 1983-07-13 1986-12-02 Landa Isaac J Rapid and automatic fluorescence immunoassay analyzer for multiple micro-samples
JPS60260830A (en) * 1984-06-07 1985-12-24 Fujisawa Pharmaceut Co Ltd Light source device for irradiating cell in automatic analysis instrument for cell
US4656790A (en) * 1984-12-04 1987-04-14 Fuji Photo Film Co., Ltd. Burnishing method and apparatus for magnetic disk
US4656790B1 (en) * 1984-12-04 1989-01-10
US4868103A (en) * 1986-02-19 1989-09-19 Enzo Biochem, Inc. Analyte detection by means of energy transfer
US4900933A (en) * 1986-09-08 1990-02-13 C. R. Bard, Inc. Excitation and detection apparatus for remote sensor connected by optical fiber

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US6246898B1 (en) 1995-03-28 2001-06-12 Sonometrics Corporation Method for carrying out a medical procedure using a three-dimensional tracking and imaging system

Also Published As

Publication number Publication date
FR2696235A1 (en) 1994-04-01
EP0591034A2 (en) 1994-04-06
EP0591034B1 (en) 1998-12-30
EP0591034A3 (en) 1994-04-27
FR2696235B1 (en) 1994-10-28
JPH08505697A (en) 1996-06-18
DE69322825D1 (en) 1999-02-11
DE69322825T2 (en) 1999-10-21

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