US20080176307A1 - Method of isolating cell or sample to be analyzed in cell - Google Patents
Method of isolating cell or sample to be analyzed in cell Download PDFInfo
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- US20080176307A1 US20080176307A1 US11/834,419 US83441907A US2008176307A1 US 20080176307 A1 US20080176307 A1 US 20080176307A1 US 83441907 A US83441907 A US 83441907A US 2008176307 A1 US2008176307 A1 US 2008176307A1
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/10—Processes for the isolation, preparation or purification of DNA or RNA
- C12N15/1003—Extracting or separating nucleic acids from biological samples, e.g. pure separation or isolation methods; Conditions, buffers or apparatuses therefor
Abstract
There is disclosed a method of irradiating a specimen which is laid on a substrate and which includes a cell with a laser beam to remove an unnecessary specimen portion other than a cell portion from the substrate, next inserting the cell portion into one opening in a cylindrical member to dispose the cylindrical member onto the substrate, next injecting a stripping agent into a container formed by the substrate and cylindrical member to strip the cell portion from the substrate, and sampling the cell portion which floats in the stripping agent.
Description
- The present application is a Divisional Application of U.S. application Ser. No. 10/348,198 filed Jan. 21, 2003, which claims the benefit of priority from the prior Japanese Patent Application No. 2002-14643, filed Jan. 23, 2002, the entire contents of which are incorporated herein by reference.
- 1. Field of the Invention
- The present invention relates to a method comprising: selecting a specific cell or cells to be analyzed from a specimen on a substrate; and isolating the cell or cells to be analyzed.
- 2. Description of the Related Art
- At present, a research to analyze a function of a gene has been a matter of greatest concern. In the research of analysis of the gene function, cells are acquired from a specimen. A sample for the analysis of the gene function is directly sampled from the cells. Various types of an apparatus for sampling the cells or sample have been developed and used.
- Various cell sampling methods for sampling the cells from a slice of a specimen will be described.
- (1) A slice of a specimen is fixed onto a thin film plate. The slice of the specimen includes, for example, the cells to be sampled for the analysis of the functions of the gene. The thin film plate is irradiated with a laser beam of an ultraviolet wavelength region (UV laser beam) along a contour of the cells. Thereby, the cells to be sampled in the specimen are cut/separated together with the thin film plate.
- Next, a cut/separated portion is irradiated with the UV laser beam out of focus with respect to the thin film plate. Thereby, the cut/separated portion in the specimen is flied. As a result, the cells to be sampled are acquired.
- (2) A transparent cap for sampling to which a transfer film is attached is used. The transparent cap is laid on the specimen which has the cells to be sampled. The portion including the cells to be sampled in the specimen are irradiated with an IR laser beam through the transparent cap. Thereby, the cell portion to be sampled adheres to a transfer film surface.
- (3) The specimen is attached onto the film. The surface to which the specimen is attached on the film is turned downwards. In this state, the UV laser beam is emitted along the contour of the cells from above. Thereby, the periphery of the cells to be sampled are cut off.
- Next, the film is irradiated with the UV laser beam. Thereby, the cells to be sampled are detached from the film and dropped into a tray which is set below.
- However, the method (1) comprises: cutting/separating the cells to be sampled together with the thin film plate; and subsequently irradiating the cut/separated portion with the UV laser beam out of focus to fly the portion. Therefore, a place of the cells to be sampled are not known. The cells are lost in many cases. Moreover, it is also expected that properties of the cells are changed by the UV laser beam.
- Moreover, foreign particles such as dust are easily mixed in the sampled cells. It is necessary to fix the specimen onto the thin film plate beforehand. Therefore, the sampling operation of the cells requires troubles and is laborious.
- The method (2) comprises: irradiating the cells to be sampled in the specimen with the IR laser beam transmitted through the transparent cap. Therefore, a spot diameter of the IR laser beam with which the cells to be sampled are irradiated cannot be reduced. The spot diameter of the IR laser beam is also sometimes larger than the size of the cells to be sampled. When the cells are irradiated with the laser beam having the spot diameter larger than the size of the cells to be sampled, together with the cells to be sampled, unnecessary surrounding cells also stick to the transfer film surface. For example, to sample a micro cell of five microns or less with a spot diameter of ten microns, the surrounding unnecessary cells also stick to the transfer film surface.
- Since the cells to be sampled are attached to the transfer film surface, depending on a dry degree of the cells, the cells cannot well stick to the transfer film surface, and the cells cannot sometimes be sampled. It is impossible to sample the cells in a wet state. The sticking of the cells to the transfer film surface is influenced by conditions of the cells such as the dry degree of the cells. In other words, the capability of sampling the cells is limited by the conditions of the cells.
- With an increase of the output of the IR laser beam with which the specimen is irradiated, an efficiency of attaching the cell portion to the transfer film surface to sample the cells is improved. On the other hand, when the output of the IR laser beam is increased, the cells burn.
- The method (3) comprises: dropping the cell portion sampled by the irradiation with the UV laser beam down to the tray which is set below. Therefore, the place of the cells to be sampled is not known. Moreover, the foreign particles such as dust are easily mixed in the acquired cells. It is also expected that the properties of the cells are changed by the UV laser beam.
- The above-described problems are summarized as follows
- (a) It takes much time to sample the cells to be sampled. Additionally, the cells are easily lost at a sampling time.
- (b) It is difficult to acquire small cells. The cells which can be sampled are limited by a dry state of the specimen.
- (c) It is impossible to acquire a large amount of cells having sufficient qualities in a short time by causes such as the change of the properties of the cells by the UV laser beam.
- According to a main viewpoint of the present invention, there is provided a method of isolating a cell or a group of cells, comprising: irradiating a specimen which is laid on a substrate and which includes the cells with a laser beam to remove an unnecessary specimen portion other than a necessary cell portion from the substrate; using a cylindrical member including openings in opposite ends; inserting the cell portion into one opening in the cylindrical member and disposing the cylindrical member onto the substrate; injecting a stripping agent into a container formed by the substrate and cylindrical member to strip the cell portion from the substrate; and sampling the cell portion which is floated in the stripping agent.
- According to the main viewpoint of the present invention, there is provided a method of isolating a cell or a group of cells or a sample to be analyzed in the cells, comprising: irradiating a specimen which is laid on a substrate and which includes the cells with a laser beam to remove an unnecessary specimen portion other than a necessary cell portion from the substrate; using a cylindrical member including openings in opposite ends; inserting the necessary cell portion into one opening in the cylindrical member and disposing the cylindrical member onto the substrate; injecting a solution for eluting the sample existing in the cells into a container formed by the substrate and cylindrical member; and sampling the sample eluted in the solution.
- According to the main viewpoint of the present invention, there is provided a method of isolating a cell or a group of cells, comprising: irradiating a specimen which is laid on a substrate and which includes the cells with a laser beam to change properties of an unnecessary specimen portion other than a necessary cell portion; using a cylindrical member including openings in opposite ends; inserting the necessary cell portion into one opening in the cylindrical member and disposing the cylindrical member onto the substrate; injecting a stripping agent into a container formed by the substrate and cylindrical member to strip the necessary cell portion from the substrate; and sampling the necessary cell portion which floats in the stripping agent.
- According to the main viewpoint of the present invention, there is provided a method of isolating a cell or a group of cells or a sample to be analyzed in the cells, comprising: irradiating a specimen which is laid on a substrate and which includes the cells with an ultraviolet light to change properties of an unnecessary specimen portion other than a necessary cell portion; using a cylindrical member including openings in opposite ends; inserting the necessary cell portion into one opening in the cylindrical member and disposing the cylindrical member onto the substrate; injecting a stripping agent into a container formed by the substrate and cylindrical member to strip the necessary cell portion from the substrate; and sampling the necessary cell portion which is floated in the stripping agent.
- Additional objects and advantages of the invention will be set forth in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. The objects and advantages of the invention may be realized and obtained by means of the instrumentalities and combinations particularly pointed out hereinafter.
- The accompanying drawings, which are incorporated in and constitute a part of the specification, illustrate presently preferred embodiments of the invention, and together with the general description given above and the detailed description of the preferred embodiments given below, serve to explain the principles of the invention.
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FIG. 1 is a constitution diagram of an inverted microscope for use in a first embodiment of a cell acquiring method of the present invention; -
FIG. 2 is a diagram showing a cylindrical member for use in sampling cells in the first embodiment; -
FIG. 3 is a diagram showing a trimming of the cells in first and second embodiments; -
FIG. 4 is a diagram showing a trimming of the cells in the first and second embodiments; -
FIG. 5 is a diagram showing an injection of a stripping agent in the first and second embodiments; -
FIG. 6 is a diagram showing a stripped state of the cells in the first and second embodiments; -
FIG. 7 is a constitution diagram of the inverted microscope for use in the second embodiment of the cell acquiring method of the present invention; -
FIG. 8 is a diagram showing irradiation of an unnecessary cells with laser in the first and second embodiments; and -
FIG. 9 is a diagram showing one example of a sampling method of the cells in the first and second embodiments. - A first embodiment of the present invention will be described hereinafter with reference to the drawings.
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FIG. 1 is a constitution diagram of an inverted microscope for use in a cell acquiring method of the present invention. In a light source forlighting 1, for example, a halogen lamp is used. The light source forlighting 1 outputs visible rays. Aview field stop 2,aperture stop 3, anddichroic mirror 4 are disposed on a light path of visible rays outputted from the light source forlighting 1. - The
view field stop 2 andaperture stop 3 constitute an optical system of a conventional microscope. Thedichroic mirror 4 reflects the visible rays toward a lower part of an optical axis P. - An objective lens for
laser 5 is disposed on a reflected light path (optical axis P) of thedichroic mirror 4. The objective lens forlaser 5 collects the visible rays outputted from the light source forlighting 1 to irradiate aspecimen 7. The objective lens forlaser 5 functions as a condenser lens. - A
stage 6 is disposed on the reflected light path of thedichroic mirror 4. Aslide glass 8 on which thespecimen 7 is laid is laid on thestage 6. Thestage 6 includesmotors motor 6 a of the X direction is driven to move thestage 6 in the X direction. Themotor 6 b of the Y direction is driven to move thestage 6 in the Y direction. Astage driving unit 9 drives therespective motors - The
specimen 7 is a tissue slice or cultured cells for use, for example, in analyzing a gene function. When a nucleus of the cell is fluori-dyed to observe fluorescence and to acquire the cell, thespecimen 7 is applied with fluorescence dyestuff beforehand. - An objective lens for
observation 10,dichroic mirror 11,UV barrier filter 12,image forming lens 13, andhalf mirror 14 are disposed on the optical axis P transmitted through thespecimen 7. - A
CCD camera 15 is disposed on the reflected light path of thehalf mirror 14. TheCCD camera 15 is disposed in an image forming position of theimage forming lens 13 via thehalf mirror 14. TheCCD camera 15 picks up an image formed by theimage forming lens 13 to output an image signal. - An image processing and
stage control apparatus 16 inputs the image signal outputted from theCCD camera 15, image-processes the image signal to acquire image data, and displays the image data in adisplay 17. - The image processing and
stage control apparatus 16 is connected tooperation units 18 such as a keyboard and mouse. Theoperation unit 18 receives an operation of a movement direction and movement amount of thestage 6 by an observer, and generates this operation command. The image processing andstage control apparatus 16 receives the operation command from theoperation unit 18, and transmits a movement control signal of thestage 6 to thestage driving unit 9 in accordance with the operation command. - A
mirror 19 is disposed on a transmitted light path of thehalf mirror 14.Relay lenses 20, UV cutfilter 21, andeyepiece lens 22 are disposed on the reflected light path. - On the other hand, a
UV laser head 23 is disposed on the optical axis P passed through the objective lens forlaser 5 anddichroic mirror 4. A UVimage forming lens 24 is disposed on the optical axis p between theUV laser head 23 anddichroic mirror 4. - The
UV laser head 23 includes alaser beam source 23 a. TheUV laser head 23 outputs a laser beam having a wavelength of an ultraviolet region. The laser beam outputted from theUV laser head 23 removes unnecessary cells other than the cell to be sampled among a plurality of cells included in thespecimen 7 from theslide glass 8. - Energy of the laser beam outputted from the
UV laser head 23 is set to such a degree that the unnecessary cell on theslide glass 8 is removed from theslide glass 8. - The
UV laser head 23 includes avariable slit 25 andLED 26. The variable slit 25 includes, for example, a circular or quadrilateral opening. The variable slit 25 can vary the diameter of the opening. The variable slit 25 varies the spot diameter of the laser beam outputted from theUV laser head 23. - The
LED 26 outputs an LED light. The LED light is a light having a wavelength of the visible ray, such as a red visible ray. - A
dichroic mirror 27 is disposed on the light path of the LED light. Thedichroic mirror 27 reflects the LED light outputted from theLED 26 toward the optical axis p of the laser beam outputted from theUV laser head 23. - Thereby, the LED light passes through the
dichroic mirror 27 which has a transmission property with respect to the wavelengths of the LED light and UV laser beam, and the same position as the irradiation position of the laser beam is irradiated in thespecimen 7. The red LED light with which thespecimen 7 is irradiated displays the irradiation position of the laser beam with which thespecimen 7 is irradiated. - A light source for
fluorescence excitation 28 and UVband pass filter 29 are disposed on the reflected light path of thedichroic mirror 11. The light source forfluorescence excitation 28 is disposed below thestage 6. The light source forfluorescence excitation 28 outputs an ultraviolet light. The light source forfluorescence excitation 28 is, for example, a mercury lamp. - The ultraviolet light outputted from the light source for
fluorescence excitation 28 is transmitted through the UVband pass filter 29, and reflected by thedichroic mirror 11, and thespecimen 7 is irradiated through the objective lens forobservation 10. The ultraviolet light outputted from the light source forfluorescence excitation 28 excites the fluorescence dyestuff applied to thespecimen 7 beforehand. - An
air blow 30 is disposed obliquely above thestage 6. Theair blow 30 sprays air toward thespecimen 7 laid on thestage 6. When the unnecessary cells in thespecimen 7 is removed from theslide glass 8, a broken piece of the unnecessary cells is generated. The broken piece of the unnecessary cells is blown/flied from theslide glass 8 by the air sprayed by theair blow 30. - An
air suction unit 31 is disposed obliquely above thestage 6. Theair suction unit 31 is disposed opposite to theair blow 30 via thespecimen 7 on thestage 6. Theair suction unit 31 sucks air, and sucks the broken pieces of the cells flied by the air spraying of theair blow 30. - Next, a sampling method of the cells existing in the
specimen 7 will be described. - The
specimen 7 is laid on theslide glass 8. Theslide glass 8 is laid on thestage 6. - The light source for
lighting 1 outputs the visible ray. The visible ray is passed through theview field stop 2 andaperture stop 3, and reflected by thedichroic mirror 4, and thespecimen 7 is irradiated through the objective lens forlaser 5. A part of the visible ray with which thespecimen 7 is irradiated passes through thespecimen 7. - The light passed through the
specimen 7 passes through the objective lens forobservation 10, is transmitted through thedichroic mirror 11 andUV barrier filter 12, and is formed into the image by theimage forming lens 13. - The
CCD camera 15 picks up an enlarged image of thespecimen 7 formed by theimage forming lens 13 via thehalf mirror 14 to output the image signal. - The image processing and
stage control apparatus 16 inputs the image signal outputted from theCCD camera 15, image-processes the image signal to acquire the image data, and displays the image data in thedisplay 17. - Additionally, the image of the
specimen 7 formed by theimage forming lens 13 is reflected by themirror 19, and incident upon theeyepiece lens 22 through therelay lenses 20 and UV cutfilter 21. The observer sees through theeyepiece lens 22 to visually observe the enlarged image of thespecimen 7. - The observer observes the
specimen 7 displayed in thedisplay 17 or sees through theeyepiece lens 22 to observe the enlarged image of thespecimen 7 and determines the cell to be sampled from thespecimen 7. - To sample cells a from the
specimen 7, acylindrical member 40 shown inFIG. 2 is used. Thecylindrical member 40 is formed, for example, of a plastic, metal, or glass material. Thecylindrical member 40 is formed in a cylindrical shape, and includes circular openings formed in opposite ends. The diameter of thecylindrical member 40 may preferably be larger than that of the cells a to be sampled. For the size of thecylindrical member 40, in one example an outer diameter is 4 to 11 mm, inner diameter is 3 to 10 mm, and length is 5 to 10 mm. The size of thecylindrical member 40 is sometimes smaller than that of the example. - When the
specimen 7 is larger than the inner diameter of thecylindrical member 40, a specimen portion d including the cells a to be sampled is trimmed. When the shape of the opening of thecylindrical member 40 is circular, the specimen portion d including the sampled cells a in thespecimen 7 is trimmed in a circular shape as shown inFIG. 3 . When thespecimen 7 is smaller than the inner diameter of thecylindrical member 40, the trimming is not necessary. - The
cylindrical member 40 is not limited to a member including each circular opening, and may also have another shape opening. For example, when the opening of thecylindrical member 40 has a quadrilateral shape, the shape of the specimen portion d to be trimmed is a quadrilateral shape including the cells a to be sampled in thespecimen 7 as shown inFIG. 4 . - The trimming of the specimen portion d including the cells a to be sampled will be described.
- In the variable slit 22 of the
UV laser head 21, for example, the circular opening is used. For the variable slit 22, the slit diameter is set, for example, to a spot diameter s1 which is substantially equal to a thickness S1 of a wall of thecylindrical member 40 as shown inFIG. 2 . - On the other hand, the
LED 26 outputs the red LED light. The LED light is reflected by thedichroic mirror 27, and incident upon thevariable slit 25. When the LED light passes through the opening of thevariable slit 25, a circular LED light having the spot diameter s1 is shaped. - The
specimen 7 is irradiated with the LED light having the spot diameter s1 through the UVimage forming lens 24 and objective lens forlaser 5. At this time the LED light with the spot diameter s1 transmitted through thedichroic mirror 4 proceeds on the optical axis P of the laser beam outputted from theUV laser head 23. The same position as the irradiation position of the laser beam in thespecimen 7 is irradiated with the LED light with the spot diameter s1 in red. - The
CCD camera 15 picks up the enlarged image of thespecimen 7 to output the image signal. The image processing andstage control apparatus 16 inputs the image signal outputted from theCCD camera 15, image-processes the image signal to acquire the image data, and displays the image data of thespecimen 7 irradiated with the red LED light with the spot diameter s1 in thedisplay 17. - The observer observes the
specimen 7 displayed in theCCD camera 15 while leaving the specimen portion d, and determines an irradiation route R1 of the laser beam to remove the portion of thespecimen 7 around the specimen portion d from theslide glass 8. - When the
cylindrical member 40 with the circular opening is used, the irradiation route R1 of the laser beam is circular as shown inFIG. 3 . When thecylindrical member 40 having the quadrilateral opening is used, the irradiation route R1 of the laser beam is quadrilateral as shown inFIG. 4 . - Next, the
UV laser head 23 outputs the laser beam, for example, every shot. The energy of the laser beam is set to such a degree that the outer peripheral portion of the unnecessary specimen portion d on theslide glass 8 is removed from theslide glass 8. It is to be noted that the energy of the laser beam is preferably set to such a degree that the unnecessary portion other than the cells a to be sampled is completely blown off even with slight humidity in ambient environment. - The irradiation route R1 of the
specimen 7 is irradiated with the laser beam through the UVimage forming lens 24,dichroic mirror 4, and objective lens forlaser 5. - The observer confirms the irradiation position of the LED light on the
specimen 7 displayed in thedisplay 17 or sees through theeyepiece lens 22 to confirm the irradiation position of the LED light. The observer confirms that the irradiation position of the LED light is on the irradiation route R1 while operating theoperation unit 18 such as the keyboard and mouse. - The image processing and
stage control apparatus 16 receives the operation command in accordance with the operation of the movement direction and movement amount of thestage 6 from theoperation unit 18, and sends the movement control signal of thestage 6 to thestage driving unit 9 in accordance with the operation command. - The
stage 6 is moved in the XY direction, for example, every shot of laser beam so that the irradiation position of the LED light on thespecimen 7 is disposed on the irradiation route R1. By the movement of thestage 6 in the XY direction, each shot of the laser beam moves on the irradiation route R1 to irradiate thespecimen 7. - Each portion of the
specimen 7 on the irradiation route R1 irradiated with the laser beam is removed from theslide glass 8. Thereby, as shown inFIG. 3 , the specimen portion d including the cells a to be sampled is trimmed in a circular shape. Moreover, as shown inFIG. 4 , the specimen portion d including the cells a to be sampled is trimmed in a quadrilateral shape. - It is to be noted that XY coordinate positions of irradiation routes R1, R2 may also be stored in the image processing and
stage control apparatus 16 beforehand. That is, after the respective irradiation routes R1, R2 of the laser beam are determined, thespecimen 7 is irradiated with the LED light. The observer operates theoperation unit 18 to move thestage 6 in the XY direction, and the LED light is moved along the irradiation routes R1, R2. - At this time, the image processing and
stage control apparatus 16 reads the movement direction and amount of thestage 6 from theoperation unit 18, and calculates the XY coordinate positions of thestage 6 on the irradiation routes R1, R2 from the movement direction and amount. The image processing andstage control apparatus 16 stores the calculated XY coordinate positions of the irradiation routes R1, R2 in an internal memory. - The image processing and
stage control apparatus 16 reads the XY coordinate positions of the respective irradiation routes R1, R2 stored in the internal memory, and sends a movement control signal of thestage 6 to thestage driving unit 9 in accordance with the XY coordinate positions. - The
stage 6 automatically moves the irradiation position with the laser beam in the XY direction on the irradiation routes R1, R2. Additionally, theUV laser head 23 outputs one shot of laser beam every predetermined period. - The portions of the
specimen 7 on the irradiation routes R1, R2 irradiated with the laser beam are removed from theslide glass 8. Thereby, the specimen portion d including the cells a to be sampled is automatically trimmed in the circular shape as shown inFIG. 3 . Moreover, the specimen portion d including the cells a to be sampled is automatically trimmed in the quadrilateral shape as shown inFIG. 4 . - When the
specimen 7 is irradiated with the laser beam, the broken piece of thespecimen 7 is cut off. The cut broken piece flies/scatters, for example, above theslide glass 8. - The
air blow 30 blows air above theslide glass 8. The broken piece of thespecimen 7 flying/scattering above theslide glass 8 is flied from above theslide glass 8 by the air blown from theair blow 30. - The
air suction unit 31 sucks the air, and sucks the broken piece of thespecimen 7 flied by the air blowing of theair blow 30. - Thereby, the broken piece of the
specimen 7 flying/scattering above theslide glass 8 is removed from theslide glass 8. - The specimen portion d may also be trimmed in the following method.
- The
specimen 7 is applied with the fluorescence dyestuff beforehand. - The light source for
fluorescence excitation 28 emits an ultraviolet light. The ultraviolet light emitted from the light source forfluorescence excitation 28 is transmitted through the UVband pass filter 29 and incident upon thedichroic mirror 11. The ultraviolet light is reflected upwards by thedichroic mirror 11, and thespecimen 7 is irradiated with the light through the objective lens forobservation 10. The fluorescence dyestuff applied to thespecimen 7 is excited and emits. - The fluorescence from the
specimen 7 is transmitted through thedichroic mirror 11 andUV barrier filter 12 from the objective lens forobservation 10, and formed into the image by theimage forming lens 13. - The
CCD camera 15 picks up the enlarged image and fluorescence of thespecimen 7 to output the image signal. The image processing andstage control apparatus 16 inputs the image signal outputted from theCCD camera 15, image-processes the image signal to acquire the image data, and displays the image data of thespecimen 7 emitting the fluorescence in thedisplay 17. - Additionally, the image and fluorescence of the
specimen 7 are reflected by themirror 19, and pass through therelay lenses 20 and UV cutfilter 21 to be incident upon theeyepiece lens 22. Thereby, the observer sees through theeyepiece lens 22 to visually observe the enlarged image of thespecimen 7 emitting the fluorescence. - The observer observes the enlarged image of the
specimen 7 displayed in thedisplay 17, or sees through theeyepiece lens 22 to observe the enlarged image and fluorescence of thespecimen 7. The observer can observe the enlarged image and fluorescence of thespecimen 7 to determine the cells a to be sampled from thespecimen 7. - Since the fluorescence dyestuff applied to the
specimen 7 emits the light, as compared with the use of the visible ray, the cells a to be sampled is further easily determined. The image can also be recognized using a computer to automatically select the cells a to be sampled. - Next, a procedure of leaving only the cells a to be sampled and removing other cells from the specimen d including the cell a to be sampled.
- For the variable slit 25 maximizes the diameter of the opening. The variable slit 25 enlarges the spot diameter of the laser beam outputted from the
UV laser head 23. As shown inFIG. 8 , the cells a detached from the cells a to be sampled are irradiated with the laser beam which has a large spot diameter s2. - Next, the variable slit 25 reduces the diameter of the opening. The periphery of the cells a to be sampled are irradiated with the laser beam having a small spot diameter s3 more carefully, that is, correctly along the periphery of the cells a. Only the cells a to be sampled are left.
- Next, the
cylindrical member 40 shown inFIG. 2 is prepared. An adhesive 41 is applied to one opening of thecylindrical member 40. Thecylindrical member 40 is bonded to the substrates such as theslide glass 8 using grease, paraffin, and various adhesives which are not influenced by the solution for stripping or extracting. Moreover, a disposable thick seal or cylindrical member which is coated with the adhesive beforehand may also be used. - The
cylindrical member 40 is disposed vertically to the upper surface of theslide glass 8 while one operation is directed downwards. When thecylindrical member 40 is disposed on the upper surface of theslide glass 8, the specimen portion d including the cells a are inserted into one opening. - Next, the
cylindrical member 40 is lightly pressed against theslide glass 8 side. This secures the adhesion of thecylindrical member 40 to theslide glass 8. When the adhesive 41 hardens, a liquid-tight container is formed by thecylindrical member 40 andslide glass 8. Thecylindrical member 40 andslide glass 8 are removed from thestage 6. - Next, a stripping
agent 42 is injected into the container formed by thecylindrical member 40 andslide glass 8 as shown inFIG. 5 . Examples of the strippingagent 42 include organic solvents such as xylene and surfactants. The strippingagent 42 promotes the stripping of the cells a from theslide glass 8.FIG. 6 shows the cells a stripped from theslide glass 8. The stripped cells a are floated in the strippingagent 42. - Next, the cells a floating in the stripping
agent 42 is sucked together with the strippingagent 42 by a dropper. The cells a sucked by the dropper is transferred into another new container. As a result, the sampling of the cells a ends. - In this manner, according to the first embodiment, the method comprises: inserting the specimen portion d smaller than the inner diameter of the opening of the
cylindrical member 40 into the opening and disposing thecylindrical member 40 on theslide glass 8; forming a portion between thecylindrical member 40 andslide glass 8 by the adhesive 41 in the liquid-tight manner; injecting the strippingagent 42 into the container formed by thecylindrical member 40 andslide glass 8; and stripping the cells a to be sampled from theslide glass 8 and floating the cells in the strippingagent 42 to sample the cells. - Therefore, the cells a to be sampled included in the
specimen 7 can securely be sampled. This prevents the position of the cells a from being unknown as in the related art. The cells a cannot be lost. The foreign particles such as dust can securely be prevented from being mixed in the sampled cells a. - In the first embodiment, since the
specimen 7 is first laid on theslide glass 8, any special thin film plate or film is not used as in the related art. The operation of attaching thespecimen 7 to the thin film plate or film can be eliminated. Thereby, in the first embodiment, the sampling operation of the cells a can be simplified and efficiently performed. Additionally, this is economically advantageous. - In the above-described first embodiment, since the transparent cap for sampling the cells in the related art is not used, the spot diameter of the laser beam can be reduced. Therefore, even when the cell a to be sampled is micro, the cells a to be sampled is left, and the other cells can be irradiated with the laser beam.
- In the first embodiment, special materials such as a transfer film are not used. Therefore, a trouble of operation is saved, and expenses of consumables can be saved.
- In general, when the function of the cell or the gene included in the cells is analyzed, the mixture of the cells other than the targeted cells or the gene is requested to be reduced to enhance the purity of the sample. Moreover, since it takes much time to select and acquire the necessary cells from the specimen cut piece, there has been a demand for efficiency in acquiring the cells.
- On the other hand, according to the first embodiment, since the unnecessary cells are removed and only a large number of necessary cells can be acquired at once, it is possible to efficiently acquire the high-purity sample.
- The above-described embodiment comprises: injecting the stripping
agent 42 into thecylindrical member 40; and stripping the cells a to be sampled from theslide glass 8 and floating the cells to sample the cells. This cell sampling method is effective in a research in which the shape of the nucleus of the cells a need to be held as completely as possible, for example, analysis of a flow cytometry. - The first embodiment can appropriately be modified and carried out.
- For example, the adhesive 41 is not necessarily used between the
cylindrical member 40 andslide glass 8. When the adhesive 41 is not used, for example, the grease may thinly be applied to the opening of thecylindrical member 40. This may prevent the strippingagent 42 from leaking between thecylindrical member 40 andslide glass 8. - The material of the
cylindrical member 40 is not limited to glass, and other materials such as plastic and metal can also be used. - Moreover, as shown in
FIG. 9 , atransparent film 60 formed by a thin resin can also be attached to one opening of thecylindrical member 40 for use. In this case, a stripping agent/extract agent 61 is injected/recovered, for example, using a dropper, pipette, andsyringe 62. - Next, the
transparent film 60 is stripped, and the stripping agent/extract agent 61 is injected into thecylindrical member 40. - Next, the stripping agent/
extract agent 61 is reacted in a sealed state by thetransparent film 60. - Thereafter, the
transparent film 60 is stripped, and the solution is recovered, for example, by the dropper, pipette, andsyringe 62. - This can prevent the mixture of the foreign particles into the
cylindrical member 40. - It has been described that the first embodiment is effective in the analysis such as the flow cytometry. On the other hand, there are researches such as analysis of nucleic acid (DNA, RNA) and protein. This research does not require the stripping of the cells a from the
slide glass 8. - That is, as shown in
FIG. 2 , thecylindrical member 40 is bonded/fixed onto theslide glass 8. Next, theslide glass 8 is removed from thestage 6. - Next, as shown in
FIG. 5 , instead of the strippingagent 42, a solution for directly extracting the sample to be sampled such as the nucleic acids and protein from the cells a is injected into thecylindrical member 40. Examples of the solution include a commercially available solution such as a nucleic acid extract kit. - When the samples such as the nucleic acid (DNA, RNA) and protein are eluted in the solution, the samples eluted in the solution are sucked with the dropper and transferred into another new container in the same manner as in the first embodiment.
- For example, the nucleic acid (DNA, RNA) and protein sampled in this manner can be used to analyze the nucleic acid (DNA, RNA) and protein.
- Next, a second embodiment of the present invention will be described. It is to be noted that the same components as those of
FIG. 1 are denoted with the same reference numerals, and the detailed description thereof is omitted. -
FIG. 7 is a constitution diagram of the inverted microscope for use in the cell acquiring method of the present invention. TheUV laser head 23 outputs the laser beam having the wavelength of the ultraviolet region. The laser beam outputted from theUV laser head 23 is set to the energy for leaving the cells a to be sampled among a plurality of cells included in thespecimen 7 and damaging properties of the unnecessary specimen portion other than the cells a to be sampled. The damaging of the properties of the unnecessary specimen portion means that the nucleic acid is fractionated to change the basic properties of the nucleic acid or cell. - The
stage 6 includes a measuringunit 50. The measuringunit 50 measures an XY coordinate which moves in the XY direction of thestage 6, and outputs an XY coordinate measuring signal. - The image processing and
stage control apparatus 16 is inputted the XY coordinate measuring signal outputted from the measuringunit 50, and calculates the irradiation position of the laser beam with which thespecimen 7 is irradiated from the XY coordinate measuring signal. - The image processing and
stage control apparatus 16 image-processes the image signal outputted from theCCD camera 15 to acquire the image data of thespecimen 7, and superimposes image data of a colored display portion indicating the irradiation position of the laser beam upon the image data. The colored display portion is preferably, for example, in red. - The image processing and
stage control apparatus 16 superimposes the image data of thespecimen 7 upon the image data of the colored display portion indicating the irradiation position of the laser beam to display the data in thedisplay 17. - Next, the sampling method of the cell a existing in the
specimen 7 will be described. - In the same manner as in the first embodiment, the observer observes the enlarged image of the
specimen 7 displayed in thedisplay 17 or sees through theeyepiece lens 22 to observe the enlarged image of thespecimen 7 to determine each cells a to be sampled from thespecimen 7, for example, shown inFIG. 8 . - Next, each cells a to be sampled in the
specimen 7 are left, and the other cells are irradiated with the laser beam. - The cells detached from the cells a to be sampled are irradiated with the laser beam with the large spot diameter s2.
- Next, the variable slit 25 forms the small spot diameter s3, and irradiates the periphery of the cells a to be sampled carefully, that is, correctly along the periphery of the cells a to leave only the cells a to be sampled.
- In the first embodiment, the laser spot diameter is adjusted by the variable slit 25, but in the second embodiment, the laser energy may only be supplied to fractionate the nucleic acid of the unnecessary cell. The laser energy for fractionating the nucleic acid is smaller than the energy for removing the whole cell. Thereby, instead of the
UV laser head 23, the mercury lamp is used to fractionate the nucleic acid of the unnecessary cell, and the basic properties of the nucleic acid or cell can be damaged. Moreover, the objective lens forlaser 5 can be changed to a lens with a low magnification to enlarge the laser spot diameter. - The objective lens for
laser 5 is changed to the magnification such as about five times. - Moreover, the measuring
unit 50 measures the XY coordinate which moves in the XY direction of thestage 6, and outputs the XY coordinate measuring signal. - The image processing and
stage control apparatus 16 inputs the XY coordinate measuring signal outputted from the measuringunit 50, and calculates the irradiation position of the laser beam with which thespecimen 7 is irradiated from the XY coordinate measuring signal. - The image processing and
stage control apparatus 16 image-processes the image signal outputted from theCCD camera 15 to acquire the image data of thespecimen 7, and superimposes the image data upon the image data of the colored display portion indicating the irradiation position of the laser beam, for example, in red. - The image processing and
stage control apparatus 16 superimposes the image data of thespecimen 7 upon the image data of the colored display portion indicating the irradiation position of the laser beam to display the data in thedisplay 17. - The observer operates the
operation unit 18, confirms the image of thespecimen 7 displayed in thedisplay 17 and the colored display portion indicating the irradiation position of the laser beam, and moves the irradiation position of the laser beam with the spot diameter s2 to the peripheral portion of each cell a every shot. - The
specimen 7 around each cell a is irradiated with the laser beam with the spot diameter s2. It is to be noted thatFIG. 8 does not show all the irradiation positions of the laser beam with the spot diameter s2. - For the portion of the
specimen 7 irradiated with the laser beam with the spot diameter s2, when the nucleic acid is fractionated, the basic properties of the nucleic acid or cell are damaged. - Next, for the variable slit 22 of the UV cut
filter 21, the diameter of the circular opening is set, for example, to the slit width S3 smaller than the cells a as shown inFIG. 8 . Thereby, the spot diameter of the laser beam with which thespecimen 7 is irradiated is s3. - It is to be noted that the objective lens for
laser 5 is changed to a high magnification such as about five times, and an ND filter may also be disposed before the objective lens forlaser 5. - The observer observes the
specimen 7 displayed in thedisplay 17 to determine irradiation routes R3, R4 of the laser beam for leaving each cells a and damaging the properties of theambient specimen 7 disposed adjacent to each cells a. - The observer confirms the image of the
specimen 7 displayed in thedisplay 17 and the colored display portion indicating the irradiation position of the laser beam, and operates theoperation unit 18 so as to dispose the irradiation position of the laser on the irradiation route R3 or R4. - The image processing and
stage control apparatus 16 receives the operation command in accordance with the operation of the movement direction and movement amount of thestage 6 from theoperation unit 18, and sends the movement control signal of thestage 6 to thestage driving unit 9 in accordance with the operation command. - Thereby, the
stage 6 is moved in the XY direction, for example, every shot of laser beam so as to dispose the irradiation position on the irradiation route R1 or R3. - When the laser beam is shot from the
UV laser head 23, for example, every shot, the irradiation route R3 is irradiated with the laser beam passed through the UVimage forming lens 24,dichroic mirror 4, and objective lens forlaser 5, and subsequently the irradiation route R4 is irradiated. It is to be noted that the irradiation route R4 and then the irradiation route R3 may be irradiated with the laser beam. - The energy of the laser beam is set to such an extent that the properties of the
surrounding specimen 7 disposed adjacent to each cell a are damaged. Therefore, for the portions of thespecimen 7 on the respective irradiation routes R3, R4 irradiated with the laser beam, when the nucleic acid is fractionated, the basic properties of the nucleic acid or cell are damaged. - It is to be noted that the XY coordinate positions of the irradiation routes R3, R4 may also be stored in the image processing and
stage control apparatus 16 beforehand. That is, after the irradiation routes R3, R4 of the laser beam are determined, the observer operates theoperation unit 18 to move thestage 6 in the XY direction, and moves the colored display portion indicating the irradiation position of the laser beam along each of the irradiation routes R3, R4. - At this time, the image processing and
stage control apparatus 16 reads the movement direction and amount of thestage 6 from theoperation unit 18, and calculates the XY coordinate positions of thestage 6 in the irradiation routes R3, R4 from the movement direction and amount. The image processing andstage control apparatus 16 stores the calculated XY coordinate positions on the irradiation routes R3, R4 in the internal memory. - The image processing and
stage control apparatus 16 reads the XY coordinate positions for each of the irradiation routes R3, R4 stored in the internal memory, and sends the movement control signal of thestage 6 to thestage driving unit 9 in accordance with the XY coordinate positions. - The
stage 6 automatically moves the irradiation position with the laser beam in the XY direction for each of the irradiation routes R3, R4. Additionally, theUV laser head 23 outputs one shot of laser beam every predetermined period. - Thereby, the portions of the
specimen 7 on the irradiation routes R3, R4 are automatically irradiated with the laser beam, and the basic properties of the nucleic acid or cell are damaged the fractionation of the nucleic acid in the cell. - Next, the
cylindrical member 40 shown inFIG. 2 is prepared in the same manner as in the first embodiment. Thecylindrical member 40 is bonded/fixed onto theslide glass 8. - Next, the solution for extracting the sample such as the nucleic acid (DNA, RNA) and protein from the cells a is injected into the
cylindrical member 40 as shown inFIG. 5 . - When the sample such as the nucleic acid (DNA, RNA) and protein is eluted in the solution, the sample is sucked with the dropper and transferred into new another container.
- On the other hand, when the cells a are stripped from the
slide glass 8, thecylindrical member 40 is bonded/fixed onto theslide glass 8 by the adhesive 41 as shown inFIG. 2 . - Next, the stripping
agent 42 is injected into the container formed bycylindrical member 40 andslide glass 8 as shown inFIG. 5 . The strippingagent 42 promotes the stripping of the cells a from theslide glass 8 as shown inFIG. 6 . - Next, the cells a floating in the stripping
agent 42 is sucked with the dropper together with the strippingagent 42. The cell a sucked with the dropper is transferred into new another container. As a result, the sampling of the cell a ends. - In this manner, according to the second embodiment, the unnecessary specimen portion d other than the cells a to be sampled in the
specimen 7 are irradiated with the laser beam, the nucleic acid of the unnecessary specimen portion d is fractionated, and the basic properties of the nucleic acid or cell are changed. - Thereafter, the solution for extracting the samples such as the nucleic acid (DNA, RNA) and protein from the cells a is injected into the
cylindrical member 40, and the samples such as nucleic acid (DNA, RNA) and protein are eluted. Moreover, the strippingagent 42 is injected into the container formed by thecylindrical member 40 andslide glass 8, and the cells a to be sampled is stripped from theslide glass 8, floated in the strippingagent 42, and sampled. - In this manner, in the second embodiment, the nucleic acid of the cells of the unnecessary specimen portion d is irradiated with the laser beam and fractionated to change the basic properties of the nucleic acid or cell. Therefore, the embodiment is effective mainly in extracting the samples such as the nucleic acid (DNA, RNA) and protein from the
raw specimen 7. - The raw specimen contains moisture. When the
raw specimen 7 is irradiated with the laser beam having a large energy, thespecimen 7 cannot be cut because of the boiled moisture in thespecimen 7. - On the other hand, the energy of the laser beam in the second embodiment is an energy necessary for fractionating the nucleic acid of the unnecessary specimen portion d to damage the basic properties of the nucleic acid or cell. For the energy of the laser beam, as an experiment result, for example, in the fractionation of DNA, an energy of 0.075 mJ is obtained at a wavelength of 266 nm in a square area of 60 μm sides. It is to be noted that even with the energy of the laser beam smaller than 0.075 mJ, the nucleic acid can be fractionated to change the basic properties of the nucleic acid or cell.
- When the properties of the unnecessary specimen portion other than the cells a necessary for the sampling are changed, first the unnecessary specimen portion d around each cell a is irradiated with the laser beam having the large spot diameter s2, and next the unnecessary surrounding specimen portion d disposed adjacent to each cell a is irradiated with the laser beam having the small spot diameter s3. This can reduce the number of laser shots with which the unnecessary specimen portion d is irradiated. The properties of the unnecessary specimen portion d in a broad region can be changed in a short time.
- When the unnecessary specimen portion d is flied from the
slide glass 8 as in the first embodiment, in general, the objective lens forlaser 5 with a magnification of about 50 times is used. On the other hand, in the second embodiment the objective lens forlaser 5 with a magnification of about five times may also be used. Thereby, in the second embodiment, as compared with the use of the objective lens forlaser 5 with the magnification of 50 times, the spot region of the laser beam can be enlarged 100 times. - Since the properties of the cells of the unnecessary specimen portion d are damaged, the broken pieces of the
specimen 7 irradiated with the laser beam are not flied/scattered. Thereby, theair blow 30 andair suction unit 31 are unnecessary. - In the
display 17, the image data of the colored display portion, for example, in red indicating the irradiation position of the laser beam is superimposed and displayed upon the image data of thespecimen 7. The observer can confirm the image of thedisplay 17 while thespecimen 7 is irradiated with the laser beam. Thereby, the periphery disposed adjacent to the cells a to be sampled and other unnecessary specimen portion d can securely be changed in properties. - In the second embodiment, the energy of the laser beam is set to such an extent that the properties of the cells are damaged. As a light source, for example, the mercury lamp or Xenon lamp may also be used. Even when the cell is irradiated with the light emitted from the mercury lamp or Xenon lamp, it is possible to fractionate the nucleic acid and to change the basic properties of the nucleic acid or cell.
- Additional advantages and modifications will readily occur to those skilled in the art. Therefore, the invention in its broader aspects is not limited to the specific details and representative embodiments shown and described herein. Accordingly, various modifications may be made without departing from the spirit or scope of the general invention concept as defined by the appended claims and their equivalents.
Claims (25)
1. A cell isolating method comprising:
irradiating a specimen which is laid on a substrate and which includes cells with an ultraviolet light to change properties of an unnecessary specimen portion other than a cell portion;
using a cylindrical member including openings in opposite ends, and inserting the cell portion into one opening in the cylindrical member to dispose the cylindrical member onto the substrate;
injecting a stripping agent into a container formed by the substrate and cylindrical member to strip the cell portion from the substrate; and
sampling the cell portion which floats in the stripping agent.
2. A method of isolating cells or samples to be analyzed in the cells, comprising:
irradiating a specimen which is laid on a substrate and which includes cells with an ultraviolet light to change properties of an unnecessary specimen portion other than a cell portion;
using a cylindrical member including openings in opposite ends, and inserting the cell portion into one opening in the cylindrical member to dispose the cylindrical member onto the substrate;
injecting a solution for eluting the sample existing in the cells into a container formed by the substrate and cylindrical member; and
sampling the sample eluted in the solution.
3. The cell isolating method according to claim 1 , wherein the stripping agent comprises an organic solvent.
4. The cell isolating method according to claim 3 , wherein the organic solvent comprises xylene.
5. The cell isolating method according to claim 1 , wherein the sampling of the cells floating in the stripping agent comprises:
sucking the cells in the stripping agent with one of a dropper, pipette, and syringe; and
transferring the cells in the one of the dropper, pipette, and syringe to a container other than the container formed by the substrate and cylindrical member.
6. The cell isolating method according to claim 1 , wherein the ultraviolet light is a laser beam having a wavelength of an ultraviolet region.
7. The cell isolating method according to claim 1 , wherein the cylindrical member is formed of one of glass, metal, and plastic.
8. The cell isolating method according to claim 1 , wherein the cylindrical member is formed by attaching a thin film of resin to one opening.
9. The cell isolating method according to claim 1 , further comprising:
disposing the cylindrical member on the substrate in a liquid-tight manner.
10. The cell isolating method according to claim 1 , further comprising:
fixing the cylindrical member onto the substrate by an adhesive.
11. The cell isolating method according to claim 1 , further comprising:
sealing a gap between the cylindrical member and the substrate by grease applied onto the substrate.
12. The cell isolating method according to claim 1 , further comprising:
applying a fluorescence dyestuff to the specimen to dye the cells;
irradiating the specimen with ultraviolet light to emit a light of the fluorescence dyestuff of the dyed cells; and
observing the cells having emitted the light to determine the cells to be sampled.
13. The cell isolating method according to claim 1 , further comprising:
disposing a microscope which enlarges an image of the specimen so that an optical axis of the microscope agrees with an optical axis of the ultraviolet light;
outputting a marking light from a marking light source on the same axis as the optical axis of the ultraviolet light;
picking up the specimen image enlarged by the microscope and the marking light with which the specimen is irradiated by an image pickup apparatus to display the specimen image and the marking light on a display apparatus; and
observing the specimen image and the marking light displayed on the display apparatus to irradiate the specimen with the ultraviolet light, changing the properties of the unnecessary specimen portion other than the cell portion to be sampled.
14. The cell isolating method according to claim 1 , wherein an irradiation diameter of the ultraviolet light is changeable, and when the unnecessary specimen portion is changed in properties, the method further comprises:
setting the irradiation diameter of the ultraviolet light for irradiating the unnecessary specimen portion disposed adjacent to the cell portion to a first diameter smaller than a size of the cell portion; and
setting the irradiation diameter of the ultraviolet light for irradiating the unnecessary specimen portion disposed apart from the cell portion by a predetermined distance to a second diameter larger than the first diameter.
15. The cell isolating method according to claim 2 , wherein the sampling of the sample eluted in the solution comprises:
sucking the sample in the solution with one of a dropper, pipette, and syringe; and
transferring the cells in the one of the dropper, pipette, and syringe to a container other than the container formed by the substrate and cylindrical member.
16. The cell isolating method according to claim 2 , further comprising:
extracting the sample including at least one of DNA, RNA, and protein from the cells by the solution.
17. The cell isolating method according to claim 2 , further comprising:
setting a wavelength of the laser beam to an ultraviolet region.
18. The cell isolating method according to claim 2 , wherein the cylindrical member is formed of one of glass, metal, and plastic.
19. The cell isolating method according to claim 2 , wherein the cylindrical member is formed by attaching a thin film of resin to one opening.
20. The cell isolating method according to claim 2 , further comprising:
disposing the cylindrical member on the substrate in a liquid-tight manner.
21. The cell isolating method according to claim 2 , further comprising:
fixing the cylindrical member onto the substrate by an adhesive.
22. The cell isolating method according to claim 2 , further comprising:
sealing a gap between the cylindrical member and the substrate by grease applied onto the substrate.
23. The cell isolating method according to claim 2 , further comprising:
applying a fluorescence dyestuff to the specimen to dye the cells;
irradiating the specimen with ultraviolet light to emit a light of the fluorescence dyestuff of the dyed cells; and
observing the cells having emitted the light to determine the cells to be sampled.
24. The cell isolating method according to claim 2 , further comprising:
disposing a microscope which enlarges an image of the specimen so that an optical axis of the microscope agrees with an optical axis of the ultraviolet light;
outputting a marking light from a marking light source on the same axis as the optical axis of the ultraviolet light;
picking up the specimen image enlarged by the microscope and the marking light with which the specimen is irradiated by an image pickup apparatus to display the specimen image and the marking light on a display apparatus; and
observing the specimen image and the marking light displayed on the display apparatus to irradiate the specimen with the ultraviolet light, changing the properties of the unnecessary specimen portion other than the cell portion to be sampled.
25. The cell isolating method according to claim 2 , wherein an irradiation diameter of the ultraviolet light is changeable, and when the unnecessary specimen portion is changed in properties, the method further comprises:
setting the irradiation diameter of the ultraviolet light for irradiating the unnecessary specimen portion disposed adjacent to the cell portion to a first diameter smaller than a size of the cell portion; and
setting the irradiation diameter of the ultraviolet light for irradiating the unnecessary specimen portion disposed apart from the cell portion by a predetermined distance to a second diameter larger than the first diameter.
Priority Applications (1)
Application Number | Priority Date | Filing Date | Title |
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US11/834,419 US20080176307A1 (en) | 2002-01-23 | 2007-08-06 | Method of isolating cell or sample to be analyzed in cell |
Applications Claiming Priority (4)
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JP2002014643 | 2002-01-23 | ||
JP2002-014643 | 2002-01-23 | ||
US10/348,198 US7264966B2 (en) | 2002-01-23 | 2003-01-21 | Method of isolating cell or sample to be analyzed in cell |
US11/834,419 US20080176307A1 (en) | 2002-01-23 | 2007-08-06 | Method of isolating cell or sample to be analyzed in cell |
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US10/348,198 Division US7264966B2 (en) | 2002-01-23 | 2003-01-21 | Method of isolating cell or sample to be analyzed in cell |
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US20080176307A1 true US20080176307A1 (en) | 2008-07-24 |
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US10/348,198 Expired - Lifetime US7264966B2 (en) | 2002-01-23 | 2003-01-21 | Method of isolating cell or sample to be analyzed in cell |
US11/834,419 Abandoned US20080176307A1 (en) | 2002-01-23 | 2007-08-06 | Method of isolating cell or sample to be analyzed in cell |
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US10/348,198 Expired - Lifetime US7264966B2 (en) | 2002-01-23 | 2003-01-21 | Method of isolating cell or sample to be analyzed in cell |
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US7264966B2 (en) * | 2002-01-23 | 2007-09-04 | Olympus Optical Co., Ltd. | Method of isolating cell or sample to be analyzed in cell |
JP6291972B2 (en) | 2014-03-31 | 2018-03-14 | 三菱マテリアル株式会社 | Sampling position display device and sampling method |
JP6541160B2 (en) | 2017-02-13 | 2019-07-10 | 株式会社片岡製作所 | Cell processing apparatus and method for processing object |
JP6532063B2 (en) | 2017-02-13 | 2019-06-19 | 株式会社片岡製作所 | Cell processing equipment |
JP6574028B1 (en) | 2018-06-29 | 2019-09-11 | 株式会社片岡製作所 | Cell processing apparatus and cell laser processing method |
Citations (12)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5037754A (en) * | 1985-10-19 | 1991-08-06 | Daikin Industries Ltd. | Culture vessel |
US5585116A (en) * | 1989-11-22 | 1996-12-17 | Ost-Developpment | Method of manufacture of a material for osteoplasty from a natural bone tissue and material obtained thereby |
US5859699A (en) * | 1997-02-07 | 1999-01-12 | Arcturus Engineering, Inc. | Laser capture microdissection analysis vessel |
US5998129A (en) * | 1996-02-05 | 1999-12-07 | P.A.L.M. Gmbh | Method and device for the contactless laser-assisted microinjection, sorting and production of biological objects generated in a planar manner |
US6040139A (en) * | 1995-09-19 | 2000-03-21 | Bova; G. Steven | Laser cell purification system |
US6100051A (en) * | 1997-06-27 | 2000-08-08 | The United States Of America As Represented By The Department Of Health And Human Services | Method utilizing convex geometry for laser capture microdissection |
US6194157B1 (en) * | 1997-02-03 | 2001-02-27 | Hamamatsu Photonics K.K. | Method for separating biological substances by using photoresist |
US20020048747A1 (en) * | 2000-09-01 | 2002-04-25 | Michael Ganser | Method and apparatus for laser microdissection |
US20020056345A1 (en) * | 2000-09-01 | 2002-05-16 | Michael Ganser | Method and apparatus for laser microdissection |
US6455309B2 (en) * | 1999-02-11 | 2002-09-24 | Crosscart, Inc. | Proteoglycan-reduced soft tissue xenografts |
US20020146816A1 (en) * | 2000-10-25 | 2002-10-10 | Vellinger John C. | Bioreactor apparatus and cell culturing system |
US7264966B2 (en) * | 2002-01-23 | 2007-09-04 | Olympus Optical Co., Ltd. | Method of isolating cell or sample to be analyzed in cell |
Family Cites Families (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US48747A (en) * | 1865-07-11 | Improved solution for saturating natural and artificial stone | ||
US56345A (en) * | 1866-07-17 | Improvement in extension corset-springs | ||
JP2000266649A (en) | 1999-03-15 | 2000-09-29 | Olympus Optical Co Ltd | Sampling cap |
-
2003
- 2003-01-21 US US10/348,198 patent/US7264966B2/en not_active Expired - Lifetime
-
2007
- 2007-08-06 US US11/834,419 patent/US20080176307A1/en not_active Abandoned
Patent Citations (13)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5037754A (en) * | 1985-10-19 | 1991-08-06 | Daikin Industries Ltd. | Culture vessel |
US5585116A (en) * | 1989-11-22 | 1996-12-17 | Ost-Developpment | Method of manufacture of a material for osteoplasty from a natural bone tissue and material obtained thereby |
US6316234B1 (en) * | 1995-09-19 | 2001-11-13 | G. Steven Bova | Laser cell purification system |
US6040139A (en) * | 1995-09-19 | 2000-03-21 | Bova; G. Steven | Laser cell purification system |
US5998129A (en) * | 1996-02-05 | 1999-12-07 | P.A.L.M. Gmbh | Method and device for the contactless laser-assisted microinjection, sorting and production of biological objects generated in a planar manner |
US6194157B1 (en) * | 1997-02-03 | 2001-02-27 | Hamamatsu Photonics K.K. | Method for separating biological substances by using photoresist |
US5859699A (en) * | 1997-02-07 | 1999-01-12 | Arcturus Engineering, Inc. | Laser capture microdissection analysis vessel |
US6100051A (en) * | 1997-06-27 | 2000-08-08 | The United States Of America As Represented By The Department Of Health And Human Services | Method utilizing convex geometry for laser capture microdissection |
US6455309B2 (en) * | 1999-02-11 | 2002-09-24 | Crosscart, Inc. | Proteoglycan-reduced soft tissue xenografts |
US20020048747A1 (en) * | 2000-09-01 | 2002-04-25 | Michael Ganser | Method and apparatus for laser microdissection |
US20020056345A1 (en) * | 2000-09-01 | 2002-05-16 | Michael Ganser | Method and apparatus for laser microdissection |
US20020146816A1 (en) * | 2000-10-25 | 2002-10-10 | Vellinger John C. | Bioreactor apparatus and cell culturing system |
US7264966B2 (en) * | 2002-01-23 | 2007-09-04 | Olympus Optical Co., Ltd. | Method of isolating cell or sample to be analyzed in cell |
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US7264966B2 (en) | 2007-09-04 |
US20040043392A1 (en) | 2004-03-04 |
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