CN1445523A - Method for preparing tissue chip - Google Patents
Method for preparing tissue chip Download PDFInfo
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- CN1445523A CN1445523A CN 03113771 CN03113771A CN1445523A CN 1445523 A CN1445523 A CN 1445523A CN 03113771 CN03113771 CN 03113771 CN 03113771 A CN03113771 A CN 03113771A CN 1445523 A CN1445523 A CN 1445523A
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Abstract
A process for preparing paraffin wax embedded tissue microarray chip with oridinary microscope includes such steps as installing special perforating needle and sampling needle to the lens holder of the ordinary optical microscope, putting special paraffin wax block fixing box on object carrier, turning the lens holder and regulating the distance between lens and object carrier while perforating, sample and checking sample, and observing while locating. Its advantages are simple operation and low cost.
Description
Affiliated technical field
The present invention relates to a kind of method for making of biochip, further relate to a kind of method for making of paraffin embedded tissue chip.
Technical background
Organization chip (tissue microarray, TMA, claim " micro-array tissue " again) be a kind of important biochip technology that after genetic chip, protein-chip, occurs, because it can detect a large amount of tissue specimens simultaneously, the science and the comparability of testing result are strong, contain much information, the efficient height, consumption reagent is few, has broad application prospects at genome times afterwards comprehensively gene and protein research, molecular pathology research, drug screening, health and aspects such as environmental monitoring and national defense and military.
Making the paraffin-embedded tissue array is one of gordian technique of organization chip.Battifora and Wan etc. once reported the manual assembled arrangement at random of the tissue specimen of the dehydration that will dewax, embedding again, conventional section (1.Battifora et al.Lab Invest 1986,55:244-248; 2.Wan et al, J Immol Meth, 1987,103:121-129).The tissue specimen limited amount that the method is placed varies in size, and spacing differs, arrangement disorder, and out-of-shape is difficult to carry out automated analysis.(Kononen Jet al.Nature Med such as Kononen, 1998,4 (7): 844-847) and (Chinese invention patent application number: 01128783.7) propose to utilize special instrument such as Li Jun, on paraffin mass, punch earlier, obtain with special sampling probe again and organize microlith, it is implanted in the paraffin mass that has punched, then section.This method needs special instrument, the expense costliness (Liu Lianxin. foreign medical oncology credit volume, 2000,27:83-85), and can not utilize microscope to carry out accurate sampling point observation and location simultaneously, the installation of its perforating needle and sampling probe, adjusting and replacing inconvenience, the whole operation process is more loaded down with trivial details, has limited this broad application.Propositions such as Hu Guo utilize the semi-molten state of paraffin to make organization chip (Chinese invention patent application number: 02113230.5).This method need be kept paraffin semi-molten state for a long time, is not easy to use microscope equally and carries out sampling point observation and location in sampling, point sample process, is difficult to make high density, high-precision organization chip.
Summary of the invention
The purpose of this invention is to provide a kind of both easy, science, technology acuracy height, tissue chip production method easy to utilize again.This method is that special perforating needle and sampling probe are installed on the ordinary optical microscope lens mount, on microscope stage, settle special paraffin mass fixed bin, utilize simple microscope to carry out the accurate sampling and the point sample of paraffin-embedded tissue, thereby make high-quality organization chip.
The technological merit of this method is:
(1) described tissue chip production method, 03113734.2) and a kind of special paraffin mass fixed bin (Chinese invention patent application number: 03113733.4) that is installed on the microscope stage employing is installed in special perforating needle on the microscope lens headstock and sampling probe (Chinese invention patent application number:, utilize moving horizontally of vertical moving between microscope camera lens and the objective table and microslide device to carry out sampling, the point sample of punching of acceptor paraffin mass and paraffin-embedded tissue, its method is easy, technology is reliable, expense is cheap, promotes easily.
(2) described tissue chip production method, its punching and sampling process do not need the operator directly firmly to push perforating needle and sampling probe, but take to regulate the method for the adjustable screw of microscope stage or camera lens, make perforating needle and sampling probe thrust and withdraw from paraffin mass.It is firmly slight, and perforating needle and sampling probe are difficult for generation slip, shake, and punching and sampling depth control are easy, the precision height.
(3) described tissue chip production method, its paraffin mass fixed bin is installed on the microscope stage, utilizes paraffin mass fixed bin " U " shape bayonet socket and respective fixation screw rod, setting nut to install and fix, and realizes and the interlock of microslide device, its degree of regulation height, easy to loading and unloading.
(4) described tissue chip production method utilizes the rotation of microscope lens headstock to carry out replacing of perforating needle, sampling probe and camera lens working position.Perforating needle and sampling probe bearing accuracy and collimation height, it is convenient to change, and can also take disposable perforating needle and sampling probe, further improves the precision of punching and sampling.
(5) described tissue chip production method can be taken a sample and observation of point sample position and location accurately by microscope; After microscope fixing body visor head and the illumination, can under the stereoscope direct-view, take a sample and point sample.This method positional accuracy height can save to conventional organization biopsy marker sampling point and with the donor paraffin mass and carry out the operating process of contraposition.
(6) described tissue chip production method, the replacement of its punching, location, sampling and point sample operating process, do not need to change back and forth operator's console, do not need to add other microscopes, only needing rotation to change the microscope lens headstock can finish with the horizontal level of regulating the paraffin mass fixed bin, can carry out on the same plane of same instrument, easy and simple to handle, accurate.
Description of drawings
Below in conjunction with accompanying drawing the preferred embodiment of the present invention is described.
Fig. 1 is the repacking of ordinary optical microscope according to the preferred embodiment of the invention synoptic diagram.
Fig. 2 for punch according to the preferred embodiment of the invention, locate, sampling and point sample process synoptic diagram.
Fig. 3 is for determining the synoptic diagram at sample of tissue position according to the preferred embodiment of the present invention.
Fig. 4 is the paraffin-embedded tissue dot chart of making according to the preferred embodiment of the present invention.
Fig. 5 is the organization chip picture of making according to the preferred embodiment of the present invention (HE dyeing) in kind
Fig. 6 is the amplification picture (amplifying 10 times) of Fig. 5 institute favored area
Among the figure, 1. microscope camera lens, 2. sampling probe, 3. perforating needle, 4. paraffin mass fixed bin, 5. paraffin mass standing screw and nut thereof, the 6. standing screw of paraffin mass fixed bin and nut, 7. paraffin mass fixed bin " U " shape bayonet socket, 8. microscope stage, 9. microslide device, 10. paraffin mass, 11. control objective tables or camera lens move up and down the adjusting screw(rod) and the nut of scope, 12. donor paraffin mass and investing tissue thereof (stereoscope finding), 13. section of the routine of donor paraffin mass and investing tissue thereof and HE dyeing (stereoscope finding), matter between 14. tissues (few cell) zone or inorganization zone, 15. sampling points.
Embodiment
Needed equipment of the tissue chip production method of preferred embodiment and article comprise: the logical optical microscope of a Daepori is also suitably reequiped; One cover can be installed in special perforating needle, sampling probe and associated component thereof (the Chinese invention patent application number: 03113734.2) on microscope camera lens or the lens mount; Special paraffin mass fixed bin (the Chinese invention patent application number: 03113733.4) that can be installed on the microscope stage; Control objective table or camera lens move up and down the adjusting screw(rod) and the nut of scope and control the microslide device in length and breadth to the fixedly card that moves; Donor paraffin mass and paraffin-embedded tissue; The acceptor paraffin mass; The baking box of controllable temperature; Apparatus, equipment and reagent that conventional organization section, dyeing and immunohistochemistry are relevant.
Fig. 1 is the repacking of simple microscope according to the preferred embodiment of the invention synoptic diagram.Its method of modifying comprises:
1. special sampling probe 2 and perforating needle 3 are installed on microscope camera lens 1 or lens mount;
2. special paraffin mass fixed bin 4 is installed on microscope stage 8;
3. adjusting screw(rod) and the nut 11 that control objective table or camera lens move up and down scope is installed on microscope column;
4. control microslide device fixedly card vertical or that laterally move at microscope stage one side along installing.
Fig. 2 (A, B, C, D) for punch according to the preferred embodiment of the invention, locate, sampling and point sample process synoptic diagram.The operating process of its preferred embodiment comprises:
A. punching.Its method is, acceptor paraffin mass 10 is fixed in special paraffin mass fixed bin 4 one sides, rotation perforating needle 3 is to working position, regulate objective table or camera lens and move up and down scope, change the vertical range of acceptor paraffin mass 10 and perforating needle 3, make perforating needle thrust the position that acceptor paraffin mass 10 is selected, the withdraw of the needle then, push the perforating needle nook closing member, extrude the paraffin in the perforating needle, finish the punch operation of position, a hole.
B. sampling point is located.Its method is, the donor paraffin mass is placed special paraffin mass fixed bin 4 opposite sides, and sampling point is observed and selected to rotation microscope camera lens 1 to working position, make the tissue that to take a sample be positioned at centre under the mirror, then anchor stone wax stone fixed bin 4 and fixing donor paraffin mass; Perhaps under stereoscope, select sampling point, and make sampling point be centered close to sampling probe needle point relative position.
C. sampling.Its method is, rotation sampling probe 2 is to working position, regulate microscope stage 8 or camera lens and move up and down scope, change the vertical range of donor paraffin mass and sampling probe 2, the selected position that makes perforating needle thrust donor paraffin mass 10, and manual suitably rotation donor paraffin mass, the withdraw of the needle then, as seen sampling point stays next thin cylindrical cavities, shows that the tissue particles of getting is positioned at sampling probe.
D. point sample.Its method is, regulates mobile paraffin mass fixed bin 4, makes sampling probe 2 aim at acceptor paraffin mass 10 relevant positions of having punched, and pushes the sampling probe nook closing member with suitable strength and speed, and the tissue particles that cuts in the sampling probe is vertically implanted acceptor paraffin mass corresponding hole site.
In the punching of preferred embodiment, sampling, point sample operating process, for the density that improves tissue array with arrange precision, should select thin, sharp and supporting perforating needle and the sampling probe of needle point for use, can select disposable perforating needle and sampling probe for use; To before the acceptor paraffin mass punching, available high-precision angle square and fine needle are drawn the system array grid earlier on paraffin mass, so that the collimation of check and correction dot matrix arrangement when punching and point sample; Simultaneously can utilize the fixedly card that is installed in the microscope stage edge, locking microslide device avoids paraffin mass arbitrarily laterally or vertically move; In the withdraw of the needle process of punching and sampling, if do not strengthen the anchor stone wax stone in vertical direction, the operator can live paraffin mass fixed bin or paraffin mass itself with a hand, teetertotters to avoid paraffin mass.
Fig. 3 is for determining the synoptic diagram at sample of tissue position according to the preferred embodiment of the present invention.Its method comprises: 1. take off the paraffin mass fixed bin, the routine of donor paraffin organization is cut into slices places microscopically, and rotation microscope camera lens 1 is observed and selected sampling point to working position; 2. utilize external light source direct irradiation donor paraffin mass, by the tissue in the wide-angle magnifier head Direct observation donor paraffin mass, selected sampling point; 3. utilize stereoscope camera lens and extraneous illumination, under the stereoscope direct-view, determine sampling point.These methods can improve the correctness of sampling, reduce blindness, thereby improve the quality of organization chip.When selecting sampling point,, determine the sampling tissue site according to different research and testing goal.Generally choose diseased region, contain the more position of cell, contain particular organization's eucaryotic cell structure and form position, the intersection of the blister cavities of cystic lesion and cyst wall intersection, normal and abnormality etc., the position of avoiding inorganization and necrosis taking place fully, normal control should be chosen the normal structure position.
Preferred embodiment is paraffin-embedded glands of large intestine tumor tissue 12, and the purpose of making chip is the research adenoma cell, when determining sampling point, should avoid the zone 14 of choosing no adenoma cell or not having the adenoma pathology.Its step is, under ordinary optical perspective microscope or stereoscope, observe routine section (the HE dyeing) 12 of this tissue, the selected sampling point 15 of intending, seek the corresponding site of donor paraffin mass 12 then, make it place central field of vision position under the mirror, fixing donor paraffin mass, rotate sampling probe again to working position, take a sample.
After finishing the punching and point sample of acceptor paraffin mass, from the paraffin mass fixed bin, take out the acceptor paraffin mass, again the acceptor paraffin mass heated to 58 ℃-65 ℃, make it suitably softening, and with slide slightly by flattening the interlacing point battle array; Make the acceptor paraffin mass freezing then, cut into slices again, dye or carry out immunohistochemistry, in situ hybridization, original position PCR equimolecular pathology and detect.
Fig. 4 is the paraffin-embedded tissue dot chart of making according to the preferred embodiment of the present invention.The paraffin mass size of preferred embodiment is 24mm * 35mm, and its dot matrix quantity is 111 (8 * 14-1) points.Single dot matrix diameter 1.5mm, the spacing 0.5mm of organizing.The arrangement mode of paraffin-embedded tissue dot matrix can carry out combination in any according to the research needs and arrange.
Fig. 5 is the organization chip pictorial diagram (HE dyeing) of making according to the preferred embodiment of the present invention.The blank dot matrix in lower left corner is a witness marker among the preferred embodiment figure.
Fig. 6 is that the stereoscope of Fig. 5 institute favored area amplifies picture (amplifying 8 times).The dot matrix marshalling of preferred embodiment among the figure, the clear-cut margin of sampling tissue does not have obviously tissue extruding and malformation, and institute gets and organizes dot matrix is the interest groups tissue region that meets the research requirement.
Claims (5)
1. tissue chip production method is characterized in that:
Special perforating needle and sampling probe are installed on ordinary optical microscope or stereoscope lens mount;
On microscope stage, settle special paraffin mass fixed bin;
Rotation microscope lens headstock carries out replacing of perforating needle, sampling probe and microscope camera lens working position;
By regulating the adjustable screw of microscope stage or camera lens, finish punching, sampling and point sample operation;
Take a sample and the observation and the location of point sample by same microscope or stereoscope.
2. tissue chip production method as claimed in claim 1, it is characterized in that, take to regulate the method for microscope stage or camera lens up-down adjustment spiral, make perforating needle or sampling probe thrust and withdraw from paraffin mass, its punching and sampling depth are subjected to the accurate adjusting and the control of microscope column adjustable screw.
3. tissue chip production method as claimed in claim 1, it is characterized in that, utilize the rotation of microscope lens headstock to carry out replacing of perforating needle, sampling probe and microscope camera lens working position, utilize the position of accurate adjusting donor of moving horizontally of objective table and acceptor paraffin mass.
4. tissue chip production method as claimed in claim 1 is characterized in that, described tissue chip production method can by same microscope or stereoscope be taken a sample accurately and the point sample position is observed and the location.
5. tissue chip production method as claimed in claim 1 is characterized in that, its punching, location, sampling and point sample operation and substitute are carried out at the same working face of same microscope or stereoscope, and be easy and simple to handle.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 03113771 CN1445523A (en) | 2003-02-20 | 2003-02-20 | Method for preparing tissue chip |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN 03113771 CN1445523A (en) | 2003-02-20 | 2003-02-20 | Method for preparing tissue chip |
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| CN1445523A true CN1445523A (en) | 2003-10-01 |
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Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101319971B (en) * | 2008-07-01 | 2010-12-01 | 南通大学 | Paraffin tissue chip preparation method |
| CN101187604B (en) * | 2007-12-14 | 2011-06-15 | 大连医科大学 | Freezing and paraffin embedded tissue chip production device |
| CN102157096A (en) * | 2011-03-14 | 2011-08-17 | 毛静涛 | Teaching (biological) chip |
| CN101294951B (en) * | 2007-04-28 | 2012-09-12 | 漆楚波 | Tissue chip production method |
| CN102798562A (en) * | 2012-08-02 | 2012-11-28 | 王虎 | Paraffin texture chip preparation method |
| CN104497595A (en) * | 2015-01-13 | 2015-04-08 | 中国科学院生物物理研究所 | Composite receptor paraffin for tissue chip and preparation process thereof |
| CN110132702A (en) * | 2019-06-11 | 2019-08-16 | 河南科技学院 | An embedding method for specific pathological tissues |
-
2003
- 2003-02-20 CN CN 03113771 patent/CN1445523A/en active Pending
Cited By (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101294951B (en) * | 2007-04-28 | 2012-09-12 | 漆楚波 | Tissue chip production method |
| CN101187604B (en) * | 2007-12-14 | 2011-06-15 | 大连医科大学 | Freezing and paraffin embedded tissue chip production device |
| CN101319971B (en) * | 2008-07-01 | 2010-12-01 | 南通大学 | Paraffin tissue chip preparation method |
| CN102157096A (en) * | 2011-03-14 | 2011-08-17 | 毛静涛 | Teaching (biological) chip |
| CN102798562A (en) * | 2012-08-02 | 2012-11-28 | 王虎 | Paraffin texture chip preparation method |
| CN104497595A (en) * | 2015-01-13 | 2015-04-08 | 中国科学院生物物理研究所 | Composite receptor paraffin for tissue chip and preparation process thereof |
| CN110132702A (en) * | 2019-06-11 | 2019-08-16 | 河南科技学院 | An embedding method for specific pathological tissues |
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