CN105938064B - The group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry - Google Patents
The group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry Download PDFInfo
- Publication number
- CN105938064B CN105938064B CN201610404344.8A CN201610404344A CN105938064B CN 105938064 B CN105938064 B CN 105938064B CN 201610404344 A CN201610404344 A CN 201610404344A CN 105938064 B CN105938064 B CN 105938064B
- Authority
- CN
- China
- Prior art keywords
- pen
- cell
- glass slide
- ware
- culture dish
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 230000009194 climbing Effects 0.000 title claims abstract description 24
- 238000003364 immunohistochemistry Methods 0.000 title claims abstract description 19
- 239000003153 chemical reaction reagent Substances 0.000 title claims abstract description 11
- 238000001514 detection method Methods 0.000 title claims abstract description 6
- 239000011521 glass Substances 0.000 claims abstract description 21
- 238000004043 dyeing Methods 0.000 claims abstract description 14
- 230000001744 histochemical effect Effects 0.000 claims abstract description 7
- 239000007788 liquid Substances 0.000 claims abstract description 7
- 238000004113 cell culture Methods 0.000 claims abstract description 5
- 239000011148 porous material Substances 0.000 claims abstract description 3
- QGJOPFRUJISHPQ-UHFFFAOYSA-N Carbon disulfide Chemical compound S=C=S QGJOPFRUJISHPQ-UHFFFAOYSA-N 0.000 claims description 21
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 15
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 claims description 14
- VZGDMQKNWNREIO-UHFFFAOYSA-N tetrachloromethane Chemical compound ClC(Cl)(Cl)Cl VZGDMQKNWNREIO-UHFFFAOYSA-N 0.000 claims description 10
- 230000000694 effects Effects 0.000 claims description 8
- 239000012188 paraffin wax Substances 0.000 claims description 8
- 238000013115 immunohistochemical detection Methods 0.000 claims description 7
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 claims description 6
- JHIVVAPYMSGYDF-UHFFFAOYSA-N cyclohexanone Chemical compound O=C1CCCCC1 JHIVVAPYMSGYDF-UHFFFAOYSA-N 0.000 claims description 6
- RSWGJHLUYNHPMX-UHFFFAOYSA-N Abietic-Saeure Natural products C12CCC(C(C)C)=CC2=CCC2C1(C)CCCC2(C)C(O)=O RSWGJHLUYNHPMX-UHFFFAOYSA-N 0.000 claims description 5
- ZAFNJMIOTHYJRJ-UHFFFAOYSA-N Diisopropyl ether Chemical compound CC(C)OC(C)C ZAFNJMIOTHYJRJ-UHFFFAOYSA-N 0.000 claims description 5
- KHPCPRHQVVSZAH-HUOMCSJISA-N Rosin Natural products O(C/C=C/c1ccccc1)[C@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 KHPCPRHQVVSZAH-HUOMCSJISA-N 0.000 claims description 5
- 235000013871 bee wax Nutrition 0.000 claims description 5
- 239000012166 beeswax Substances 0.000 claims description 5
- KHPCPRHQVVSZAH-UHFFFAOYSA-N trans-cinnamyl beta-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OCC=CC1=CC=CC=C1 KHPCPRHQVVSZAH-UHFFFAOYSA-N 0.000 claims description 5
- WZUVPPKBWHMQCE-XJKSGUPXSA-N (+)-haematoxylin Chemical compound C12=CC(O)=C(O)C=C2C[C@]2(O)[C@H]1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-XJKSGUPXSA-N 0.000 claims description 4
- WZUVPPKBWHMQCE-UHFFFAOYSA-N Haematoxylin Natural products C12=CC(O)=C(O)C=C2CC2(O)C1C1=CC=C(O)C(O)=C1OC2 WZUVPPKBWHMQCE-UHFFFAOYSA-N 0.000 claims description 4
- 238000004140 cleaning Methods 0.000 claims description 4
- 108090000790 Enzymes Proteins 0.000 claims description 3
- 102000004190 Enzymes Human genes 0.000 claims description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 3
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 3
- 229930040373 Paraformaldehyde Natural products 0.000 claims description 3
- 229920004890 Triton X-100 Polymers 0.000 claims description 3
- 239000013504 Triton X-100 Substances 0.000 claims description 3
- 239000012122 aqueous mounting media Substances 0.000 claims description 3
- 238000004821 distillation Methods 0.000 claims description 3
- 239000004615 ingredient Substances 0.000 claims description 3
- 239000007791 liquid phase Substances 0.000 claims description 3
- 229920002866 paraformaldehyde Polymers 0.000 claims description 3
- 238000000926 separation method Methods 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 3
- 239000012224 working solution Substances 0.000 claims description 3
- 239000000203 mixture Substances 0.000 claims description 2
- 239000011435 rock Substances 0.000 claims 1
- 238000011532 immunohistochemical staining Methods 0.000 abstract description 7
- 238000011081 inoculation Methods 0.000 abstract description 2
- 239000004793 Polystyrene Substances 0.000 description 3
- 238000010586 diagram Methods 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 229920002223 polystyrene Polymers 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 239000001293 FEMA 3089 Substances 0.000 description 2
- 241000283973 Oryctolagus cuniculus Species 0.000 description 2
- 235000021355 Stearic acid Nutrition 0.000 description 2
- RWGFKTVRMDUZSP-UHFFFAOYSA-N cumene Chemical group CC(C)C1=CC=CC=C1 RWGFKTVRMDUZSP-UHFFFAOYSA-N 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- RTZKZFJDLAIYFH-UHFFFAOYSA-N ether Substances CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 2
- 238000003365 immunocytochemistry Methods 0.000 description 2
- 239000003350 kerosene Substances 0.000 description 2
- 150000002576 ketones Chemical class 0.000 description 2
- QIQXTHQIDYTFRH-UHFFFAOYSA-N octadecanoic acid Chemical compound CCCCCCCCCCCCCCCCCC(O)=O QIQXTHQIDYTFRH-UHFFFAOYSA-N 0.000 description 2
- OQCDKBAXFALNLD-UHFFFAOYSA-N octadecanoic acid Natural products CCCCCCCC(C)CCCCCCCCC(O)=O OQCDKBAXFALNLD-UHFFFAOYSA-N 0.000 description 2
- 238000012545 processing Methods 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 239000008117 stearic acid Substances 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 1
- 239000000470 constituent Substances 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 238000011065 in-situ storage Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
Classifications
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
- G01N1/31—Apparatus therefor
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Biomedical Technology (AREA)
- Molecular Biology (AREA)
- Chemical & Material Sciences (AREA)
- General Physics & Mathematics (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- Analytical Chemistry (AREA)
- Physics & Mathematics (AREA)
- Pathology (AREA)
- Urology & Nephrology (AREA)
- Hematology (AREA)
- Cell Biology (AREA)
- Microbiology (AREA)
- Biotechnology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Investigating Or Analysing Biological Materials (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
- Sampling And Sample Adjustment (AREA)
Abstract
The invention discloses a kind of group pens for reducing reagent dosage to apply in the detection of cell climbing sheet immunohistochemistry, and cell inoculation is carried out cell climbing sheet in glass slide formula culture dish, immunocyte histochemical stain identification is carried out after cell culture；The glass slide formula culture dish is equipped with culture dish body and ware lid, and culture dish figure number is divided to two kinds of single hole room and porous room；Each pore chamber area is greater than glass slide, and ware body bottom is used directly as glass slide, and ware body bottom lateral surface is equipped with rectangle frame trace line chase corresponding with glass slide size, under external force can deviate from the bottom of ware body along rectangle frame trace line chase.The various immunohistochemical stainings that the present invention is suitable for glass slice are tested, and antibody and reagent dosage can be substantially reduced, and are avoided liquid trickling when dyeing and are spread, improve service speed.It is especially applicable for carrying out extensive, the other immunohistochemical staining of large sample multiple groups in the experimental study of cell climbing sheet or cell smear.
Description
The application is application number: 201410500701.1, the applying date: 2014.9.26, title: " multipurpose immunohistochemistry pen
The divisional application of application in cell climbing sheet Immunohistochemical detection ".
Technical field
The present invention relates to a kind of application of immunohistochemistry pen in cell climbing sheet Immunohistochemical detection.
Background technique
With specific antibody to the labels of certain chemical constituents analysis in histotomy and its cell climbing sheet and its content into
Row tissue and cell in-situ are qualitative, position or quantitative study, this technology are known as immunocytochemistry
(immunocytochemistry) technology.
Usual cell climbing sheet Immunohistochemical detection is to carry out dying operation one by one to individual cell climbing sheet, generally
A kind of antibody, the detection of albumen and gene can only be done to single sample, it is difficult to which competent extensive, multisample and the other medicine of multiple groups are real
Test scientific research.
Summary of the invention
It is suitable for various immunohistochemical stainings the purpose of the present invention is to provide one kind to test, antibody can be substantially reduced
And reagent dosage, it avoids liquid trickling when dyeing and spreads, improve the multipurpose immunohistochemistry pen of service speed in cell climbing sheet
Application in Immunohistochemical detection.
The technical solution of the invention is as follows:
The application of a kind of multipurpose immunohistochemistry pen in cell climbing sheet Immunohistochemical detection, it is characterized in that: by thin
Born of the same parents are inoculated in glass slide formula culture dish and carry out cell climbing sheet, and immunocyte histochemical stain mirror is carried out after cell culture
It is fixed；The glass slide formula culture dish is equipped with culture dish body and ware lid, and culture dish figure number is divided to two kinds of single hole room and porous room；Each hole
Room area is greater than glass slide, and ware body bottom is used directly as glass slide, and ware body bottom lateral surface is equipped with corresponding with glass slide size
Rectangle frame trace line chase under external force can deviate from the bottom of ware body along rectangle frame trace line chase；
The immunocyte histochemical stain identification successively includes the following steps:
(1) 15 min are fixed with ice acetone or 4% paraformaldehyde is fixed；
(2) flowing water rinses, and is cleaned sample 3 times with PBS；
(3) 10 min are incubated for 0.5%Triton X-100；
(4) 0.3%H2O2It is incubated for 10 min；
(5) after cleaning sample 3 times with PBS, hot wind is sufficiently dry；
(7) ink is drawn with immunohistochemistry pen on cell climbing sheet and divides 2-20 separation dyeing area；
(8) air is sufficiently dry；
(9) it is closed with normal two antiserum and is incubated for 10 min；
(10) mouse is added dropwise respectively in each cell dyeing area or rabbit-anti first antibody is incubated for 30~60 min；
(11) it absorbs each cell dyeing area liquid and is cleaned sample 3 times with PBS；
(12) enzyme-linked mouse is added dropwise or rabbit secondary antibody working solution is incubated for 30~60 min；
(13) PBS is cleaned sample 3 times；
(14) DAB develops the color, and is protected from light, under the microscope；
(15) distillation washing；
(16) haematoxylin lining dye；
(17) hydrochloride alcohol breaks up, and originally washes；
(18) aqueous mounting medium mounting.
The ink is prepared from the following ingredients in percentage by mixing and forms: rosin 2 ~ 6%, beeswax 18 ~ 26%, paraffin 3 ~ 8%,
Isopropyl ether 5 ~ 10%, methylene chloride 5 ~ 10%, gasoline 25 ~ 30%, carbon disulfide 12 ~ 17%, carbon tetrachloride 2 ~ 6%, chloroform 2 ~ 6%, ring
Ketone 2 ~ 6%；The sum of above-mentioned dosage of each component is 100%；
Or the ink is made of following component mixing: polystyrene 2 ~ 4%, DMF9 ~ 11%, THF9 ~ 11%, rosin 2 ~ 4%,
Beeswax 13 ~ 16%, paraffin 2 ~ 4%, isopropyl ether 4 ~ 6%, methylene chloride 4 ~ 6%, gasoline 18 ~ 22%, carbon disulfide 9 ~ 11%, carbon tetrachloride
9 ~ 11%, chloroform 1 ~ 2%, cyclohexanone 1 ~ 2%, kerosene 2 ~ 4%, turpentine oil 1 ~ 2%；The sum of above-mentioned dosage of each component is 100%；
Or the ink is made of following component mixing: stearic acid 30 ~ 36%, 15 ~ 20%, 46 DEG C of paraffin 9 ~ 15% of benzene, isopropyl
Ether 7 ~ 11%, carbon disulfide 15 ~ 20%, chloroform 15 ~ 20%；The sum of above-mentioned dosage of each component is 100%.
It under external force can be by the bottom of ware body before the abjection of rectangle frame trace line chase, first with recessed along sharp keen cutter
Ditch is drawn, is hooked, and then adds to be deviate from the single hole slot bottom of ware body with external force in ware bottom surface.
The immunohistochemistry pen includes pen container, the spongioid cylindrical body pen core of setting in pen container, is perfused with ink in pen container,
Wooden pen core is arranged in spongioid cylindrical body pen core front end；Spongioid cylindrical body pen core is placed in pen container, and pen container is rectangular
Body or cylindrical body, it is identical that pen container mouth outside has screw thread to match with the internal screw thread of interior pen cap；Stainless shot one is placed in pen container, is shaken
Make the effect for mixing ink when shaking pen container；The interior pen cap rise closing pen container mouth, prevent ink volatilization keep its liquid phase state and
Couple the effect of outer pen cap, inner surface has the internal screw thread with the conjunction of pen container tone, can spiral cover pen container mouth, the outer surface of interior pen cap is set
There is the fore shaft for matching and coincideing with outer pen cap inner lip recessed, cylindrical body pen core end side is equipped with ink brush；The inner lip of outer pen cap
It is recessed to be equipped with fore shaft week, matches and coincide with interior pen cap outer lip, pen cap and fixed interior pen cap are integrated unlatching pen container mouth in closing,
And play a part of to propose ink brush.
One angle of porous room culture dish is in bevel-faced form, facilitates bearing mark；An angle for cultivating ware lid is in and culture
The bevel-faced form that ware cooperates in the angle on inclined-plane；Cultivate interleave depth >=10mm of ware lid and culture dish previous anastomotic periphery.
The present invention is conducive to the dyeing of immunohistochemistry, and staining procedure is similar to Immunohistochemistry, but sample process
Temperature, the time, the standards such as reagent concentration are consistent, and the primary antibody type of label is more, and comparativity and reliability are significantly increased！Processing
Sample size it is more, efficient quick.PBS cleaning, serum are incubated for, mark secondary antibody, and lining dye (haematoxylin) etc. does not need the reality separated
It tests operation and can synchronize and handled, convenient and efficient, experiment condition standard is consistent and easy to control.It is carried out using large area cell climbing sheet
The temperature of its processing of cell culture, the time, the standards such as reagent concentration are consistent, keep the comparativity of experimental result and reliability aobvious
It writes and improves.The quantity of Tissue Culture Dish hole slot is reduced, easy to operate, efficient quick.Specific ink formulations have fully ensured that work
Make effect.It is tested suitable for various immunohistochemical stainings, including；Paraffin tissue sections, frozen tissue section and cell are climbed
The immunohistochemical staining of piece is tested, and antibody and reagent dosage can be substantially reduced, and is avoided liquid trickling when dyeing and is spread, mentions
High service speed；It is especially applicable for carrying out on a large scale in the experimental study of cell climbing sheet or cell smear, large sample multiple groups are other
Immunohistochemical staining.
The group pen is suitable for carrying out on the cell climbing sheet of the histotomy that glass is carrier and polystyrene material carrier
Various immunohistochemical staining experiments, can substantially reduce antibody and reagent dosage, avoid liquid trickling when dyeing and spread, mention
High service speed.It can carry out extensive, multisample and the other medical experiment scientific research of multiple groups.
Detailed description of the invention
Present invention will be further explained below with reference to the attached drawings and examples.
Fig. 1 is the structural schematic diagram of culture dish of the present invention.
Fig. 2 is the structural schematic diagram of immunohistochemistry pen.
Fig. 3 is ware body bottom abjection schematic diagram.
Specific embodiment
The application of a kind of multipurpose immunohistochemistry pen in cell climbing sheet Immunohistochemical detection, by cell inoculation in load
Cell climbing sheet is carried out in slide formula culture dish, immunocyte histochemical stain identification is carried out after cell culture；The load
Slide formula culture dish is equipped with culture dish body 1 and ware lid 2, and culture dish figure number is divided to two kinds of single hole room and porous room；Each pore chamber area
Greater than glass slide, ware body bottom 3 is used directly as glass slide, and ware body bottom lateral surface is equipped with rectangle corresponding with glass slide size
Frame trace line chase 4 under external force can deviate from the bottom of ware body along rectangle frame trace line chase；
The immunocyte histochemical stain identification successively includes the following steps:
(1) 15 min are fixed with ice acetone or 4% paraformaldehyde is fixed；
(2) flowing water rinses, and is cleaned sample 3 times with PBS；
(3) 10 min are incubated for 0.5%Triton X-100；
(4) 0.3%H2O2It is incubated for 10 min；
(5) after cleaning sample 3 times with PBS, hot wind is sufficiently dry；
(7) ink is drawn with immunohistochemistry pen on cell climbing sheet and divides 2-20 separation dyeing area；
(8) air is sufficiently dry；
(9) it is closed with normal two antiserum and is incubated for 10 min；
(10) mouse is added dropwise respectively in each cell dyeing area or rabbit-anti first antibody is incubated for 30~60 min；
(11) it absorbs each cell dyeing area liquid and is cleaned sample 3 times with PBS；
(12) enzyme-linked mouse is added dropwise or rabbit secondary antibody working solution is incubated for 30~60 min；
(13) PBS is cleaned sample 3 times；
(14) DAB develops the color, and is protected from light, under the microscope；
(15) distillation washing；
(16) haematoxylin lining dye；
(17) hydrochloride alcohol breaks up, and originally washes；
(18) aqueous mounting medium mounting.
The ink is prepared from the following ingredients in percentage by mixing and forms: rosin 2 ~ 6%, beeswax 18 ~ 26%, paraffin 3 ~ 8%,
Isopropyl ether 5 ~ 10%, methylene chloride 5 ~ 10%, gasoline 25 ~ 30%, carbon disulfide 12 ~ 17%, carbon tetrachloride 2 ~ 6%, chloroform 2 ~ 6%, ring
Ketone 2 ~ 6%；The sum of above-mentioned dosage of each component is 100%；
Or the ink is made of following component mixing: polystyrene 2 ~ 4%, DMF9 ~ 11%, THF9 ~ 11%, rosin 2 ~ 4%,
Beeswax 13 ~ 16%, paraffin 2 ~ 4%, isopropyl ether 4 ~ 6%, methylene chloride 4 ~ 6%, gasoline 18 ~ 22%, carbon disulfide 9 ~ 11%, carbon tetrachloride
9 ~ 11%, chloroform 1 ~ 2%, cyclohexanone 1 ~ 2%, kerosene 2 ~ 4%, turpentine oil 1 ~ 2%；The sum of above-mentioned dosage of each component is 100%；
Or the ink is made of following component mixing: stearic acid 30 ~ 36%, 15 ~ 20%, 46 DEG C of paraffin 9 ~ 15% of benzene, isopropyl
Ether 7 ~ 11%, carbon disulfide 15 ~ 20%, chloroform 15 ~ 20%；The sum of above-mentioned dosage of each component is 100%.
It under external force can be by the bottom of ware body before the abjection of rectangle frame trace line chase, first with recessed along sharp keen cutter
Ditch is drawn, is hooked, and then adds to be deviate from the single hole slot bottom of ware body with external force in ware bottom surface.
The immunohistochemistry pen includes pen container 5, the spongioid cylindrical body pen core 6 of setting in pen container, is perfused with oil in pen container
Wooden pen core 10 is arranged in ink, spongioid cylindrical body pen core front end；Spongioid cylindrical body pen core is placed in pen container, and pen container is
Cuboid or cylindrical body, it is identical that pen container mouth outside has screw thread to match with the internal screw thread of interior pen cap 7；Stainless shot 8 is placed in pen container
One, make the effect for mixing ink when rocking pen container；The interior pen cap plays closing pen container mouth, prevents ink volatilization from keeping its liquid phase
State and the effect for coupling outer pen cap, inner surface have the internal screw thread closed with pen container tone, can spiral cover pen container mouth, interior pen cap outside
Surface is recessed equipped with identical fore shaft is matched with outer 9 inner lip of pen cap, and cylindrical body pen core end side is equipped with ink brush 11；Outer pen cap
To be equipped with fore shaft inner lip week recessed, match and coincide with interior pen cap outer lip, act pen cap and fixed interior pen cap in closing and be integrated out
Pen container mouth is opened, and plays a part of to propose ink brush.Ink is set on interior pen cap and exchanges hole 12.
One angle of porous room culture dish is in bevel-faced form, facilitates bearing mark；An angle for cultivating ware lid is in and culture
The bevel-faced form that ware cooperates in the angle on inclined-plane；Cultivate interleave depth >=10mm of ware lid and culture dish previous anastomotic periphery.
Claims (2)
1. a kind of group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry, it is characterized in that: cell is connect
Kind carries out cell climbing sheet in glass slide formula culture dish, and immunocyte histochemical stain identification is carried out after cell culture；
The glass slide formula culture dish is equipped with culture dish body and ware lid, and culture dish figure number is divided to two kinds of single hole room and porous room；Each pore chamber
Area is greater than glass slide, and ware body bottom is used directly as glass slide, and ware body bottom lateral surface is equipped with length corresponding with glass slide size
Square box trace line chase under external force can deviate from the bottom of ware body along rectangle frame trace line chase, and acquisition has culture thin
Born of the same parents face is ready for use on various Immunohistochemical detections just as the ware bottom of glass slide；
The immunocyte histochemical stain identification successively includes the following steps:
(1) 15 min are fixed with ice acetone or 4% paraformaldehyde is fixed；
(2) flowing water rinses, and is cleaned sample 3 times with PBS；
(3) 10 min are incubated for 0.5%Triton X-100；
(4) 0.3%H2O2It is incubated for 10 min；
(5) after cleaning sample 3 times with PBS, hot wind is sufficiently dry；
(7) ink is drawn with immunohistochemistry pen on cell climbing sheet and divides 2-20 separation dyeing area；
(8) air is sufficiently dry；
(9) it is closed with normal two antiserum and is incubated for 10 min；
(10) mouse is added dropwise respectively in each cell dyeing area or rabbit-anti first antibody is incubated for 30~60 min；
(11) it absorbs each cell dyeing area liquid and is cleaned sample 3 times with PBS；
(12) enzyme-linked mouse is added dropwise or rabbit-anti secondary antibody working solution is incubated for 30~60 min；
(13) PBS is cleaned sample 3 times；
(14) DAB develops the color, and is protected from light, under the microscope；
(15) distillation washing；
(16) haematoxylin lining dye；
(17) hydrochloride alcohol breaks up, and originally washes；
(18) aqueous mounting medium mounting；
The ink is prepared from the following ingredients in percentage by mixing composition: rosin 2 ~ 6%, beeswax 18 ~ 26%, paraffin 3 ~ 8%, isopropyl
Ether 5 ~ 10%, methylene chloride 5 ~ 10%, gasoline 25 ~ 30%, carbon disulfide 12 ~ 17%, carbon tetrachloride 2 ~ 6%, chloroform 2 ~ 6%, cyclohexanone 2
~6%；The sum of above-mentioned dosage of each component is 100%；
The immunohistochemistry pen includes pen container, the spongioid cylindrical body pen core of setting in pen container, is perfused with ink, sponge in pen container
Wooden pen core is arranged in the cylindrical body pen core front end of sample；Spongioid cylindrical body pen core is placed in pen container, pen container be cuboid or
Cylindrical body, it is identical that pen container mouth outside has screw thread to match with the internal screw thread of interior pen cap；Stainless shot one is placed in pen container, rocks pen
Make the effect for mixing ink when cylinder；The interior pen cap plays closing pen container mouth, prevents ink volatilization from keeping its liquid phase state and connection
The effect of outer pen cap, inner surface have the internal screw thread closed with pen container tone, can spiral cover pen container mouth, the outer surface of interior pen cap be equipped with
The identical fore shaft of outer pen cap inner lip matching is recessed, and cylindrical body pen core end side is equipped with ink brush；The inner lip week of outer pen cap sets
Have that fore shaft is convex, coincide with recessed match of interior pen cap fore shaft, rise in closing pen cap and it is fixed in pen cap be integrated and open pen container mouth, and rise
It is proposed the effect of ink brush.
2. the group pen according to claim 1 for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry,
It is characterized in: under external force can be by the bottom of ware body before the abjection of rectangle frame trace line chase, first with recessed along sharp keen cutter
Ditch is drawn, is hooked, and then slightly uses external force that can deviate from the single hole slot bottom of ware body in ware bottom surface.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201610404344.8A CN105938064B (en) | 2014-09-26 | 2014-09-26 | The group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry |
Applications Claiming Priority (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201410500701.1A CN104359741B (en) | 2014-09-26 | 2014-09-26 | Application of multi-functional PAP pen in cell climbing immunohistochemical detection |
| CN201610404344.8A CN105938064B (en) | 2014-09-26 | 2014-09-26 | The group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry |
Related Parent Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201410500701.1A Division CN104359741B (en) | 2014-09-26 | 2014-09-26 | Application of multi-functional PAP pen in cell climbing immunohistochemical detection |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN105938064A CN105938064A (en) | 2016-09-14 |
| CN105938064B true CN105938064B (en) | 2019-03-19 |
Family
ID=52527024
Family Applications (5)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610404345.2A Active CN105938065B (en) | 2014-09-26 | 2014-09-26 | The fast group pen of service speed is applied in the detection of cell climbing sheet immunohistochemistry |
| CN201610404343.3A Active CN106092703B (en) | 2014-09-26 | 2014-09-26 | Application of the easy to operate groupization pen in the detection of cell climbing sheet immunohistochemistry |
| CN201410500701.1A Active CN104359741B (en) | 2014-09-26 | 2014-09-26 | Application of multi-functional PAP pen in cell climbing immunohistochemical detection |
| CN201610404344.8A Active CN105938064B (en) | 2014-09-26 | 2014-09-26 | The group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry |
| CN201610404342.9A Active CN106092702B (en) | 2014-09-26 | 2014-09-26 | The group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry |
Family Applications Before (3)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610404345.2A Active CN105938065B (en) | 2014-09-26 | 2014-09-26 | The fast group pen of service speed is applied in the detection of cell climbing sheet immunohistochemistry |
| CN201610404343.3A Active CN106092703B (en) | 2014-09-26 | 2014-09-26 | Application of the easy to operate groupization pen in the detection of cell climbing sheet immunohistochemistry |
| CN201410500701.1A Active CN104359741B (en) | 2014-09-26 | 2014-09-26 | Application of multi-functional PAP pen in cell climbing immunohistochemical detection |
Family Applications After (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201610404342.9A Active CN106092702B (en) | 2014-09-26 | 2014-09-26 | The group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry |
Country Status (1)
| Country | Link |
|---|---|
| CN (5) | CN105938065B (en) |
Families Citing this family (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105865879A (en) * | 2016-05-19 | 2016-08-17 | 四川金域医学检验中心有限公司 | Operation method for immunohistochemical staining of frozen sections |
| CN106248464B (en) * | 2016-09-02 | 2020-06-09 | 四川大学 | Cell climbing sheet efficient dyeing device |
| CN107699486A (en) * | 2017-09-22 | 2018-02-16 | 山东省农业科学院畜牧兽医研究所 | For dyeing the Tissue Culture Dish and cell culture dyeing, observational technique of observation |
| CN109100503B (en) * | 2018-09-20 | 2021-02-02 | 同济大学 | Immunohistochemical pen liquid and preparation method thereof |
| CN110887720A (en) * | 2019-12-11 | 2020-03-17 | 武汉原谷生物科技有限责任公司 | Immunohistochemical pen semisolid pen paste and preparation method thereof |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1458284A (en) * | 2002-05-15 | 2003-11-26 | 上海新世界基因技术开发有限公司 | Cell line chip |
| CN201402229Y (en) * | 2009-03-18 | 2010-02-10 | 南通大学 | Combination tool for paraffin wax tissue chip |
| CN102466729A (en) * | 2010-11-05 | 2012-05-23 | 北京工业大学 | Method for screening tumor specificity target and targeting ligand based on tissue chip |
| CN202322860U (en) * | 2011-11-14 | 2012-07-11 | 江苏百奥特医疗仪器科技有限公司 | High-resolution imaged cell culture dish with glass bottom |
| CN103037970A (en) * | 2010-05-21 | 2013-04-10 | Lab901有限公司 | Membrane incubation device |
| CN203324275U (en) * | 2013-05-13 | 2013-12-04 | 湖南光琇高新生命科技有限公司 | Immunochemical staining water-blocking device, cell culture dish and cell culture plate |
| CN103525773A (en) * | 2013-10-18 | 2014-01-22 | 武汉市畜牧兽医科学研究所 | Method for improving porcine reproductive and respiratory syndrome virus target cell infection titer and application of method |
| CN104101701A (en) * | 2014-07-15 | 2014-10-15 | 上海中医药大学附属曙光医院 | Efficient cell slide immunohistochemical method |
Family Cites Families (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN2209044Y (en) * | 1994-09-29 | 1995-10-04 | 广西大学 | Pen writing on plate |
| CN2338749Y (en) * | 1998-06-17 | 1999-09-15 | 罗祝泉 | Pencil for marking with circle for immunoassary |
| JP2008533214A (en) * | 2004-12-07 | 2008-08-21 | イッサム リサーチ ディベロップメント カンパニー オブ ザ ヘブリュー ユニバーシティー オブ エルサレム | Nanoparticle capture spherical composites, their manufacturing process and use |
| CN201386105Y (en) * | 2009-04-02 | 2010-01-20 | 武汉大学 | Folded-angle groove-type cell slide |
| CN101629953A (en) * | 2009-08-07 | 2010-01-20 | 中国海洋大学 | On-site detection immuno-chip and preparation method thereof and application |
| CN201740686U (en) * | 2010-06-30 | 2011-02-09 | 中国人民解放军第三军医大学第三附属医院 | Universal pathological histocyte chemical staining glass slide |
| CN202757821U (en) * | 2012-07-11 | 2013-02-27 | 南京金斯瑞生物科技有限公司 | Reserving device for liquid on solid surface |
| CN203238261U (en) * | 2013-05-21 | 2013-10-16 | 中国人民解放军南京军区南京总医院 | Cell culture dish specially used for cell climbing piece |
| CN103352005B (en) * | 2013-07-30 | 2015-12-23 | 中国人民解放军第三军医大学第二附属医院 | A kind of culture dish |
-
2014
- 2014-09-26 CN CN201610404345.2A patent/CN105938065B/en active Active
- 2014-09-26 CN CN201610404343.3A patent/CN106092703B/en active Active
- 2014-09-26 CN CN201410500701.1A patent/CN104359741B/en active Active
- 2014-09-26 CN CN201610404344.8A patent/CN105938064B/en active Active
- 2014-09-26 CN CN201610404342.9A patent/CN106092702B/en active Active
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1458284A (en) * | 2002-05-15 | 2003-11-26 | 上海新世界基因技术开发有限公司 | Cell line chip |
| CN201402229Y (en) * | 2009-03-18 | 2010-02-10 | 南通大学 | Combination tool for paraffin wax tissue chip |
| CN103037970A (en) * | 2010-05-21 | 2013-04-10 | Lab901有限公司 | Membrane incubation device |
| CN102466729A (en) * | 2010-11-05 | 2012-05-23 | 北京工业大学 | Method for screening tumor specificity target and targeting ligand based on tissue chip |
| CN202322860U (en) * | 2011-11-14 | 2012-07-11 | 江苏百奥特医疗仪器科技有限公司 | High-resolution imaged cell culture dish with glass bottom |
| CN203324275U (en) * | 2013-05-13 | 2013-12-04 | 湖南光琇高新生命科技有限公司 | Immunochemical staining water-blocking device, cell culture dish and cell culture plate |
| CN103525773A (en) * | 2013-10-18 | 2014-01-22 | 武汉市畜牧兽医科学研究所 | Method for improving porcine reproductive and respiratory syndrome virus target cell infection titer and application of method |
| CN104101701A (en) * | 2014-07-15 | 2014-10-15 | 上海中医药大学附属曙光医院 | Efficient cell slide immunohistochemical method |
Non-Patent Citations (1)
| Title |
|---|
| 细胞爬片免疫组化;海风吹海浪起;《百度文库》;https://wenku.baidu.com/view/4de5ca39ba1aa8114431d963.html;20131222;第2-3页 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN105938065B (en) | 2019-03-12 |
| CN106092702B (en) | 2018-07-06 |
| CN105938065A (en) | 2016-09-14 |
| CN106092703A (en) | 2016-11-09 |
| CN106092702A (en) | 2016-11-09 |
| CN104359741A (en) | 2015-02-18 |
| CN105938064A (en) | 2016-09-14 |
| CN104359741B (en) | 2017-01-11 |
| CN106092703B (en) | 2018-07-17 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN105938064B (en) | The group pen for reducing reagent dosage is applied in the detection of cell climbing sheet immunohistochemistry | |
| Liu et al. | Evaluation of decalcification techniques for rat femurs using HE and immunohistochemical staining | |
| Montgomery et al. | A laboratory method for testing the toxicity of protective fungicides | |
| Shi et al. | Co‐culture of human primary hepatocytes and nonparenchymal liver cells in the emulate® liver‐chip for the study of drug‐induced liver injury | |
| CN104101701B (en) | High efficiency cell creep plate ImmunohistochemistryMethods Methods | |
| Fichtman et al. | Imaging metazoan nuclear pore complexes by field emission scanning electron microscopy | |
| CN102590488B (en) | Method for regulating and controlling vitellogenin level of zebra fishes | |
| Harris | Cytology and immunocytochemistry | |
| Klieneberger-Nobel | Methods for the study of the cytology of bacteria and pleuropneumonia-like organisms | |
| Frost et al. | Migration assays for oligodendrocyte precursor cells | |
| CN104593320A (en) | Sweat gland differential induction medium and applications thereof | |
| TW202000797A (en) | Tissue staining composition suitable for inkjet printing and method for tissue staining capable of preventing the composition from being oxidized and keeping consistent color rendering of the tissue | |
| Teixeira et al. | Methacrylate: An alternative fixing agent for identifying the botanical origin of propolis | |
| Goldstein | Transplantation of human embryonic stem cells to the chick embryo | |
| Kowalska et al. | Development of pancreatic acini in embryos of the grass snake Natrix natrix (Lepidosauria, Serpentes) | |
| Inooka | Aggregation of sphingosine-DNA and cell construction using components from egg white | |
| CN110261206A (en) | A kind of flaking method of great Ye gold waist chromosome karyotype analysis | |
| CN103013437B (en) | Interior label stationary liquid for biological specimen preserved through formalin, preparation method and application thereof | |
| CN109182253A (en) | A method of improving oocyte in vitro maturation quality and efficiency | |
| Goldstein | Transplantation of human embryonic stem cells and derivatives to the chick embryo | |
| Maneval | Rapid staining methods | |
| Nigam | Experimental animal physiology and biochemistry | |
| Oberlender et al. | Application of Functional Blocking Antibodies: N-Cadherin and Chick Embryonic Limb Development | |
| Nayaka | Studying lichens | |
| Anderson et al. | Preparing SC spreads with RNs for EM analysis |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| TR01 | Transfer of patent right | ||
| TR01 | Transfer of patent right |
Effective date of registration: 20200817 Address after: 201306 2nd floor, no.979 Yunhan Road, Lingang New Area, China (Shanghai) pilot Free Trade Zone, Pudong New Area, Shanghai Patentee after: Shanghai zhudian Biotechnology Co., Ltd Address before: 226019 Jiangsu city of Nantong province sik Road No. 9 Patentee before: NANTONG University |