CN1818649A - Agarose gel plastic substrate, its production and use - Google Patents
Agarose gel plastic substrate, its production and use Download PDFInfo
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- CN1818649A CN1818649A CN 200610065057 CN200610065057A CN1818649A CN 1818649 A CN1818649 A CN 1818649A CN 200610065057 CN200610065057 CN 200610065057 CN 200610065057 A CN200610065057 A CN 200610065057A CN 1818649 A CN1818649 A CN 1818649A
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Abstract
本发明公开了一种琼脂糖凝胶膜基片及其制备方法与应用。本发明所提供的琼脂糖凝胶膜基片，是在基片表面形成琼脂糖凝胶膜，其中，所述琼脂糖凝胶膜被分隔成不少于两个的独立区域。本发明利用琼脂糖凝胶的良好成膜性和生物相容性，并将基片采用隔断方法将基片分隔为多个独立区域，由于分隔物的存在，使不同区域内的不同生物样品之间不会产生交叉污染。用含有多个独立分隔点样区域的琼脂糖凝胶片制备芯片时，可以实现多项指标样品和多份相同、不同生物样品的同时检测，大大提高了芯片检测效率。The invention discloses an agarose gel film substrate, a preparation method and application thereof. The agarose gel film substrate provided by the present invention forms an agarose gel film on the surface of the substrate, wherein the agarose gel film is divided into no less than two independent regions. The present invention utilizes the good film-forming properties and biocompatibility of agarose gel, and divides the substrate into a plurality of independent regions by partition method. Due to the existence of partitions, different biological samples in different regions There will be no cross-contamination. When the chip is prepared with an agarose gel sheet containing multiple independently separated sampling areas, the simultaneous detection of multiple index samples and multiple identical and different biological samples can be realized, which greatly improves the detection efficiency of the chip.
Description
Technical field
The present invention relates to a kind of agarose gel plastic substrate and preparation method thereof and application.
Background technology
The notion of biochip stems from computer chip.The biochip of narrow sense is meant the microarray of bioactivators such as the high-density DNA that is coated on the solid phase carrier, protein, cell, mainly comprises cDNA microarray, oligonucleotide microarray and protein microarray.For the broad sense biochip, except above-mentioned passive type micro-array chip, also comprise utilize photoetching technique and micro-processing technology make up on the solid substrate surface fluid analysis unit and system with realize that biomolecule is carried out fast, the slim device of miniature solid of large information capacity parallel processing and analysis.Comprise nucleic acid amplification chip, capillary array electrophoresis chip, active electromagnetism biochip etc.
Protein chip technology became and carried out a kind of technology that is widely used of albumen correlative study along with the protein function arrival in research epoch recent years.Influence the protein chip technology application many limiting factors are arranged, wherein most critical is because how the existence of protein structure diversity and albumen multilevel hierarchy is fixed in substrate surface with protein molecular and is kept the sufacing of protein active to become guardian technique in the protein chip technology research.Be used for the fixing surface of protein sample at present and can be divided into one dimension, 2 and 3 dimensional organization by the chemical modification form, the one dimension substrate mainly is to modify one deck polylysine (poly-L-lysine in surface of glass slide, PLL) carry out amino, aldehyde radical, epoxy, mercapto groups etc. on the silylation modification or on the surface, the one dimension substrate is because its surface is comparatively hydrophobic, be fixed in the protein molecular changeableness of substrate surface, stronger non-special absorption is arranged.The two-dimensional substrate surface mainly contains two types, and a kind of is polyglycol (polyethyleneglycol, PEG) compound in the various functionalization of finishing; Another is the self assembly surface, carry out self assembly at gold-plated surface with the compound of the pure and mild chain alkyl mercapto of chain alkyl mercapto alcohol functionalization, two-dimensional substrate surface performance aspect the ankyrin molecule is preferable, but exists compound to synthesize a difficult problem, is not suitable for mass preparation.Three-dimensional substrate surface mainly contains two types, and a kind of is dendritic macromole compound on surface of glass slide is modified, and bifunctional group linking arm in the dendritic macromole finishing is used for the protein molecular covalent bond then." protein slides " as U.S. Fullmoon company promptly belongs to such protecpectic substrate that is used for.The three-dimensional substrate surface of this class is comparatively stable, and protein molecular same substrate surface is by the mode combination of covalent effect or high-affinity, in conjunction with also very firm.Another kind of three-dimensional structure substrate is the three-dimensional substrate of filtration membrane (filter membrane) and hydrogel (hydrogel).Filtration membrane mainly contains polyvinylidene fluoride (Polyvinylidene fluoride, PVDF), cellulose nitrate (Nitrocellulose) film is used for protein chip research, this class film by acting forces such as electrostatic force and hydrophobic forces with the protein molecular combination, have the high characteristics of proteopexy amount, but the non-special absorption in surface of this class film is stronger, is a kind of cellulose nitrate film substrate as the Fast Slides film substrate of German Schleicher and schuell company.Hydrogel (Hydrogel) substrate is that a class is used more three-dimensional substrate at present, and the hydrogel substrate is by the mode ankyrin of diffusion.The hydrogel molecule is easy to keep protein active for protein molecular provides the environment of half wet (semi-wet); The hydrogel substrate itself has the autofluorescence background very low and non-special absorption is low, and is better in the surface of glass slide physical strength.Be a kind of in this type of substrate as the Hydrogel polyacrylamide hydrophilic gel substrate of U.S. Perkin-Elmer Life Scienc company.
Summary of the invention
The purpose of this invention is to provide a kind of agarose gel plastic substrate that has a plurality of isolated areas and preparation method thereof and application.
Agarose gel plastic substrate provided by the present invention is to form agarose-gel film at substrate surface, and wherein, described agarose-gel film is separated into and is no less than two isolated area.
Here, the commonly used separation adopted rubber or the silk-screen oar material that is not higher than 1mm.Substrate is selected glass substrate for use.
The preparation method of this agarose gel plastic substrate has two kinds of methods
Method one comprises the steps:
1) on substrate, prepares agarose-gel film;
2) rubber that has a plurality of separated regions that will be pasted with double faced adhesive tape is attached to the agarose-gel film surface and forms and be no less than two isolated area, then, 40～90 ℃ of bakings 5 minutes～2 hours down, obtains described Ago-Gel substrate; Perhaps, adopt method for printing screen to form on the agarose-gel film surface and be no less than two isolated area, then, cure pastes obtains described Ago-Gel substrate.
Method two comprises the steps:
1) rubber that will be pasted with a plurality of separated regions of tool of double faced adhesive tape is attached on the substrate, substrate is divided into be no less than two isolated area; Perhaps, on substrate slurry is printed in substrate surface by method for printing screen, cure pastes is divided into substrate and is no less than two isolated area;
2) in each zone, add agarose solution, drying and forming-film, through the sodium periodate solution oxidation, the washing drying obtains described Ago-Gel substrate.
Here, slurry can be selected 5100 serial polypropylene inks, white spirit metal ink, glass ink etc. for use.
Agarose gel plastic substrate of the present invention is a kind of biochip base material of function admirable, can be widely used in the preparation of various biochips.
Agarose itself is a kind of polyphosphazene polymer sugar compounds, and agarose solution is a kind of hydrogel, can be at solid (as slide/plastic sheet/silicon chip etc.) surface filming; The agarose film can better remain secured to biological substance activity wherein.The present invention utilizes the good filming and the biocompatibility of Ago-Gel, and adopts the partition method that substrate is divided into a plurality of isolated areas substrate, because the existence of separator makes between the different biological samples in the zones of different and can not produce cross pollution.When preparing chip, can realize that the many index sample is identical with many parts, detect different biological sample the time, improve chip detection efficient greatly with the Ago-Gel sheet that contains a plurality of independently divided point samples zone.
Description of drawings
Fig. 1 is a kind of front view of multiple spot battle array Ago-Gel substrate, and dot matrix is set to 2 * 6;
Fig. 2 is a upward view shown in Figure 1;
Fig. 3 is a kind of front view of multiple spot battle array Ago-Gel substrate, and dot matrix is set to 1 * 2;
Fig. 4 is a kind of front view of multiple spot battle array Ago-Gel substrate, and dot matrix is set to 3 * 8;
Fig. 5 is a kind of front view of multiple spot battle array Ago-Gel substrate, and dot matrix is set to 4 * 4;
Fig. 6 is that dot matrix is set to 1 * 2 dot matrix Ago-Gel substrate preparation protein chip figure as a result;
Fig. 7 is that dot matrix is set to 2 * 5 dot matrix Ago-Gel substrate preparation protein chips figure as a result;
Fig. 8 is that dot matrix is set to 2 * 6 dot matrix Ago-Gel substrate preparation antinuclear antibodies detection chip figure as a result.
Embodiment
The invention provides a kind of Ago-Gel substrate that contains a plurality of independently divided point samples zone, it contains independently divided number of regions and is not less than 2; These separators can be the rubber that is pasted with double faced adhesive tape, also can be hydrophobicity serigraphy slurry, and these separators can not destroy the activity of biological sample.
Its structure such as Fig. 1-shown in Figure 5, wherein, Fig. 1 is a kind of front view of multiple spot battle array Ago-Gel substrate, dot matrix is set to 2 * 6; Fig. 2 is a upward view shown in Figure 1; Fig. 3 is a kind of front view of multiple spot battle array Ago-Gel substrate, and dot matrix is set to 1 * 2; Fig. 4 is a kind of front view of multiple spot battle array Ago-Gel substrate, and dot matrix is set to 3 * 8; Fig. 5 is a kind of front view of multiple spot battle array Ago-Gel substrate, and dot matrix is set to 4 * 4.
The preparation method of this Ago-Gel substrate can adopt following two kinds of methods to prepare:
Method one at first forms the independently divided zone that a plurality of areas equate at cleaning slide or substrate surface, adds the finite concentration agarose solution respectively then in a plurality of independently divided zones, and preparation contains the Ago-Gel substrate in a plurality of independently divided zones.In order at first to form the independently divided zone that a plurality of areas equate, can adopt following method, but be not limited only to following method at cleaning slide or substrate surface.
Certain thickness rubber surface paste can with slide or substrate surface can in conjunction with double faced adhesive tape, double faced adhesive tape and rubber thickness sum can not be higher than 1mm, adopt process for stamping to form the separated region that needed a plurality of area equates being pasted with the double faced adhesive tape rubber surface; This rubber is pasted on cleaning slide or substrate surface; Then, add agarose solution in these zones of separating by rubber, the substrate that contains agarose solution is placed drying and forming-film, and the agarose substrate is put into the sodium periodate solution oxidation after the film forming, ultrapure water washing then, centrifuge dripping drying.
Equally, also can form the independently divided zone that a plurality of areas equate with method for printing screen at cleaning slide or substrate surface, to have certain hydrophobicity slurry and be printed in slide or substrate surface by method for printing screen, be cured under uniform temperature and time conditions by selected hydrophobicity slurry character then, solidify disposed slurry thickness and can not be higher than 1mm.Add agarose solution at these in by hydrophobicity slurry separated region, will contain the agarose solution substrate and place drying and forming-film, the agarose substrate is put into the sodium periodate solution oxidation after the film forming, ultrapure water washing, centrifuge dripping drying.Here, slurry can be selected 5100 serial polypropylene inks, white spirit metal ink, glass ink etc. for use.
Method two forms full wafer agarose film at cleaning slide or substrate, pastes at the agarose substrate surface then and contains a plurality of independently divided rubber fences or form a plurality of independently divided zones by method for printing screen at the agarose film surface.
The agarose that contains full wafer can adopt infiltration method, dropping method, scrape the method preparation of film method and whirl coating: the infiltration method will clean slide or substrate exactly and immerse in the certain density agarose solution, then slide is taken out from solution, according to requirement film thickness needs, can repeated multiple times, to form than thick film; The dropping method is that certain concentration solution is dripped in cleaning slide or substrate surface, allows agarose solution fully sprawl in whole surface of glass slide, then drying and forming-film.Scraping film method is to get finite concentration and a certain amount of agarose solution dropping slide one end, uses certain instrument (as glass rod) that solution is uniformly distributed in slide or substrate surface, drying and forming-film by the method for scraping with agarose solution then; The whirl coating method is that agarose solution is dripped in slide or substrate surface, by photoresist spinner solution is evenly distributed drying and forming-film then at slide or substrate surface.
In order full wafer agarose film to be separated into a plurality of independently divided zones, can adopt following method, but be not limited only to following method.Adopt process for stamping to form the separated region that needed a plurality of area equates on the surface the certain thickness rubber that is pasted with double faced adhesive tape, it is pasted on slide or substrate surface, double faced adhesive tape and rubber thickness sum can not surpass 1mm.Because the agarose aquogel characteristic, the rubber that is pasted on agarose film surface comes off in water easily, can cause the pollution between the sample like this, makes in the detection that can not realize various product with a slice agarose substrate surface; In order the rubber fence firmly can be pasted on agarose film surface, and guarantee in the chip course of reaction, not come off, can after multiple spot battle array rubber fence be pasted on agarose film surface, put into baking oven and toast certain hour at a certain temperature.Baking temperature is 40～90 ℃ in the method, and the time of baking is 5 minutes～2 hours; Can firmly be pasted on agarose film surface and in the chip course of reaction, do not come off through rubber fence behind the overbaking.
Equally, also can form a plurality of independently divided zone that needs on agarose film surface by method for printing screen.By method for printing screen with oar material silk-screen in agarose film surface, be cured by the slurry requirement then; Should have biocompatibility by the oar material that is screen-printed to agarose film surface, and can not cause the infringement of agarose film when it solidifies, oar material thickness is not higher than 1mm after the serigraphy.
The used glass substrate of the present invention normally is of a size of the general microslide of microscope of 75.6mm * 25mm * 1mm, can use not modified microslide to be base material, also can carry out finishing to microslide.
A plurality of independently divided point sample zone Ago-Gel substrate of the present invention is a kind of biochip base material of function admirable.When preparing chip with the Ago-Gel sheet that contains a plurality of independently divided point samples zone, can by certain rule various product be arranged in a plurality of independently divided Ago-Gel substrate surfaces with array format at the microarray point sample instrument that companies such as adopting U.S. Cartesian, Genomic Instrumentation, the rich biology difficult to understand of China produces, when carrying out the chip reaction, identical or different biological sample can be added in each independently divided Ago-Gel substrate dot matrix and carry out biological respinse.Because the existence of separator, make between the different biological samples of adding and can not produce cross pollution.When preparing chip, can realize that the many index sample is identical with many parts, detect different biological sample the time with the Ago-Gel sheet that contains a plurality of independently divided point samples zone.
Below with specific embodiment the present invention is described.
Embodiment 1, dripping method prepare full wafer Ago-Gel substrate, prepare multiple spot battle array agarose substrate with method for printing screen then
Weighing 0.01 gram agarose adds in the 1000mL beaker, adds the 500mL ultrapure water subsequently, puts into micro-wave oven moderate heat heating for dissolving; The agarose solution of getting 1000 μ L with the application of sample rifle is added drop-wise to uses H
2O
2/ H
2SO
4The common slide surface that (3: 7) washing lotion is cleaned allows agarose solution evenly distribute in surface of glass slide; Place drying and forming-film under the room temperature then.
With 10 dot matrix silk screen plates of photochemical method preparation 5 row * 2 row, the every dot matrix size of silk screen is 10 * 10mm, and two dot matrix centre distances are 12mm; With agarose substrate surface silk-screen 5 row * 2 row 10 dot matrix of the hydrophobicity slurry L8026 of Du Pont (E.I.Du Pont Company) in above-mentioned full wafer dropping, the Ago-Gel substrate is put into baking oven and is heating and curing after the serigraphy, clean with ultrapure water then, clean the sodium metaperiodate (NaIO that meron is put into 0.1M
4) oxidation reaction 30min in the solution; Centrifuge dripping after the oxidation reaction obtains 10 dot matrix size Ago-Gel substrate.
By different sizes and different dot matrix wire printing plates such as same light chemical method preparation 1 row * 2 row, 2 row * 2 row, 2 row * 3 row, 3 row * 3 row, 4 row * 4 row, 5 row * 2 row, 6 row * 2 row, 8 row * 3 row, carry out serigraphy and sodium metaperiodate carries out oxidation by above-mentioned same procedure, prepare the multiple spot battle array Ago-Gel substrate of different latticed forms.
By above same procedure, weighing 0.05,0.1,0.2,0.3,0.4,0.5,1.0,2.0,3.0,4.0 and 5.0 gram agaroses are dissolved in the 500ml ultrapure water, preparation variable concentrations agarose solution, drip as stated above and prepare full wafer in the cleaning surface of glass slide and drip agarose, carry out the Ago-Gel substrate of silk-screen preparation 1 row * 2 row, 2 row * 2 row, 2 row * 3 row, 3 row * 3 row, 4 row * 4 row, 5 row * 2 row, 6 row * 2 row, the different dot matrix sizes of 8 row * 3 row and different latticed forms then by above-mentioned same procedure.
Embodiment 2, infusion method prepare full wafer Ago-Gel substrate, prepare multiple spot battle array agarose substrate with method for printing screen then
Weighing 0.01 gram agarose adds in the 1000mL beaker, adds the 500mL ultrapure water subsequently, puts into micro-wave oven moderate heat heating for dissolving.Use H
2O
2/ H
2SO
430 of the common microslides that (3: 7) washing lotion is cleaned insert 30 slide framves on chip, and the slide frame is immersed in the good agarose solution of dissolving, and the slide frame proposes solution rapidly, so repeatedly, allow the agarose solution surface of glass slide that evenly distributes; Place nature volatile dry film forming then under the room temperature.
Press the agarose solution of same procedure dissolving variable concentrations among the embodiment 1, carry out the serigraphy and the oxidation of substrate surface, the Ago-Gel substrate of preparation 1 row * 2 row, 2 row * 2 row, 2 row * 3 row, 3 row * 3 row, 4 row * 4 row, 5 row * 2 row, 6 row * 2 row, the different dot matrix sizes of 8 row * 3 row and different latticed forms.
Embodiment 3, lacquering technique prepare full wafer Ago-Gel substrate, prepare multiple spot battle array agarose substrate with method for printing screen then
Weighing 0.01 gram agarose adds in the 1000mL beaker, adds the 500mL ultrapure water subsequently, puts into micro-wave oven moderate heat heating for dissolving.To use H
2O
2/ H
2SO
4The common microslide that (3: 7) washing lotion is cleaned is put into photoresist spinner, and the agarose solution of 800 μ L is added drop-wise to slide central authorities, with the even glue of 500 rev/mins of speed, with 1000 rev/mins of whirl coatings, allows agarose solution be evenly distributed on surface of glass slide; Place nature volatile dry film forming then under the room temperature.
Press the agarose solution of same procedure dissolving variable concentrations among the embodiment 1, carry out the serigraphy and the oxidation of substrate surface, the Ago-Gel substrate of preparation 1 row * 2 row, 2 row * 2 row, 2 row * 3 row, 3 row * 3 row, 4 row * 4 row, 5 row * 2 row, 6 row * 2 row, the different dot matrix sizes of 8 row * 3 row and different latticed forms.
Embodiment 4, elder generation's preparation full wafer Ago-Gel substrate prepare multiple spot battle array Ago-Gel substrate with pasting the fence method then
Weighing 0.01 gram agarose adds in the 1000mL beaker, adds the 500mL ultrapure water subsequently, puts into micro-wave oven moderate heat heating for dissolving 10min; Keeping the agarose solution temperature is 40～80 ℃; Prepare full wafer by same procedure among embodiment 1 or embodiment 2 or the embodiment 3 and drip the agarose substrate, press the sodium periodate solution oxidation reaction 30min of method usefulness 0.1M among the embodiment 1.
Paste double faced adhesive tape on the natural rubber surface, adopt the mechanical stamping method to stamp out the rubber fence of 1 row * 2 row, 2 row * 2 row, 2 row * 3 row, 3 row * 3 row, 4 row * 4 row, 5 row * 2 row, 6 row * 2 row, the different dot matrix sizes of 8 row * 3 row and different latticed forms at rubber surface then, fence is pasted on the agarose substrate surface, puts into baking oven and toast; Dry by the fire back agarose substrate room temperature and placed natural cooling, obtained the agarose substrate that multiple spot battle array fence is separated.
Embodiment 5, elder generation prepare multiple spot battle array Ago-Gel substrate with dripping method then in the surface of glass slide serigraphy
With 10 dot matrix silk screen plates of photochemical method preparation 5 row * 2 row, the every dot matrix size of silk screen is 10 * 10mm, and two dot matrix centre distances are 12mm; With 10 dot matrix of the hydrophobicity slurry L8026 of Du Pont at cleaning slide surface silk-screen 5 row * 2 row, put into baking oven and be heating and curing, clean with ultrapure water then, centrifuge dripping obtains being divided into the slide of 10 dot matrix.Similar the method can have multiple spot battle array silk-screen fence at the surface of glass slide silk-screen.
Weighing 0.01 gram agarose adds in the 1000mL beaker, adds the 500mL ultrapure water subsequently, puts into micro-wave oven moderate heat heating for dissolving.Add 30 μ L agarose solutions with the application of sample rifle in the little lattice of each silk-screen of slide, room temperature is placed dry; Dry meron is put into 0.1M sodium metaperiodate (NaIO
4) oxidation reaction 30min in the solution; Centrifuge dripping after the oxidation reaction obtains 10 dot matrix Ago-Gel substrates.
Adopt the Ago-Gel substrate of above similar approach preparation 1 row * 2 row, 2 row * 2 row, 2 row * 3 row, 3 row * 3 row, 4 row * 4 row, 5 row * 2 row, 6 row * 2 row and different dot matrix sizes of 8 row * 3 row and different latticed forms.
Multiple spot battle array fence is pasted in surface of glass slide by embodiment 6, elder generation, prepares multiple spot battle array Ago-Gel substrate with dripping method then
With thickness is the natural rubber surface stickup double faced adhesive tape of 0.5mm, adopts mechanical stamping then, stamps out 10 dot matrix fences of 5 row * 2 row, and the every dot matrix size of fence is 10 * 10mm, and two dot matrix centre distances are 12mm; Then 10 dot matrix fences are pasted on surface of glass slide.
Weighing 0.01 gram agarose adds in the 1000mL beaker, adds the 500mL ultrapure water subsequently, puts into micro-wave oven moderate heat heating for dissolving.Add 30 μ L agarose solutions with the application of sample rifle in each dot matrix of slide, room temperature is placed dry; Dry meron is put into 0.1M sodium metaperiodate (NaIO
4) oxidation reaction 30min in the solution; Centrifuge dripping after the oxidation reaction obtains 10 dot matrix Ago-Gel substrates.
Adopt the Ago-Gel substrate of above similar approach preparation 1 row * 2 row, 2 row * 2 row, 2 row * 3 row, 3 row * 3 row, 4 row * 4 row, 5 row * 2 row, 6 row * 2 row, the different dot matrix sizes of 8 row * 3 row and different latticed forms.
Embodiment 7, multiple spot battle array Ago-Gel substrate are applied to the chip preparation
The preparation process of biochip is as follows: 1. (60% glycerine/40%1 * PBS) dilutions makes the human IgG sample of 1.0mg/ml, takes out 10 μ L and shifts and add in the 384 hole sample panel with 60% glycerine point sample damping fluid with the human IgG sample of 10mg/ml; 2. by automatic spot sample device ready sampling liquid is assigned to the Ago-Gel substrate surface of 10 dot matrix according to certain dot matrix way row.Every chip comprises the individual array of 10 (2 row * 5 row), and each array comprises 25 points, and each dot spacing is from being 400 μ m.Each array forms independently reaction chamber; 3. chip room temperature behind the point sample is placed and fixedly spend the night; 4. fixing back chip shakes capping 30min with 0.1%BSA solution; 5. sealing back chip cleans with ultrapure water, then centrifuge dripping; 6. the fluorescently-labeled goat anti-human antibody's solution that in each hole of agarose substrate of 10 dot matrix, adds 30 μ L, room temperature reaction 30min subsequently; 7. reaction back chip cleans centrifuge dripping with ultrapure water; 8. chip scanning and data processing.Testing result as shown in Figure 7.
Testing result shown in Figure 6 can adopt with last identical method and carry out, and only need use the agarose gel plastic substrate that contains the individual array of 2 (1 row * 2 row) instead and prepare chip and get final product.
Embodiment 8, multiple spot battle array Ago-Gel substrate are applied to the preparation of antinuclear antibodies detection chip
The preparation process of biochip is as follows: 1. (60% glycerine/40%1 * PBS) are diluted to finite concentration, and every kind of sample takes out 10 μ L and shifts and add in the 384 hole sample panel with 60% glycerine point sample damping fluid with various antinuclear antibodies samples; 2. by automatic spot sample device ready sampling liquid is assigned to the Ago-Gel substrate surface of 12 dot matrix according to certain dot matrix way row.Every chip comprises the individual array of 12 (6 row * 2 row), and each array comprises 63 points, and each lattice distance is from being 400 μ m.Each array forms independently reaction chamber; 3. chip room temperature behind the point sample is placed and fixedly spend the night; 4. fixing back chip shakes capping 30min with 0.1%BSA solution; 5. sealing back chip cleans with ultrapure water, then centrifuge dripping; 6. the blood serum sample that in each hole of agarose substrate of 12 dot matrix, adds 30 μ L, room temperature reaction 30min subsequently; 7. reaction back chip cleans centrifuge dripping with ultrapure water; 8. add fluorescently-labeled goat anti-human antibody's solution, react 30min under the room temperature; 9. reaction back chip cleans centrifuge dripping with ultrapure water; 10 chip scannings and data processing.The result as shown in Figure 8.
Claims (7)
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Cited By (5)
| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2011015131A1 (en) * | 2009-08-07 | 2011-02-10 | 中国海洋大学 | Immunoassay chip for fish lymphocystis disease virus, preparation method and application thereof |
| CN101629954B (en) * | 2009-08-07 | 2012-09-26 | 中国海洋大学 | Immunity detection chip of prawn white spot syndrome virus (WSSV) and preparation method thereof and application |
| CN109827814A (en) * | 2019-03-11 | 2019-05-31 | 南京大学 | Preparation of a Novel Atmospheric Particulate Matter Agar Sampling Membrane and Solvent-Free Extraction Cell Exposure Method |
| CN112630420A (en) * | 2020-12-07 | 2021-04-09 | 北京科技大学 | Method for realizing directional coupling by using glycosyl of antibody and solid phase carrier material |
| CN113957129A (en) * | 2021-09-01 | 2022-01-21 | 泰安市农业科学研究院 | Method for screening homozygous material from Chinese cabbage commodity hybrid |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2011015131A1 (en) * | 2009-08-07 | 2011-02-10 | 中国海洋大学 | Immunoassay chip for fish lymphocystis disease virus, preparation method and application thereof |
| CN102132159A (en) * | 2009-08-07 | 2011-07-20 | 中国海洋大学 | Immunoassay chip for fish lymphocystis disease virus, preparation method and application thereof |
| CN101629954B (en) * | 2009-08-07 | 2012-09-26 | 中国海洋大学 | Immunity detection chip of prawn white spot syndrome virus (WSSV) and preparation method thereof and application |
| CN101629955B (en) * | 2009-08-07 | 2012-09-26 | 中国海洋大学 | Fish lymphocystis disease virus (LCDV) immunity detection chip and preparation method and application thereof |
| CN102132159B (en) * | 2009-08-07 | 2013-05-29 | 中国海洋大学 | Fish lymphocyst virus immune detection chip and its preparation method and application |
| CN109827814A (en) * | 2019-03-11 | 2019-05-31 | 南京大学 | Preparation of a Novel Atmospheric Particulate Matter Agar Sampling Membrane and Solvent-Free Extraction Cell Exposure Method |
| CN112630420A (en) * | 2020-12-07 | 2021-04-09 | 北京科技大学 | Method for realizing directional coupling by using glycosyl of antibody and solid phase carrier material |
| CN113957129A (en) * | 2021-09-01 | 2022-01-21 | 泰安市农业科学研究院 | Method for screening homozygous material from Chinese cabbage commodity hybrid |
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