CN103808922B - A kind of screening technique of hybridoma supernatant stage trace antibody - Google Patents
A kind of screening technique of hybridoma supernatant stage trace antibody Download PDFInfo
- Publication number
- CN103808922B CN103808922B CN201310154372.5A CN201310154372A CN103808922B CN 103808922 B CN103808922 B CN 103808922B CN 201310154372 A CN201310154372 A CN 201310154372A CN 103808922 B CN103808922 B CN 103808922B
- Authority
- CN
- China
- Prior art keywords
- supernatant
- antibody
- screening technique
- microlens array
- array chip
- 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
Classifications
-
- 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
- G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
- G01N33/6803—General methods of protein analysis not limited to specific proteins or families of proteins
-
- 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
- G01N33/531—Production of immunochemical test materials
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Hematology (AREA)
- Chemical & Material Sciences (AREA)
- Urology & Nephrology (AREA)
- Biomedical Technology (AREA)
- Physics & Mathematics (AREA)
- Medicinal Chemistry (AREA)
- Food Science & Technology (AREA)
- Biotechnology (AREA)
- Pathology (AREA)
- Microbiology (AREA)
- General Physics & Mathematics (AREA)
- General Health & Medical Sciences (AREA)
- Cell Biology (AREA)
- Biochemistry (AREA)
- Analytical Chemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Bioinformatics & Computational Biology (AREA)
- Biophysics (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Peptides Or Proteins (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
The invention provides the screening technique of trace antibody of a kind of hybridoma supernatant stage, this screening technique comprises the steps: supernatant pre-service, supernatant point sample and aftertreatment, supernatant marks, point sample is tested, double-antibody sandwich detects.First carry out pairing screening when hybridoma cell conditioned medium stage trace antibody, utilize the advantage of the highly sensitive of chip and many flux, avoid a large amount of random monoclonal antibody purification work.When preparing the monoclonal antibody of this kind, this method can be used to filter out pairing in advance even can the effective pairing antibody of pre-sifted clinical detection, then prepare on a large scale, greatly improves efficiency, reduce cost and the risk of preparation.
Description
Technical field
The present invention relates to a kind of antibody screening method, particularly a kind of screening technique of hybridoma supernatant stage trace antibody.
Background technology
Double antibody sandwich method is the most frequently used ELISA method of clinical in vitro diagnosis in vitro detectable antigens, is applicable to the quantitative detection of the polyvalent antigen in detection molecules with at least two antigenic determinants.Its basic functional principle is: the antibody that utilization is connected on solid phase carrier and enzyme labelled antibody are detected two antigenic determinants on antigen molecule respectively and are combined in sample, form solid matrix antibody-antigen-enzyme labelled antibody immune complex.
Traditional classical double-antibody sandwich target antibody screening technique needs to adopt hybridoma technology to prepare monoclonal antibody (abbreviation monoclonal antibody), tiring and specificity of antibody is measured by ELISA method and Western blotting after monoclonal antibody purifying, also have Antibody types, relative affinity to measure, the monoclonal antibody of application of purified sets up the applicable pairing antibody detected of double-antibody sandwich elisa detection method screening.When classic method prepares monoclonal antibody, need to carry out expansion for specific immunogenic all positive colonies and cultivate and carry out monoclonal antibody purifying.But the monoclonal antibody prepared might not carry out double-antibody sandwich pairing for this immunogene, there is certain blindness in preparation.Even if filter out the antibody that can carry out double-antibody sandwich pairing for immunogene, during clinical detection serum effect, also there is very large difference.
Summary of the invention
The object of this invention is to provide a kind of efficiency high and can on purpose carry out monoclonal antibody purification work the hybridoma supernatant stage trace antibody screening technique, to solve problems of the prior art.
To achieve these goals, the present invention is by the following technical solutions:
A screening technique for hybridoma supernatant stage trace antibody, this screening technique comprises the steps:
(1) culture supernatant of the hybridoma formed after getting Fusion of Cells, by exchange buffering system after salting out method removal impurity, obtains pre-service supernatant;
(2) antibody in the pre-service supernatant obtained in step (1) is fixed to microlens array chip;
(3) add horseradish peroxidase-labeled sheep anti-mouse antibody to the microlens array chip utilization of step (2) to react, confirm antibody activity and the fixing situation on microlens array chip in supernatant;
(4) the pre-service supernatant mark horseradish peroxidase will obtained in step (1);
(5) immunogene is added in microlens array chip that step (2) processed, then the supernatant antibody adding the horseradish peroxidase mark that step (4) obtains carries out double-antibody sandwich reaction screening pairing situation respectively, then by checking, negative control confirms, native protein checking draws the pairing supernatant be applicable to.
Screening technique as above, preferably, the concrete operation method of described step (1) is: the culture supernatant of the hybridoma formed after a) getting Fusion of Cells, the saturated ammonium sulfate solution of 80% ~ 150% of supernatant volume is added to described supernatant, 8 ~ 15h is precipitated under 2 ~ 8 DEG C of conditions, in high speed freezing centrifuge, after centrifuging, be precipitated thing; B) described sediment 0.4ml buffer solution, uses super filter tube centrifuging, obtains pre-service supernatant after repeating ultrafiltration 2 ~ 6 times;
Screening technique as above, preferably, the concrete operation method of described step (2) is: after the pre-service supernatant spotting buffer that a) described step (1) obtains dilutes 5 ~ 20 times, point sample is to microlens array chip; B) wash plate 3 ~ 6 times with PBST after hatching 30 ~ 120min under 25 ~ 37 DEG C of conditions, add confining liquid, under 2 ~ 8 DEG C of conditions, hatch 8 ~ 15 hours;
Screening technique as above, preferably, the concrete operation method of described step (3) is: microlens array chip a) obtained to described step (2) adds horseradish peroxidase-labeled sheep anti mouse, hatch 30 ~ 120min under 25 ~ 37 DEG C of conditions after, PBST washing 3 ~ 6 times; B) chemiluminescence scanner scanning is used to draw the image in conjunction with situation that can reflect supernatant and microlens array chip;
Screening technique as above, preferably, the concrete operation method of described step (4) is: supernatant sodium periodate oxidizing process after the pre-service obtained in step (1) is marked upper horseradish peroxidase;
Screening technique as above, preferably, the concrete operation method of described step (5) is: a) immunogene confining liquid is diluted to 0.1 ~ 1 μ g/ml, add in the micropore of the microlens array chip that step (2) processed, under 25 ~ 37 DEG C of conditions, hatch 60 ~ 150min; B) PBST washing 2 ~ 6 times; C) add in the micropore of microlens array chip with the horseradish peroxidase-labeled supernatant that processes of step (3) of confining liquid 5 ~ 20 times dilution, hatch 60 ~ 150min for 25-37 DEG C; D) PBST washing 5 ~ 8 times; E) use chemiluminescence scanner scanning, draw image; F) according to image, microlens array chip there is the supernatant corresponding to the selecting of light signal be the supernatant that can match; G) the pairing supernatant that draws repeat in step (5) a) ~ e) reaction, simultaneously carry out step a) time increase and replace immunogene to carry out the negative control reacted with confining liquid; When negative control microlens array chip is without light signal, and when immunogene reaction has a signal, determine the authentic and valid of unpaired message; H) for the pairing supernatant filtered out, use native protein replace immunogene repeat in step (5) a) ~ e) reaction, the numerical value and the clinical detection concentration linear fit that obtain optical signals are evaluated, and filter out the pairing supernatant information of the most applicable clinical detection.
Screening technique as above, preferably, in described step (1), the centrifugal force of centrifuging operation is 10000 ~ 15000g, and working time is 10 ~ 30min.
Screening technique as above, preferably, the molecular cut off of described super filter tube is less than the molecular weight of antibody in supernatant.
Screening technique as above, preferably, described damping fluid is phosphate buffer, and the concentration of this phosphate buffer is 0.01mol/L, pH is 7.2.
Screening technique as above, preferably, described spotting buffer is the potpourri of carbonate (CBS) and 30% glycerine, and described CBS concentration is 0.05mol/L.
Screening technique as above, preferably, described confining liquid is phosphate buffer and bovine serum albumin mix, and the concentration of this phosphate buffer is 0.1mol/L, pH is 7.2, and the mass volume ratio of bovine serum albumin(BSA) and phosphate buffer is 1:100 ~ 5:100.
Beneficial effect of the present invention is:
During conventional method Dispersal risk, need the positive colony all for immunogene to expand to cultivate, carry out protein purification after obtaining abundant cell conditioned medium liquid, utilize the albumen of purifying to carry out the test of being correlated with.And if test result is undesirable, the process of needs repetition immunity, cell chulture, antibody purification, test obtains the antibody of high-quality, and manufacturing cycle is long, and Expenses Cost is high.The present invention is in positive colony cell conditioned medium during micro-antibody, utilize the advantage of the highly sensitive of microlens array chip and many flux, first dependence test is carried out, filter out the positive colony that pairing is effective, carry out expansion again to cultivate and protein purification, avoid expansion cultivation blindly and protein purification work, preparation time traditional at present can be shortened one times even more, greatly improve efficiency, and reduce cost and the risk of preparation.
Accompanying drawing explanation
Fig. 1 is the process flow diagram of the screening technique of hybridoma supernatant stage trace antibody;
Fig. 2 is that supernatant point sample is to the schematic diagram on microlens array chip;
Fig. 3 is the schematic diagram that microlens array chip supernatant fixes positive test;
Fig. 4 is the immunogene pairing schematic diagram of a supernatant;
Fig. 5 is the native protein pairing schematic diagram of supernatant in the embodiment of the present invention 1;
Fig. 6 is the schematic diagram in conjunction with situation of supernatant and microlens array chip;
Fig. 7 is the native protein pairing schematic diagram of supernatant in the embodiment of the present invention 2.
Embodiment
The process flow diagram of the screening technique of hybridoma supernatant stage trace antibody is shown in Fig. 1, is described further the inventive method below in conjunction with accompanying drawing.
Embodiment 1: the screening of hybridoma supernatant stage trace antibody
The culture supernatant of the hybridoma formed after 1, getting Fusion of Cells, by exchange buffering system after salting out method removal impurity, obtain pre-service supernatant, concrete steps are as follows:
A) 29 parts of cell conditioned medium liquid add equal-volume saturated ammonium sulfate solution respectively, and 4 DEG C precipitate 15 hours, and the centrifugal 10min of 12000g, abandons supernatant and be precipitated thing;
B) sediment is 0.01mol/L by concentration respectively, pH is that after the phosphate buffer dissolving of 7.2, being placed in volume is respectively 0.5ml, and molecular weight is the millipore super filter tube of 50KD, 12000g centrifuging 10min, obtains pre-service supernatant after repeating ultrafiltration 2 times;
2, step 1 pre-service supernatant antibody is fixed to microlens array chip:
A) described step 1 pre-service supernatant uses spotting buffer to dilute 10 times to use GSD1800 point sample instrument point sample to 58 positions (i.e. 29 parts of hybridoma supernatants of microlens array chip afterwards, every part do two parallel), as shown in Figure 2, described spotting buffer is the potpourri of CBS and 30% glycerine, and described CBS concentration is 0.05mol/L;
B) hatch 30min for 37 DEG C, wash plate 3 times with PBST; Add confining liquid 4 DEG C close 8 hours for subsequent use; The compound method of described PBST is: be add the polysorbas20 that volume is 5/10000ths of phosphate buffer volume in the phosphate buffer of 0.1mol/L to concentration; Described confining liquid is phosphate buffer and bovine serum albumin mix, and the concentration of described phosphate buffer is 0.1mol/L, pH is 7.2, and bovine serum albumin(BSA) and phosphate buffer mass volume ratio are 5:100.
3, microlens array chip supernatant fixing situation in step 2 is confirmed:
A) add with confining liquid 1:1000 dilution by volume horseradish peroxidase-labeled sheep anti mouse (note: every part is done a Duplicate Samples to microlens array chip after point sample in step 2, another Duplicate Samples is used for doing step 5, this horseradish peroxidase-labeled sheep anti mouse adopts Wuhan doctor's moral company BA1050,37 DEG C hatch 30min after, PBST washs 3 times;
B) GSS2400 chemiluminescence scanner scanning is used to draw the image in conjunction with situation that can reflect supernatant and microlens array chip, as shown in Figure 3;
4, pre-service supernatant mark horseradish peroxidase: pretreated 29 parts of supernatant sodium periodate oxidizing process are marked upper HRP respectively.
5, use immunogene to carry out double-antibody sandwich reaction screening pairing situation to supernatant, then by checking, negative control confirms, native protein checking draws the pairing supernatant be applicable to:
A) immunogene used confining liquid to be diluted to 0.5 μ g/ml, add in the microlens array chip of step 2 point sample, 37 DEG C of reaction 60, min;
B) PBST washs 3 times;
C) the corresponding step 4 horseradish peroxidase supernatant labelled antibody that a kind of confining liquid dilutes 10 times is added in each hole, 37 DEG C of reaction 60min;
D) PBST washs 6 times;
E) use GSS2400 chemiluminescence scanner scanning, draw image;
F) according to analysis result, draw the mark supernatant numbering that can match, repeat the reaction of this mark supernatant, do negative control simultaneously, determine the authentic and valid of unpaired message;
G) for the pairing supernatant determined, carry out the evaluation of native protein (such as clinical serum), filter out the unpaired message of applicable clinical detection, as shown in Figure 4, Figure 5.
Embodiment 2:
The culture supernatant of the hybridoma formed after 1, getting Fusion of Cells, by exchange buffering system after salting out method removal impurity, obtains pre-service supernatant:
A) 20 parts of cell conditioned medium liquid adds the saturated ammonium sulfate solution of 150% of supernatant volume respectively, 4 DEG C of precipitations 8 hours, and the centrifugal 10min of 12000g, abandons supernatant and be precipitated thing;
B) sediment is 0.01mol/L by concentration respectively, pH is that after the phosphate buffer dissolving of 7.2, being placed in volume is respectively 0.5ml, and molecular weight is the millipore super filter tube of 50KD, 12000g centrifuging 10min, obtains pre-service supernatant after repeating ultrafiltration 2 times;
2, pre-service supernatant protein in step 1 is fixed to microlens array chip, concrete steps are as follows:
A) described step 1 pre-service supernatant uses spotting buffer to dilute 5 times to use GSD1800 point sample instrument point sample to 60 positions of microlens array chip afterwards, described spotting buffer is the potpourri of CBS and 30% glycerine, and described CBS concentration is 0.05mol/L;
B) hatch 60min for 25 DEG C, wash plate 6 times with PBST; Add confining liquid 4 DEG C close 15 hours for subsequent use; The compound method of described PBST is: be add the polysorbas20 that volume is 5/10000ths of phosphate buffer volume in the phosphate buffer of 0.1mol/L to concentration; Described confining liquid is phosphate buffer and bovine serum albumin mix, and the concentration of described phosphate buffer is 0.1mol/L, pH is 7.2, and bovine serum albumin(BSA) and phosphate buffer mass volume ratio are: 1:100.
3, confirm microlens array chip supernatant fixing situation in step 2, concrete steps are as follows:
A) to adding with confining liquid 1:1000 dilution by volume enzyme labeling sheep anti mouse to after point sample in step 2 to microlens array chip, this horseradish peroxidase-labeled sheep anti mouse adopts Wuhan doctor's moral company BA1050,25 DEG C hatch 60min after, PBST washs 6 times;
B) GSS2400 chemiluminescence scanner scanning is used to draw the image in conjunction with situation that can reflect supernatant and microlens array chip, as shown in Figure 6;
4, by pre-service supernatant mark horseradish peroxidase in step 1:
By in step 1, pretreated 20 parts of supernatant sodium periodate oxidizing process are marked upper horseradish peroxidase respectively.
5, use immunogene to carry out double-antibody sandwich reaction screening pairing situation to supernatant, then by checking, negative control confirms, native protein checking draws the pairing supernatant be applicable to, concrete steps are as follows:
A) immunogene used confining liquid to be diluted to 1 μ g/ml, add in the microlens array chip of point sample in step 2,25 DEG C of reaction 150min;
B) PBST washs 6 times;
C) the supernatant antibody that a kind of confining liquid dilutes corresponding step 4 horseradish peroxidase-labeled of 20 times is added in each hole, 25 DEG C of reaction 150min;
D) PBST washs 8 times;
E) use GSS2400 chemiluminescence scanner scanning, draw image;
F) according to analysis result, draw the mark supernatant numbering that can match, repeat the reaction of this mark supernatant, do negative control simultaneously, determine the authentic and valid of unpaired message;
H) for the pairing supernatant determined, carry out the evaluation of native protein (such as clinical serum), filter out the unpaired message of applicable clinical detection, as shown in Figure 7.
Claims (10)
1. a screening technique for hybridoma supernatant stage trace antibody, it is characterized in that, this screening technique comprises the steps:
(1) culture supernatant of the hybridoma formed after getting Fusion of Cells, by exchange buffering system after salting out method removal impurity, obtains pre-service supernatant;
(2) antibody in the pre-service supernatant obtained in step (1) is fixed to microlens array chip;
(3) add horseradish peroxidase-labeled sheep anti-mouse antibody to the microlens array chip utilization of step (2) to react, confirm antibody activity and the fixing situation on microlens array chip in supernatant;
(4) the pre-service supernatant mark horseradish peroxidase will obtained in step (1);
(5) immunogene is added in microlens array chip that step (2) processed, then the supernatant antibody adding the horseradish peroxidase mark that step (4) obtains carries out double-antibody sandwich reaction screening pairing situation respectively, then by checking, negative control confirms, native protein checking draws the pairing supernatant be applicable to.
2. the screening technique of hybridoma supernatant stage trace antibody according to claim 1, it is characterized in that, the concrete operation method of described step (1) is: the culture supernatant of the hybridoma formed after a) getting Fusion of Cells, the saturated ammonium sulfate solution of 80% ~ 150% of supernatant volume is added to described supernatant, 8 ~ 15h is precipitated under 2 ~ 8 DEG C of conditions, in high speed freezing centrifuge, after centrifuging, be precipitated thing; B) described sediment 0.4ml buffer solution, uses super filter tube centrifuging, obtains pre-service supernatant after repeating ultrafiltration 2 ~ 6 times.
3. the screening technique of hybridoma supernatant stage trace antibody according to claim 1, it is characterized in that, the concrete operation method of described step (2) is: after the pre-service supernatant spotting buffer that a) described step (1) obtains dilutes 5 ~ 20 times, point sample is to microlens array chip; B) wash plate 3 ~ 6 times with PBST after hatching 30 ~ 120min under 25 ~ 37 DEG C of conditions, add confining liquid, under 2 ~ 8 DEG C of conditions, hatch 8 ~ 15 hours.
4. the screening technique of hybridoma supernatant stage trace antibody according to claim 1, it is characterized in that, the concrete operation method of described step (3) is: microlens array chip a) obtained to described step (2) adds horseradish peroxidase-labeled sheep anti mouse, hatch 30 ~ 120min under 25 ~ 37 DEG C of conditions after, PBST washing 3 ~ 6 times; B) chemiluminescence scanner scanning is used to draw the image in conjunction with situation that can reflect supernatant and microlens array chip.
5. the screening technique of hybridoma supernatant stage trace antibody according to claim 1, it is characterized in that, the concrete operation method of described step (4) is: the upper horseradish peroxidase of supernatant sodium periodate oxidizing process mark after the pre-service obtain step (1).
6. the screening technique of hybridoma supernatant stage trace antibody according to claim 1, it is characterized in that, the concrete operation method of described step (5) is: a) immunogene confining liquid is diluted to 0.1 ~ 1 μ g/ml, add in the micropore of the microlens array chip that described step (2) processed, under 25 ~ 37 DEG C of conditions, hatch 60 ~ 150min; B) PBST washing 2 ~ 6 times; C) add in the micropore of microlens array chip with the mark supernatant that processes of step (3) of confining liquid 5 ~ 20 times dilution, hatch 60 ~ 150min for 25-37 DEG C; D) PBST washing 5 ~ 8 times; E) use chemiluminescence scanner scanning, draw image; F) according to image, microlens array chip there is the supernatant corresponding to the selecting of light signal be the supernatant that can match; G) the pairing supernatant that draws repeat in step (5) a) ~ e) reaction, simultaneously carry out step a) time increase and replace immunogene to carry out the negative control reacted with confining liquid; When negative control microlens array chip is without light signal, and when immunogene reaction has a signal, determine the authentic and valid of unpaired message; H) for the pairing supernatant filtered out, use native protein replace immunogene repeat in step (5) a) ~ e) reaction, the numerical value and the clinical detection concentration linear fit that obtain optical signals are evaluated, and filter out the pairing supernatant information of the most applicable clinical detection.
7. the screening technique of hybridoma supernatant stage trace antibody according to claim 2, it is characterized in that, in described step (1), the centrifugal force of centrifuging operation is 10000 ~ 15000g, and working time is 10 ~ 30min.
8. the screening technique of hybridoma supernatant stage trace antibody according to claim 2, it is characterized in that, the molecular cut off of described super filter tube is less than the molecular weight of antibody in supernatant.
9. the screening technique of hybridoma supernatant stage trace antibody according to claim 3, it is characterized in that, described spotting buffer is the potpourri of carbonate and 30% glycerine, and the concentration of described carbonate is 0.05mol/L.
10. the screening technique of hybridoma supernatant stage trace antibody according to claim 6, it is characterized in that, described confining liquid is phosphate buffer and bovine serum albumin mix, the concentration of this phosphate buffer is 0.1mol/L, pH is 7.2, and the mass volume ratio of bovine serum albumin(BSA) and phosphate buffer is 1:100 ~ 5:100.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310154372.5A CN103808922B (en) | 2013-04-27 | 2013-04-27 | A kind of screening technique of hybridoma supernatant stage trace antibody |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310154372.5A CN103808922B (en) | 2013-04-27 | 2013-04-27 | A kind of screening technique of hybridoma supernatant stage trace antibody |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN103808922A CN103808922A (en) | 2014-05-21 |
| CN103808922B true CN103808922B (en) | 2015-09-09 |
Family
ID=50705996
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310154372.5A Active CN103808922B (en) | 2013-04-27 | 2013-04-27 | A kind of screening technique of hybridoma supernatant stage trace antibody |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN103808922B (en) |
Families Citing this family (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN104062431A (en) * | 2014-07-01 | 2014-09-24 | 上海理工大学 | Method for screening monoclonal antibody hybridoma by utilization of visual antigen chip |
| CN104034899B (en) * | 2014-07-01 | 2015-12-30 | 上海理工大学 | The grand array of a kind of food-borne pathogens hybridoma rapid screening antigen |
| CN108700576B (en) * | 2017-01-20 | 2021-07-02 | 深圳市新产业生物医学工程股份有限公司 | Method and kit for preparing antibody pair, application of kit and system for preparing antibody pair |
| CN110484974A (en) * | 2019-07-19 | 2019-11-22 | 中国农业科学院郑州果树研究所 | A kind of preparation method of grape antibody library |
| CN111812327A (en) * | 2020-06-05 | 2020-10-23 | 武汉华美生物工程有限公司 | Screening method of monoclonal antibody cells expressing secretion recognition natural samples |
| CN112611875A (en) * | 2020-12-29 | 2021-04-06 | 珠海碳云智能科技有限公司 | Method for screening polypeptide for detecting target antibody and application of screened polypeptide |
| CN113219178A (en) * | 2021-05-12 | 2021-08-06 | 上海理工大学 | Trace antibody rapid pairing immune test paper and pairing method thereof |
Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003035824A1 (en) * | 2001-10-25 | 2003-05-01 | Bar-Ilan University | Interactive transparent individual cells biochip processor |
| CN1446923A (en) * | 2002-03-22 | 2003-10-08 | 阎小君 | Method for filtering hybridoma by using peptide chips with high flux |
| CN1659185A (en) * | 2002-04-17 | 2005-08-24 | 欧洲分子生物学实验室 | Method for producing monoclonal antibodies |
| CN1763530A (en) * | 2004-10-18 | 2006-04-26 | 上海生物芯片有限公司 | Polypeptide chip and preparation process thereof, and method for preparing monoclonal antibody group using the same |
| CN1880447A (en) * | 2005-06-17 | 2006-12-20 | 上海生物芯片有限公司 | Process for preparing high-flux monoclonal antibody |
| WO2007019024A2 (en) * | 2005-08-11 | 2007-02-15 | Sru Biosystems, Inc. | Grating-based sensor combining label-free binding detection and fluorescence amplification and readout system for sensor |
| CN102608032A (en) * | 2012-04-10 | 2012-07-25 | 无锡国盛精密模具有限公司 | Integrated micro lens array device |
| CN202886259U (en) * | 2012-04-10 | 2013-04-17 | 无锡国盛精密模具有限公司 | Integrated microlens array device |
-
2013
- 2013-04-27 CN CN201310154372.5A patent/CN103808922B/en active Active
Patent Citations (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2003035824A1 (en) * | 2001-10-25 | 2003-05-01 | Bar-Ilan University | Interactive transparent individual cells biochip processor |
| CN1446923A (en) * | 2002-03-22 | 2003-10-08 | 阎小君 | Method for filtering hybridoma by using peptide chips with high flux |
| CN1659185A (en) * | 2002-04-17 | 2005-08-24 | 欧洲分子生物学实验室 | Method for producing monoclonal antibodies |
| CN1763530A (en) * | 2004-10-18 | 2006-04-26 | 上海生物芯片有限公司 | Polypeptide chip and preparation process thereof, and method for preparing monoclonal antibody group using the same |
| CN1880447A (en) * | 2005-06-17 | 2006-12-20 | 上海生物芯片有限公司 | Process for preparing high-flux monoclonal antibody |
| WO2007019024A2 (en) * | 2005-08-11 | 2007-02-15 | Sru Biosystems, Inc. | Grating-based sensor combining label-free binding detection and fluorescence amplification and readout system for sensor |
| CN102608032A (en) * | 2012-04-10 | 2012-07-25 | 无锡国盛精密模具有限公司 | Integrated micro lens array device |
| CN202886259U (en) * | 2012-04-10 | 2013-04-17 | 无锡国盛精密模具有限公司 | Integrated microlens array device |
Non-Patent Citations (3)
| Title |
|---|
| High-content analysis of single cells directly assembled on CMOS sensor based on color imaging;Tsuyoshi Tanaka et al;《Biosensor and Bioelectronics》;20101215;第26卷(第4期);1460-1465 * |
| 一种用于筛选杂交瘤上清的改良斑点免疫结合试验;陈华青等;《国外医学免疫分册》;19920229(第1期);50-51 * |
| 斑点免疫渗滤板批量筛选杂交瘤培养上清中的抗体;徐兵等;《上海免疫学杂志》;19961031;第16卷(第5期);308-309 * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN103808922A (en) | 2014-05-21 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN103808922B (en) | A kind of screening technique of hybridoma supernatant stage trace antibody | |
| CN102735851B (en) | Mycoplasma hyopneumoniae multi-recombination antigen ELISA (enzyme-linked immunosorbent assay) detection kit | |
| AU695419B2 (en) | Method for detecting antibodies | |
| CN101968490A (en) | Indirect ELISA (Enzyme-Linked Immunosorbent Assay) method and kit for detecting haemophilus parasuis antibodies | |
| CN105651999A (en) | Molybdenum disulfide-based sensor and preparation method and application thereof | |
| CN102747040A (en) | Anti-influenza A virus nucleoprotein monoclonal antibody, its preparation and application | |
| CN103134931A (en) | Double antibody sandwich method of detecting staphylococcus aureus enterotoxin A of food | |
| CN102175853A (en) | ELISA (enzyme linked immunosorbent assay) kit for detecting pig progenitive and respiratory syndrome (PRRS) antibody | |
| CN102876634B (en) | PMSG (pregnant mare serum gonadotropin) double-antibody sandwich ELISA (enzyme-linked immunosorbent assay) kit | |
| CN101609097A (en) | The competitive enzyme-linked immune detection method of EV71 neutralizing antibody, kit or reagent and preparation method thereof | |
| CN103739675B (en) | Strawberry veinbanding virus antibody and antigenic peptide, immunogen and application | |
| CN103364552A (en) | ELISA (enzyme-linked immuno sorbent assay) detection kit for porcine parvovirus IgM (immunoglobulin m) antibodies as well as preparation method and application of ELISA detection kit | |
| CN105242044A (en) | Test method for hog cholera virus E2 protein quantification | |
| CN102539776A (en) | Enzyme-linked immunosorbent assay method for enterovirus 71-specific antibody | |
| CN105223354A (en) | Aleutian Mink Disease Parvovirus colloidal gold immunochromatographydetection detection test paper bar and preparation method thereof | |
| CN101936998A (en) | Antirabies virus IgG (Immunoglobulin G) antibody ELISA (Enzyme Linked Immunosorbent Assay) detection kit for dogs | |
| CN104422771B (en) | Detect monoclonal antibody and the kit thereof of tick-brone encephalitis virus | |
| CN103364551A (en) | ELISA (enzyme-linked immuno sorbent assay) detection kit for porcine reproductive and respiratory syndrome virus IgM (immunoglobulin m) antibodies as well as preparation method and application of ELISA detection kit | |
| CN105567643B (en) | Hybridoma cell capable of secreting anti-EV 71 virus protein VP1 monoclonal antibody, monoclonal antibody and application | |
| CN105131122A (en) | Preparation and applications of monoclonal antibodies of polychlorinated biphenyl homologs | |
| CN1851463B (en) | Method for detecting horse arteritis virus by indirect enzyme-linked immunosorbent test | |
| CN102621319B (en) | Colloidal gold quick diagnosis test paper for distinguishing classic porcine reproductive and respiratory syndrome (PRRS) from highly pathogenic PRRS (HPRRS) | |
| CN103420879B (en) | Dapsone hapten and its preparation method and application | |
| CN102778568A (en) | Preparation and application of total antibody ELISA (enzyme linked immunosorbent assay) kit for detecting fever accompanied by thrombocytopenia syndrome virus | |
| CN104614519A (en) | Soybean peroxidase labelled chemiluminescence immunoassay kit, as well as use method and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant |