CN113968905A - Bovine serum albumin and application thereof - Google Patents
Bovine serum albumin and application thereof Download PDFInfo
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- CN113968905A CN113968905A CN202111255203.1A CN202111255203A CN113968905A CN 113968905 A CN113968905 A CN 113968905A CN 202111255203 A CN202111255203 A CN 202111255203A CN 113968905 A CN113968905 A CN 113968905A
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- 108091003079 Bovine Serum Albumin Proteins 0.000 title claims abstract description 97
- 229940098773 bovine serum albumin Drugs 0.000 title claims abstract description 95
- 238000001514 detection method Methods 0.000 claims abstract description 46
- 238000007789 sealing Methods 0.000 claims abstract description 25
- 239000000427 antigen Substances 0.000 claims abstract description 20
- 102000036639 antigens Human genes 0.000 claims abstract description 20
- 108091007433 antigens Proteins 0.000 claims abstract description 20
- 239000002158 endotoxin Substances 0.000 claims abstract description 6
- 239000007788 liquid Substances 0.000 claims description 32
- 241000283073 Equus caballus Species 0.000 claims description 16
- 230000000903 blocking effect Effects 0.000 claims description 16
- 239000000126 substance Substances 0.000 claims description 12
- 208000004668 avian leukosis Diseases 0.000 claims description 8
- 206010000210 abortion Diseases 0.000 claims description 6
- 231100000176 abortion Toxicity 0.000 claims description 6
- 102000004169 proteins and genes Human genes 0.000 claims description 6
- 108090000623 proteins and genes Proteins 0.000 claims description 6
- 102000009027 Albumins Human genes 0.000 claims description 5
- 108010088751 Albumins Proteins 0.000 claims description 5
- 238000002835 absorbance Methods 0.000 claims description 5
- 208000007212 Foot-and-Mouth Disease Diseases 0.000 claims description 4
- 241000710198 Foot-and-mouth disease virus Species 0.000 claims description 4
- 241001529856 Celsia Species 0.000 claims description 3
- 208000032839 leukemia Diseases 0.000 claims 1
- 230000000694 effects Effects 0.000 abstract description 11
- 230000035945 sensitivity Effects 0.000 abstract description 9
- 230000004044 response Effects 0.000 abstract description 2
- 210000002966 serum Anatomy 0.000 description 42
- 239000000243 solution Substances 0.000 description 27
- 238000002965 ELISA Methods 0.000 description 20
- 239000000523 sample Substances 0.000 description 15
- 239000011248 coating agent Substances 0.000 description 13
- 238000000576 coating method Methods 0.000 description 13
- 238000005406 washing Methods 0.000 description 8
- 238000010790 dilution Methods 0.000 description 7
- 239000012895 dilution Substances 0.000 description 7
- 238000001035 drying Methods 0.000 description 7
- 238000007865 diluting Methods 0.000 description 6
- 239000003085 diluting agent Substances 0.000 description 6
- 238000002360 preparation method Methods 0.000 description 6
- UQLDLKMNUJERMK-UHFFFAOYSA-L di(octadecanoyloxy)lead Chemical compound [Pb+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O UQLDLKMNUJERMK-UHFFFAOYSA-L 0.000 description 5
- 102000004190 Enzymes Human genes 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 4
- 239000012888 bovine serum Substances 0.000 description 4
- 238000004519 manufacturing process Methods 0.000 description 4
- 238000000034 method Methods 0.000 description 4
- 238000002156 mixing Methods 0.000 description 4
- 239000012089 stop solution Substances 0.000 description 4
- 241000283690 Bos taurus Species 0.000 description 3
- 241001198387 Escherichia coli BL21(DE3) Species 0.000 description 3
- 241000710202 Foot-and-mouth disease virus - type A Species 0.000 description 3
- 241000144531 Foot-and-mouth disease virus - type Asia 1 Species 0.000 description 3
- 241000607142 Salmonella Species 0.000 description 3
- 201000010099 disease Diseases 0.000 description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 241001465754 Metazoa Species 0.000 description 2
- 241001494479 Pecora Species 0.000 description 2
- QAOWNCQODCNURD-UHFFFAOYSA-N Sulfuric acid Chemical compound OS(O)(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-N 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 230000008676 import Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 238000004806 packaging method and process Methods 0.000 description 2
- 239000013641 positive control Substances 0.000 description 2
- 239000000758 substrate Substances 0.000 description 2
- 241000712461 unidentified influenza virus Species 0.000 description 2
- 238000009461 vacuum packaging Methods 0.000 description 2
- 241000700664 Capripoxvirus Species 0.000 description 1
- 241000710777 Classical swine fever virus Species 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- 208000004232 Enteritis Diseases 0.000 description 1
- 241000230501 Equine herpesvirus sp. Species 0.000 description 1
- 241000713730 Equine infectious anemia virus Species 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 241000764238 Isis Species 0.000 description 1
- 208000031662 Noncommunicable disease Diseases 0.000 description 1
- 208000030852 Parasitic disease Diseases 0.000 description 1
- 241001135549 Porcine epidemic diarrhea virus Species 0.000 description 1
- 241001135989 Porcine reproductive and respiratory syndrome virus Species 0.000 description 1
- 241000607729 Salmonella enterica subsp. enterica serovar Abortus Species 0.000 description 1
- 241001632234 Senecavirus Species 0.000 description 1
- 241001428894 Small ruminant morbillivirus Species 0.000 description 1
- 241000701093 Suid alphaherpesvirus 1 Species 0.000 description 1
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- 239000013062 quality control Sample Substances 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 235000020183 skimmed milk Nutrition 0.000 description 1
- 239000003381 stabilizer Substances 0.000 description 1
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- 238000003239 susceptibility assay Methods 0.000 description 1
- 229960005486 vaccine Drugs 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/76—Albumins
- C07K14/765—Serum albumin, e.g. HSA
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- 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
- G01N33/5306—Improving reaction conditions, e.g. reduction of non-specific binding, promotion of specific binding
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- 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
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56983—Viruses
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Abstract
The invention relates to the technical field of biology, and particularly relates to bovine serum albumin and application thereof. The purity of the bovine serum albumin BSA is more than or equal to 90 percent; the content of endotoxin is less than or equal to 5 EU/ml. The molecular weight of the bovine serum albumin is 68 +/-10 kDa. The bovine serum albumin is used for a sealing solution of an ELSIA antibody and/or antigen detection kit. The invention has the advantages that: (1) high sensitivity and good specificity. (2) The sealing effect is good, and the specificity is high. (3) The batch-to-batch variation is low. (4) There was no positive response to negative standards.
Description
Technical Field
The invention relates to the technical field of biology, and particularly relates to bovine serum albumin and application thereof.
Background
Enzyme-linked immunosorbent assay (ELISA) is a bioactive substance micro-measurement technology first reported and established by Engvall and Perlman, and is successfully applied to immunodiagnosis of infectious diseases, parasitic diseases, non-infectious diseases and the like caused by various pathogenic microorganisms, and quantitative measurement of thousand macromolecular antigens and small molecular antigens. The method is not only suitable for checking thousand clinical specimens, but also suitable for thousand serum epidemiological investigation, and is widely applied to the field of life science due to the advantages of high sensitivity, good specificity and the like
In recent years, many research reports about establishment of ELISA detection methods are reported at home and abroad, but few commercial kits are available, especially at home. The main reason for this is that the detection method does not achieve the desired sensitivity and specificity. Although the ELISA detection method has simple operation steps, the influence factors are more, and the blocking is one of the most important influence factors. Improper sealing can directly affect the sensitivity and specificity of an ELISA detection method, and false positive results are generated, so that misdiagnosis of diseases is caused, and adverse results are caused; in addition, the good sealing liquid can also play the roles of a stabilizer and a protective agent, so that the storage life of the ELISA plate is improved; conversely, poor confining liquids can also interfere with the reaction of the various reagents. Therefore, the selection of the optimal blocking solution is one of the keys for improving the sensitivity and the specificity of the ELISA detection method, and has an important effect on establishing a good ELISA detection method.
The confining liquid is one of important raw materials for manufacturing the kit for detecting the animal epidemic disease, and the main function is to reduce the non-specific reaction of the kit. There are many materials that are used for sealing, such as: horse serum, Bovine Serum Albumin (BSA), skim milk, and commercial blocking solutions, etc., wherein BSA is most commonly used. Bovine Serum Albumin (BSA) is an important animal-derived material and is widely applied to vaccine manufacture, detection kits and production and manufacture of molecular biological reagents.
However, BSA which is specially aimed at the quality of the detection kit does not exist, so that when the BSA is used for producing the detection kit, the sealing effect difference is large, the quality of the kit is unstable, and the commercialized sealing solution has the problems of large batch difference, unstable supply and the like. In addition, the current BAS standard can only meet the universality requirement, but cannot well meet the individual requirement. When the detection kit for epidemic diseases is prepared, BSA sold in the market at present cannot meet the requirements, and the method is mainly characterized in that the kit has poor blocking effect, shows positive reaction on negative standard substances, has low reaction value on positive standard substances, and causes the detection result of the kit to have deviation and poor repeatability.
Disclosure of Invention
The invention aims to provide bovine serum albumin which has good sealing effect, high sensitivity and good specificity and does not have positive reaction to a negative standard substance and application thereof.
In order to achieve the purpose, the invention adopts the technical scheme that:
bovine Serum Albumin (BSA) with the purity of more than or equal to 90 percent; the content of endotoxin is less than or equal to 5 EU/ml.
Further, the molecular weight of the bovine serum albumin is 68 +/-10 kDa.
Further, bovine serum albuminPreparing to 5% concentration with PBS, clarifyingThe clarity is less than or equal to 0.2 percent.
Furthermore, the clarity of the bovine serum albumin solution is that the 403nm absorbance value of the 1% albumin solution is less than or equal to 0.2%.
Further, the bovine serum albumin is detected by an avian leukosis P26 protein coated plate, the OD value of a diluted 12-fold sensitive standard substance is more than or equal to 1.0, and the OD value of a negative sample is less than or equal to 0.06.
The invention also provides application of the bovine serum albumin.
The application of bovine serum albumin is to a confining liquid of an ELSIA antibody and/or antigen detection kit.
Furthermore, the bovine serum albumin is used for a confining liquid of an avian leukosis cELSIA antigen detection kit, a confining liquid of a foot-and-mouth disease competition ELSIA antibody detection kit and a confining liquid of a equine abortion indirect ELSIA antibody detection kit.
Further, the concentration of the bovine serum albumin in a sealing solution of the ELSIA antibody and/or antigen detection kit is 0.5-1%.
Further, the bovine serum albumin is used as a blocking solution of an ELSIA antibody and/or antigen detection kit, and is 5% BSA solution prepared by 10mM PBS.
The bovine serum albumin provided by the invention is used for detecting BSA (bovine serum albumin) used for coating of the kit. The enclosed coated plate prepared from the bovine serum albumin provided by the invention has low detection result on sensitive serum. The coated plate sealed by the sealing liquid prepared by the bovine serum albumin has high detection result on specific serum.
The bovine serum albumin provided by the invention is used for sealing the ELSIA antibody and/or antigen detection kit, and the sealing effect is good.
The ELISA plate is prepared by using avian leukosis P26 monoclonal antibody, the confining liquid prepared by using the bovine serum albumin provided by the invention is closed, the OD value of a diluted 12-fold sensitive standard substance is more than or equal to 1.0, and the OD value of a negative sample is less than or equal to 0.06.
The kit prepared by the bovine serum albumin provided by the invention has the batch difference of less than or equal to 5 percent.
When the BSA provided by the invention is applied to the sealing process of other kits, compared with other commercial sealing solutions, the OD value of a sensitive quality control sample is higher than 10%, and for a negative sample, the OD value is less than 0.1, so that the sealing effect is good, and the specificity of the kit is improved by 10%.
Compared with the prior art, the bovine serum albumin and the application thereof provided by the invention have the advantages that:
(1) high sensitivity and good specificity.
(2) The sealing effect is good, and the specificity is high.
(3) The batch-to-batch variation is low.
(4) There was no positive response to negative standards.
Detailed Description
In order to make those skilled in the art better understand the technical solution of the present invention, the following examples further describe the present invention in detail, and the following examples are only used for illustrating the present invention, but not for limiting the scope of the present invention.
Bovine Serum Albumin (BSA) with the purity of more than or equal to 90 percent; the content of endotoxin is less than or equal to 5 EU/ml.
Further, the molecular weight of the bovine serum albumin is 68 +/-10 kDa.
Further, the bovine serum albuminThe white was adjusted to 5% concentration with PBSThe clarity is less than or equal to 0.2 percent.
Furthermore, the clarity of the bovine serum albumin solution is that the 403nm absorbance value of the 1% albumin solution is less than or equal to 0.2%.
Further, the bovine serum albumin is detected by an avian leukosis P26 protein coated plate, the OD value of a diluted 12-fold sensitive standard substance is more than or equal to 1.0, and the OD value of a negative sample is less than or equal to 0.06.
The invention also provides application of the bovine serum albumin.
The application of bovine serum albumin is to a confining liquid of an ELSIA antibody and/or antigen detection kit.
Furthermore, the bovine serum albumin is used for a confining liquid of an avian leukosis cELSIA antigen detection kit, a confining liquid of a foot-and-mouth disease competition ELSIA antibody detection kit and a confining liquid of a equine abortion indirect ELSIA antibody detection kit.
Further, the concentration of the bovine serum albumin in a sealing solution of the ELSIA antibody and/or antigen detection kit is 0.5-1%.
Further, the bovine serum albumin is used as a confining liquid of an ELSIA antibody and/or antigen detection kit, and the confining liquid isIs 10mM 5% BSA in PBS.
Example 1
Bovine serum albumin, wherein the purity of the bovine serum albumin BSA is 90%; the endotoxin content was 4 EU/ml. The molecular weight of the bovine serum albumin is 68 kDa.
The clarity of the bovine serum albumin solution is 1% albumin solution 403nm absorbance value, 0.09%.
The bovine serum albumin is detected by an avian leukosis P26 protein coated plate, the OD value of a diluted 12-fold sensitive standard substance is more than or equal to 1.0, and the OD value of a negative sample is less than or equal to 0.06.
Example 2
Bovine serum albumin, wherein the purity of the bovine serum albumin BSA is 95%; the endotoxin content was 2 EU/ml. The molecular weight of the bovine serum albumin is 68 kDa.
The clarity of the bovine serum albumin solution is 1% albumin solution 403nm absorbance value, 0.09%.
The bovine serum albumin is detected by an avian leukosis P26 protein coated plate, the OD value of a diluted 12-fold sensitive standard substance is more than or equal to 1.0, and the OD value of a negative sample is less than or equal to 0.06.
Example 3
Preparation of ELSIA antibody detection kit for foot-and-mouth disease competition
1. Preparation of coated plates
1.1 preparation of coating antibody: the purified monoclonal antibody 3D9 strain was diluted with coating solution to working concentration and mixed well with a stirrer (at least 10 minutes, taking care not to generate excessive vortexes).
1.2 coating: taking the uniformly mixed coating antibody, incubating overnight (16-20 hours) at the temperature of 4 ℃ in a hole of 100 mul;
1.3 washing the plate: the overnight coated ELISA plate was removed, washed 3 times with PBST, 300. mu.l/well and patted dry;
1.4 addition of antigen: diluting antigen 40 times with PBST, mixing well, adding 100 μ l/well into ELISA plate, incubating for 1.5 hr at 37 deg.C;
1.5 plate washing: the coated plate was removed, washed 3 times with PBST, 300. mu.l/well and patted dry.
1.6 sealing: the prepared coated plate was added with bovine serum albumin provided by the present invention, BSA1# (commercially available from China), BSA2# (commercially available from China) and BSA3# (commercially available from import) to prepare blocking solutions, each of which was 150. mu.l/well, and blocked at 37 ℃ for 1.5 hours.
1.7 airing and packaging: and pouring liquid in the holes of the sealed ELISA plate, drying the ELISA plate by using a drying barrel, putting the ELISA plate on a clean table for drying, keeping the ELISA plate on the table for at least 1 half hour, and carrying out vacuum packaging.
2. Detection of sensitivity
2.1 treatment of the samples: taking a pig serum standard substance, and respectively diluting the pig serum standard substance by 16 times, 32 times, 64 times, 128 times, 256 times, 512 times and 1024 times by using a sample diluent. (taking 25. mu.l serum to add 375. mu.l dilution, namely 16-fold dilution; taking 200. mu.l 16-fold dilution to add 200. mu.l dilution, namely 32-fold dilution; and so on)
2.2 sample adding: taking a detachable coating plate, adding 50 mu l of the diluted serum sample into each hole, simultaneously adding 2 holes of negative and positive control serum, immediately adding 50 mu l of enzyme-labeled antibody, and uniformly mixing;
2.3 incubation: sealing the coated plate, and placing in a constant temperature incubator at 37 deg.C for 60 min;
2.4 washing: discarding liquid in the hole, adding 300 μ l of washing solution into each hole, washing for 5 times, and drying each time;
2.5 color development: adding 50 mu l of substrate solution into each hole, and incubating for 20 minutes at 37 ℃ in the dark;
2.6 termination: adding 50 mu L of stop solution (2mol/L sulfuric acid solution) into each hole, slightly shaking and mixing uniformly, and reading the OD450nm value at the wavelength of 450nm (the reading should be completed within 10 minutes after the stop solution is added);
2.7 results:
the results are detailed in table 1.
The test result shows that: the detection result of the coated plate closed by the blocking liquid prepared by the bovine serum albumin provided by the invention on sensitive serum is lower in each dilution than that of the coating liquid prepared by other BSA, and the blocking effect is better than that of BSA of other brands. The effect of the BSA provided by the invention in a control is obviously better than that of other BSA.
TABLE 1 sensitivity assay results (OD 450nm)
3. Specificity detection
3.1 dilution of the sample: using the sample diluent to treat porcine epidemic diarrhea virus positive serum, porcine reproductive and respiratory syndrome virus positive serum, porcine pseudorabies virus positive serum, hog cholera virus positive serum, seneca virus positive serum, bovine epidemic heat positive serum, peste des petits ruminants virus positive serum, capripox virus positive serum, negative porcine serum and negative bovine serum, diluting 1:32 times by using negative sheep serum, escherichia coli BL21(DE3) positive pig serum, escherichia coli BL21(DE3) positive bovine serum, escherichia coli BL21(DE3) positive sheep serum, foot-and-mouth disease virus A type positive serum (ovine source), foot-and-mouth disease virus A type positive serum (bovine source), foot-and-mouth disease virus A type positive serum (porcine source), foot-and-mouth disease virus Asia 1 type positive serum (ovine source), foot-and-mouth disease virus Asia 1 type positive serum (bovine source) and foot-and-mouth disease virus Asia 1 type positive serum (porcine source);
3.2, detection: performing the treatment according to 2.2-2.6 items;
3.3 results:
the results are detailed in table 2.
The test result shows that: the detection results of the coating plate closed by the blocking liquid prepared by the bovine serum albumin provided by the invention on specific serum are all higher than those of the coating liquid prepared by other BSA, and the blocking effect is better than that of other BSA. The effect of the BSA provided by the invention in a control is obviously better than that of other BSA.
TABLE 2 detection results (OD 450nm) for specific serum samples
Example 4
Preparation of indirect ELSIA antibody detection kit for equine abortion
1. Preparation of coated plates
1.1 preparation of coating antigen: the purified Salmonella abortus equine protein was diluted to working concentration with coating solution and mixed well with a stirrer (at least 10 minutes, taking care not to generate excessive vortexes).
1.2 coating: taking the uniformly mixed envelope antigen, incubating at 100 mu l/hole for overnight (16-20 hours) at 4 ℃;
1.3 washing the plate: the overnight coated ELISA plate was removed, washed 3 times with PBST, 300. mu.l/well and patted dry;
1.4 sealing: the prepared coated plate was added with bovine serum albumin provided by the present invention, BSA1# (commercially available from China), BSA2# (commercially available from China) and BSA3# (commercially available from import) to prepare blocking solutions, each of which was 150. mu.l/well, and blocked at 37 ℃ for 2 hours. Adding sealing liquid, sealing at 150 μ l/hole for 2 hr at 37 deg.C;
1.5 airing and packaging: and pouring liquid in the holes of the sealed ELISA plate, drying the ELISA plate by using a drying barrel, putting the ELISA plate on a clean table for drying, keeping the ELISA plate on the table for at least 1 half hour, and carrying out vacuum packaging.
2. Detection of sensitivity
2.1 treatment of the samples: taking positive serum of the equine abortion salmonella, and diluting the positive serum by 1: 100-1: 3200 times respectively by using sample diluent.
2.2 sample adding: taking a detachable coating plate, adding 100 mu l of the diluted serum sample into each hole, simultaneously adding 2 holes of negative and positive control serum, 100 mu l/hole, and incubating for 45 minutes in a constant-temperature incubator at 37 ℃;
2.3 washing: the liquid was discarded and wash solution (PBST) was added at 300. mu.l/well and washed 4 times, each time patted dry.
2.4 addition of enzyme-labeled antibody: diluting the horse-resistant enzyme labeled antibody with a diluent (Hubei Yingchuang HRP diluent) according to a ratio of 1: 30000 times, adding the diluted horse-resistant enzyme labeled antibody into a 96-well plate at a concentration of 100 mu l/well, and incubating for 30 minutes at 37 ℃. (ii) a
2.5 washing: the same as 2.3;
2.6 addition of substrate solution: 100 μ l/well, incubated at 37 ℃ for 10 min in the absence of light;
2.7 adding stop solution: after mixing well with gentle shaking at 50. mu.l/well, the reading was done at a wavelength of 450nm (the reading should be done within 5 minutes after addition of stop solution) and the results were recorded.
2.8 results:
the results are shown in Table 3.
The test result shows that: the blocking liquid prepared from the bovine serum albumin provided by the invention has a good blocking effect, and is superior to other BSA. The effect of the BSA provided by the invention in a control is obviously better than that of other BSA.
TABLE 3 detection of sensitive sera (OD 450nm)
3. Specificity detection
3.1 treatment of the samples: taking equine herpesvirus positive serum, equine escherichia coli positive serum, equine influenza virus H3 subtype positive serum, equine influenza virus H7 subtype positive serum, equine infectious anemia virus positive serum, equine dublin salmonella, equine typhimurium positive serum, equine enteritis salmonella positive serum and equine abortion negative serum, and diluting by 1:200 times with a sample diluent.
3.2, detection: the method is carried out according to the items 2.2 to 2.7 of the present examples.
3.3 results:
the results are shown in Table 4.
The detection result shows that: the blocking liquid prepared from the bovine serum albumin provided by the invention has a good blocking effect, is superior to other BSA (bovine serum albumin), and has an OD (optical density) value not higher than 0.1. The effect of the BSA provided by the invention in a control is obviously better than that of other BSA.
TABLE 4 specific assay results (OD 450nm)
The preferred embodiments of the present invention have been described in detail, however, the present invention is not limited to the specific details of the above embodiments, and various changes may be made to the technical solution of the present invention within the technical idea of the present invention, and these simple modifications are included in the protective scope of the present invention.
It should be noted that, in the foregoing embodiments, various specific technical features and steps described in the above embodiments can be combined in any suitable manner, and in order to avoid unnecessary repetition, various possible combinations of the features and steps are not described separately.
In addition, any combination of the various embodiments of the present invention is also possible, and the same should be considered as the disclosure of the present invention as long as it does not depart from the spirit of the present invention.
Claims (9)
1. A bovine serum albumin, characterized by: the purity of the bovine serum albumin BSA is more than or equal to 90 percent; the content of endotoxin is less than or equal to 5 EU/ml.
2. The bovine serum albumin of claim 1, wherein: the molecular weight of the bovine serum albumin is 68 +/-10 kDa.
3. The bovine serum albumin of claim 1, wherein: the bovine serum albumin is prepared by PBS to have a concentration of 5% and the clarity of the bovine serum albumin is less than or equal to 0.2%.
4. The bovine serum albumin of claim 1, wherein: the clarity of the bovine serum albumin solution is that the 403nm absorbance value of 1% albumin solution is less than or equal to 0.2%.
5. The bovine serum albumin of claim 1, wherein: the bovine serum albumin is detected by an avian leukosis P26 protein coated plate, the OD value of a diluted 12-fold sensitive standard substance is more than or equal to 1.0, and the OD value of a negative sample is less than or equal to 0.06.
6. The application of bovine serum albumin is to a confining liquid of an ELSIA antibody and/or antigen detection kit.
7. The use of bovine serum albumin according to claim 6, wherein: the bovine serum albumin is used for sealing liquid of a fowl leukemia cELSIA antigen detection kit, sealing liquid of a foot-and-mouth disease competition ELSIA antibody detection kit and sealing liquid of a equine abortion indirect ELSIA antibody detection kit.
8. The use of bovine serum albumin according to claim 6, wherein: the bovine serum albumin is used for the confining liquid of the ELSIA antibody and/or antigen detection kit, and the concentration of the confining liquid is 0.5-1%.
9. Use of a bovine serum albumin according to any one of claims 6 to 10, wherein: the bovine serum albumin is used as a blocking solution of an ELSIA antibody and/or antigen detection kit, and is a solution with BSA concentration of 5% prepared by 10mM PBS.
Priority Applications (1)
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