CN112083158A - Preparation method of bovine serum for confining liquid - Google Patents
Preparation method of bovine serum for confining liquid Download PDFInfo
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- CN112083158A CN112083158A CN202010890348.8A CN202010890348A CN112083158A CN 112083158 A CN112083158 A CN 112083158A CN 202010890348 A CN202010890348 A CN 202010890348A CN 112083158 A CN112083158 A CN 112083158A
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- 239000012888 bovine serum Substances 0.000 title claims abstract description 65
- 239000007788 liquid Substances 0.000 title claims abstract description 23
- 238000002360 preparation method Methods 0.000 title claims abstract description 9
- 238000001914 filtration Methods 0.000 claims abstract description 21
- 238000007789 sealing Methods 0.000 claims abstract description 19
- 239000012528 membrane Substances 0.000 claims abstract description 12
- 239000006228 supernatant Substances 0.000 claims abstract description 12
- 238000001042 affinity chromatography Methods 0.000 claims abstract description 11
- 230000001954 sterilising effect Effects 0.000 claims abstract description 8
- 238000001556 precipitation Methods 0.000 claims abstract description 6
- 238000000034 method Methods 0.000 claims description 17
- 210000002966 serum Anatomy 0.000 claims description 13
- 230000000903 blocking effect Effects 0.000 claims description 11
- 238000004587 chromatography analysis Methods 0.000 claims description 8
- 229920002401 polyacrylamide Polymers 0.000 claims description 8
- 238000005352 clarification Methods 0.000 claims description 6
- 229920002684 Sepharose Polymers 0.000 claims description 5
- 244000309466 calf Species 0.000 claims description 5
- 239000000919 ceramic Substances 0.000 claims description 5
- 239000002131 composite material Substances 0.000 claims description 5
- 239000000706 filtrate Substances 0.000 claims description 5
- 238000005119 centrifugation Methods 0.000 claims description 4
- 238000004659 sterilization and disinfection Methods 0.000 claims description 2
- 239000012530 fluid Substances 0.000 claims 4
- 238000001514 detection method Methods 0.000 abstract description 9
- 238000008157 ELISA kit Methods 0.000 abstract description 2
- 108060003951 Immunoglobulin Proteins 0.000 description 24
- 102000018358 immunoglobulin Human genes 0.000 description 24
- 239000000243 solution Substances 0.000 description 19
- 239000000427 antigen Substances 0.000 description 12
- 102000036639 antigens Human genes 0.000 description 12
- 108091007433 antigens Proteins 0.000 description 12
- 108090000790 Enzymes Proteins 0.000 description 8
- 102000004190 Enzymes Human genes 0.000 description 8
- 108010086710 Hydroxylamine dehydrogenase Proteins 0.000 description 8
- 239000007790 solid phase Substances 0.000 description 8
- 210000000601 blood cell Anatomy 0.000 description 7
- 238000006243 chemical reaction Methods 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- 230000000052 comparative effect Effects 0.000 description 5
- 238000002965 ELISA Methods 0.000 description 4
- 238000005259 measurement Methods 0.000 description 4
- 239000002244 precipitate Substances 0.000 description 4
- 230000001376 precipitating effect Effects 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 241000283690 Bos taurus Species 0.000 description 3
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 108010088160 Staphylococcal Protein A Proteins 0.000 description 3
- 238000004440 column chromatography Methods 0.000 description 3
- 238000003756 stirring Methods 0.000 description 3
- 238000012360 testing method Methods 0.000 description 3
- 108010088751 Albumins Proteins 0.000 description 2
- 102000009027 Albumins Human genes 0.000 description 2
- 239000012091 fetal bovine serum Substances 0.000 description 2
- 239000002245 particle Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 102000004169 proteins and genes Human genes 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 239000000758 substrate Substances 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 102000004506 Blood Proteins Human genes 0.000 description 1
- 108010017384 Blood Proteins Proteins 0.000 description 1
- FRPHFZCDPYBUAU-UHFFFAOYSA-N Bromocresolgreen Chemical compound CC1=C(Br)C(O)=C(Br)C=C1C1(C=2C(=C(Br)C(O)=C(Br)C=2)C)C2=CC=CC=C2S(=O)(=O)O1 FRPHFZCDPYBUAU-UHFFFAOYSA-N 0.000 description 1
- 102000018071 Immunoglobulin Fc Fragments Human genes 0.000 description 1
- 108010091135 Immunoglobulin Fc Fragments Proteins 0.000 description 1
- 108010071390 Serum Albumin Proteins 0.000 description 1
- 102000007562 Serum Albumin Human genes 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 description 1
- 239000003929 acidic solution Substances 0.000 description 1
- 125000000129 anionic group Chemical group 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- OHJMTUPIZMNBFR-UHFFFAOYSA-N biuret Chemical compound NC(=O)NC(N)=O OHJMTUPIZMNBFR-UHFFFAOYSA-N 0.000 description 1
- 230000003197 catalytic effect Effects 0.000 description 1
- 238000004090 dissolution Methods 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 239000012894 fetal calf serum Substances 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 239000005556 hormone Substances 0.000 description 1
- 229940088597 hormone Drugs 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 229920002521 macromolecule Polymers 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000008055 phosphate buffer solution Substances 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 230000000750 progressive effect Effects 0.000 description 1
- 238000004451 qualitative analysis Methods 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 235000020183 skimmed milk Nutrition 0.000 description 1
- AJPJDKMHJJGVTQ-UHFFFAOYSA-M sodium dihydrogen phosphate Chemical compound [Na+].OP(O)([O-])=O AJPJDKMHJJGVTQ-UHFFFAOYSA-M 0.000 description 1
- 229910000162 sodium phosphate Inorganic materials 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
Classifications
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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
- G01N33/531—Production of immunochemical test materials
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- Health & Medical Sciences (AREA)
- Immunology (AREA)
- Life Sciences & Earth Sciences (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- Biomedical Technology (AREA)
- Chemical & Material Sciences (AREA)
- Hematology (AREA)
- Urology & Nephrology (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Cell Biology (AREA)
- Food Science & Technology (AREA)
- Medicinal Chemistry (AREA)
- Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Biochemistry (AREA)
- General Health & Medical Sciences (AREA)
- General Physics & Mathematics (AREA)
- Pathology (AREA)
- Investigating Or Analysing Biological Materials (AREA)
Abstract
The invention discloses a preparation method of bovine serum for confining liquid, which comprises the following steps: taking and pretreating mature bovine serum, sequentially carrying out deep filtration and precipitation on the pretreated mature bovine serum to obtain clarified mature bovine serum, centrifuging the clarified mature bovine serum, taking supernatant, carrying out affinity column chromatography on the obtained supernatant to obtain chromatographic solution, filtering the obtained chromatographic solution by using a 0.01-0.05 mu m filter membrane, and sterilizing to obtain bovine serum for sealing solution. The confining liquid prepared by the invention reduces the background value of the enzyme linked immunosorbent assay kit during detection and improves the detection limit.
Description
Technical Field
The invention relates to the technical field of immunology, in particular to a preparation method of bovine serum for confining liquid.
Background
The enzyme-linked immunosorbent assay is a common method for detecting antigen or antibody, and the basic principle is that the antigen or antibody is connected with certain enzyme to form enzyme-labeled antigen or antibody, and the enzyme-labeled antigen or antibody retains the immunological activity and the activity of the enzyme. In the measurement, the tested specimen (the antibody or antigen to be measured) and the enzyme-labeled antigen or antibody are reacted with the antigen or antibody on the surface of the solid phase carrier according to different steps, then the antigen-antibody complex formed on the solid phase carrier is separated from other substances by a washing method, and finally the amount of the enzyme bound on the solid phase carrier is in a certain proportion to the amount of the tested substance in the specimen. After the substrate of the enzyme reaction is added, the substrate is catalyzed by the enzyme to be changed into a colored product, and the amount of the product is directly related to the amount of the detected substance in the sample, so that qualitative or quantitative analysis can be carried out according to the shade of the color reaction. Because the catalytic frequency of the enzyme is very high, the reaction effect can be greatly amplified, so that the determination method achieves very high sensitivity.
Since the solid phase carrier in the kit can be bound to the antigen or antibody to be detected, in order to avoid the binding of the solid phase carrier to the antigen or antibody to be detected, a blocking solution needs to be used for the binding to the solid phase carrier. At present, although a blocking solution mainly comprises adult bovine serum, the adult bovine serum contains a large amount of immunoglobulin, and the immunoglobulin has an antibody with an immune function, and if the immunoglobulin exists in the blocking solution in a large amount, the result of quantitative and qualitative detection of an antigen or an antibody to be detected is influenced, so that the reduction of the amount of the immunoglobulin in the adult bovine serum is necessary.
Therefore, how to provide bovine serum for a sealing solution is a technical problem that needs to be solved urgently by those skilled in the art.
Disclosure of Invention
In view of the above, the invention adopts clarification, affinity column chromatography, filtration and other modes to remove immunoglobulin in fetal calf serum, reduces the background value of the enzyme linked immunosorbent assay kit during detection, and improves the detection limit.
In order to achieve the purpose, the invention adopts the following technical scheme:
1) pretreatment: taking adult bovine serum, and pretreating the adult bovine serum to obtain pretreated adult bovine serum;
2) clarification: sequentially carrying out deep filtration and precipitation on the pretreated mature bovine serum to obtain clarified mature bovine serum;
3) centrifuging: centrifuging the clarified adult calf serum, and taking the supernatant;
4) affinity column chromatography: performing affinity column chromatography on the obtained supernatant to obtain a chromatographic solution;
5) and (3) filtering: filtering the obtained chromatography liquid with 0.01-0.05 μm filter membrane, and sterilizing to obtain bovine serum for blocking liquid.
The technical effect achieved by the technical scheme is as follows: the adult cattle serum contains BSA protein, immunoglobulin, blood cells, hormone substances, ions, etc., and is subjected to deep filtration by a deep filter to remove macromolecular substances; after centrifugation, part of the macromolecular protein can be removed; the affinity column chromatography can make part of immunoglobulin stay in the column, because the immunoglobulin mainly includes IgG, IgM and IgA, their molecular weight is 150kd, 1000kd and 400kd respectively, and the molecular weight of BSA protein is between 60-75kd, so adopt 0.01-0.05 μm filter membrane to filter, can retain most of immunoglobulin on the membrane, so as to achieve the goal of removing immunoglobulin, thus, prevent the immunoglobulin from adsorbing reaction with antigen or antibody to be detected, have improved the detection limit of the kit.
As a preferred technical solution of the present invention, in step 1), the pretreatment is: the resultant bovine serum was centrifuged at 8000rpm for 10-15min by a continuous flow centrifuge to remove blood cells.
The technical effect achieved by the technical scheme is as follows: the continuous flow centrifuge has large treatment capacity, short time and low labor cost, can treat the blood cells in the serum of the adult cattle on a large scale, can precipitate after centrifugation, and achieves the aim of removing the blood cells.
In the preferable technical scheme of the invention, in the step 2), the pH value of the filtrate is adjusted to 6.0, and polyacrylamide is adopted for precipitation.
The technical effect achieved by the technical scheme is as follows: polyacrylamide is used as an anionic flocculant and can precipitate particles with positive charges, the isoelectric point of immunoglobulin is 8.6, and the isoelectric point of BSA protein is 4.7, so that when the pH value of the solution is 6.0, the BSA protein is negatively charged, and the immunoglobulin is positively charged, at the moment, in an acidic solution, the polyacrylamide is negatively charged and can be combined with the immunoglobulin with positive charges to form a precipitate, and the purpose of primarily precipitating the immunoglobulin is achieved;
in addition, polyacrylamide can also precipitate other charged particles in serum, so that the aim of clarifying the serum is fulfilled, and the clarity of the serum is improved.
As a preferable technical scheme of the invention, in the step 3), the centrifugation is performed for 5-10min at 10000rpm by adopting a continuous flow centrifuge.
In a preferred embodiment of the present invention, in step 4), the affinity column is a Sepharose CL4B column.
The technical effect achieved by the technical scheme is as follows: staphylococcal Protein A (SPA) has the ability to bind to the Fc fragment of a variety of mammalian IgG molecules, and therefore, the SPA is wrapped in a column that removes the LgG protein from the immunoglobulin.
As a preferable technical solution of the present invention, in the step 5), the filter membrane is a ceramic composite membrane.
In a preferred embodiment of the present invention, in step 5), the sterilization is performed at 121 ℃ for 21 min.
In conclusion, compared with the bovine serum in the prior art, the preparation method of the bovine serum for the confining liquid removes the immunoglobulin to a greater extent, reduces the content of the immunoglobulin to 0.01mg/mL, effectively prevents the immunoadsorption chromogenic reaction between the immunoglobulin and a solid phase carrier, an antigen or an antibody to be detected, reduces a background value and improves the detection limit;
moreover, after the mature bovine serum is clarified by deep filtration, the clarity is reduced to 5-8NTU from 30-200NTU in the prior art, and the clarity of the confining liquid is obviously improved.
Detailed Description
The technical solutions in the embodiments of the present invention are clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1
A preparation method of bovine serum for sealing liquid comprises the following steps:
1) pretreatment: taking adult bovine serum, centrifuging the adult bovine serum by a continuous flow centrifuge at 8000rpm for 10min, and removing blood cells;
2) clarification: sequentially carrying out deep filtration on the pretreated mature bovine serum, then adjusting the pH value of a filtrate to 6.0, and precipitating by adopting a polyacrylamide flocculant to obtain the clarified mature bovine serum;
3) centrifuging: centrifuging the clarified adult calf serum by a continuous flow centrifuge at 10000rpm for 5min, and taking the supernatant;
4) affinity column chromatography: subjecting the obtained supernatant to Sepharose CL4B column chromatography to obtain chromatography solution;
5) and (3) filtering: filtering the obtained chromatography liquid with 0.05 μm ceramic composite filter membrane, and sterilizing at 121 deg.C for 21min to obtain bovine serum for sealing liquid.
Example 2
A preparation method of bovine serum for sealing liquid comprises the following steps:
1) pretreatment: taking adult bovine serum, centrifuging the adult bovine serum by a continuous flow centrifuge at 8000rpm for 15min, and removing blood cells;
2) clarification: sequentially carrying out deep filtration on the pretreated mature bovine serum, then adjusting the pH value of a filtrate to 6.0, and precipitating by adopting a polyacrylamide flocculant to obtain the clarified mature bovine serum;
3) centrifuging: centrifuging the clarified adult calf serum by a continuous flow centrifuge at 10000rpm for 10min, and taking the supernatant;
4) affinity column chromatography: subjecting the obtained supernatant to Sepharose CL4B column chromatography to obtain chromatography solution;
5) and (3) filtering: filtering the obtained chromatography liquid with 0.01 μm ceramic composite filter membrane, and sterilizing at 121 deg.C for 21min to obtain bovine serum for sealing liquid.
Example 3
A preparation method of bovine serum for sealing liquid comprises the following steps:
1) pretreatment: taking adult bovine serum, centrifuging the adult bovine serum by a continuous flow centrifuge at 8000rpm for 12min, and removing blood cells;
2) clarification: sequentially carrying out deep filtration on the pretreated mature bovine serum, then adjusting the pH value of a filtrate to 6.0, and precipitating by adopting a polyacrylamide flocculant to obtain the clarified mature bovine serum;
3) centrifuging: centrifuging the clarified adult calf serum by a continuous flow centrifuge at 10000rpm for 8min, and taking the supernatant;
4) affinity column chromatography: subjecting the obtained supernatant to Sepharose CL4B column chromatography to obtain chromatography solution;
5) and (3) filtering: filtering the obtained chromatography liquid with 0.03 μm ceramic composite filter membrane, and sterilizing at 121 deg.C for 21min to obtain bovine serum for sealing liquid.
The immunoglobulin content of the fetal bovine serum obtained in examples 1 to 3 was measured by the following method:
in each example, 3 samples of bovine serum were taken, and the total protein content of bovine serum and the albumin content of bovine serum were measured using a total protein content measurement kit (biuret method) and a serum albumin content measurement kit (bromocresol green method) provided by Nanjing institute of bioengineering, respectively, and the results are shown in Table 1.
TABLE 1 results of measurement of total protein content of fetal bovine serum in examples 1-3
| Detecting items | Example 1 | Example 2 | Example 3 |
| Immunoglobulin content (mg/mL) | 0.016 | 0.015 | 0.01 |
Wherein, the serum immunoglobulin content is the total serum protein content-serum albumin content.
As is clear from Table 1, bovine serum for blocking solutions was prepared in the same manner as in examples 1 to 3, and the immunoglobulin content was low.
The results of examining the clarity of the blocking solutions prepared in examples 1 to 3 with bovine serum by the turbidity assay are shown in Table 2.
TABLE 2 results of clarity tests in examples 1-3
| Detecting items | Example 1 | Example 2 | Example 3 |
| Clarity (NTU) | 6 | 8 | 5 |
As is clear from Table 2, the serum for the blocking solution prepared by the method of the present invention was high in clarity.
Example 4ELISA experiment
A blocking solution was further prepared from the bovine serum prepared in example 3, and 12.0g of sucrose, 7.2g of skim milk powder, and Na were weighed out separately2HPO4·12H2O 4.64g、NaH2PO4·2H2Placing 0.48g of O and 2.4g of AEP-HBC in a 1L glass container, adding 500mL of pure water, stirring at room temperature for 30min for dissolution by using a magnetic stirrer, adding 8mL of glycerol and 450mL of pure water, continuing stirring for 12h by using the magnetic stirrer, adding 32mL of bovine serum and 300240 mu L of Proclin, and stirring for 30min by using the magnetic stirrer until the volume is constant to 1L to obtain a sealing solution.
In the ELISA test for detecting HAO (hydroxylamine oxidase) using an antibody against HAO, 150. mu.L/well of a blocking solution was added to the microplate in the blocking step of the procedure, and the microplate was blocked at 37 ℃ for 2 hours and then patted dry for use. Adding 150 mu L/hole phosphate buffer solution into the ELISA plate, sealing for 2h at 37 ℃, and patting dry as a blank control ELISA plate for later use;
the HAO concentrations were set to 1mg/mL and 10-1mg/mL、10-2mg/mL、10-3mg/mL、10-4mg/mL、10- 5mg/mL、10-6mg/mL、10-7mg/mL、10-8mg/mL, after the solid phase carrier is sealed by the sealing liquid, HAO solutions with different concentrations are respectively detected, and the results show that when the concentration of the HAO is 1mg/mL and 10-1mg/mL、10-2mg/mL、10- 3mg/mL、10-4mg/mL、10-5mg/mL、10-6mg/mL、10-7The color reaction can occur at the concentration of 10 mg/mL- 8No color reaction occurs when the concentration is mg/mL, which indicates that the detection limit can reach 0.1ng/mL after the kit is sealed by the sealing liquid.
Comparative example:
comparison 1: removing the step 5) in the example 3), preparing bovine serum according to the method of the steps 1) to 4);
comparison 2: step 4) in example 3 was removed) and bovine serum was prepared according to the methods of steps 1) -3) and 5);
comparison 3: removing the step 4) and the step 5) in the example 3), and preparing bovine serum according to the method of the steps 1) to 3);
HAO was detected with the above-mentioned kit, and the concentration of HAO was set to 1mg/mL and 10 for each comparative example-1mg/mL、10- 2mg/mL、10-3mg/mL、10-4mg/mL、10-5mg/mL、10-6mg/mL、10-7mg/mL、10-8mg/mL, the result shows that the minimum limit of the test of the comparative example 1 is 10-6mg/mL, minimum 10 for comparative example 2-7mg/mL, minimum 10 for comparative example 3-5mg/mL, which indicates that the content of the immunoglobulin in the serum of the adult cattle can be effectively removed by affinity column chromatography and filtration, and the detection limit is improved.
The embodiments in the present description are described in a progressive manner, each embodiment focuses on differences from other embodiments, and the same and similar parts among the embodiments are referred to each other.
The previous description of the disclosed embodiments is provided to enable any person skilled in the art to make or use the present invention. Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention. Thus, the present invention is not intended to be limited to the embodiments shown herein but is to be accorded the widest scope consistent with the principles and novel features disclosed herein.
Claims (7)
1. The preparation method of bovine serum for the sealing liquid is characterized by comprising the following steps of:
1) pretreatment: taking adult bovine serum, and pretreating the adult bovine serum to obtain pretreated adult bovine serum;
2) clarification: sequentially carrying out deep filtration and precipitation on the pretreated mature bovine serum to obtain clarified mature bovine serum;
3) centrifuging: centrifuging the clarified adult calf serum, and taking the supernatant;
4) affinity column chromatography: performing affinity column chromatography on the obtained supernatant to obtain a chromatographic solution;
5) and (3) filtering: filtering the obtained chromatography liquid with 0.01-0.05 μm filter membrane, and sterilizing to obtain bovine serum for blocking liquid.
2. The method for preparing bovine serum for sealing fluid according to claim 1, wherein in the step 1), the pretreatment comprises: the resultant bovine serum was centrifuged at 8000rpm for 10-15min by a continuous flow centrifuge.
3. The method for preparing bovine serum for sealing fluid according to claim 1, wherein in the step 2), the precipitation is performed by adjusting the pH value of the filtrate to 6.0 and then performing precipitation by using a polyacrylamide flocculant.
4. The method for preparing bovine serum for sealing fluid according to claim 1, wherein the centrifugation in step 3) is performed by using a continuous flow centrifuge at 10000rpm for 5-10 min.
5. The method for preparing bovine serum for a sealing solution according to claim 1, wherein in the step 4), the affinity column is Sepharose CL4B column.
6. The method for preparing bovine serum for sealing fluid according to claim 1, wherein in step 5), the filter membrane is a ceramic composite membrane.
7. The method for preparing bovine serum for sealing solution according to claim 1, wherein the sterilization in step 5) is performed at 121 ℃ for 21 min.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN112920995A (en) * | 2021-03-31 | 2021-06-08 | 赵峻岭 | Mesenchymal stem cell culture serum refueling bag and application thereof |
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Cited By (1)
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| CN112920995A (en) * | 2021-03-31 | 2021-06-08 | 赵峻岭 | Mesenchymal stem cell culture serum refueling bag and application thereof |
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