CN106771016A - A kind of fast qualitative quantitative detecting method of oil-adjuvant vaccine - Google Patents
A kind of fast qualitative quantitative detecting method of oil-adjuvant vaccine Download PDFInfo
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- CN106771016A CN106771016A CN201611122911.7A CN201611122911A CN106771016A CN 106771016 A CN106771016 A CN 106771016A CN 201611122911 A CN201611122911 A CN 201611122911A CN 106771016 A CN106771016 A CN 106771016A
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- 229960005486 vaccine Drugs 0.000 title claims abstract description 53
- 238000000034 method Methods 0.000 title claims abstract description 39
- 239000002671 adjuvant Substances 0.000 title claims abstract description 33
- 239000000427 antigen Substances 0.000 claims abstract description 48
- 102000036639 antigens Human genes 0.000 claims abstract description 48
- 108091007433 antigens Proteins 0.000 claims abstract description 48
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 32
- 238000001514 detection method Methods 0.000 claims abstract description 25
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 claims abstract description 24
- 238000000746 purification Methods 0.000 claims abstract description 11
- 238000005119 centrifugation Methods 0.000 claims abstract description 6
- 239000012467 final product Substances 0.000 claims abstract description 6
- XXMFJKNOJSDQBM-UHFFFAOYSA-N 2,2,2-trifluoroacetic acid;hydrate Chemical compound [OH3+].[O-]C(=O)C(F)(F)F XXMFJKNOJSDQBM-UHFFFAOYSA-N 0.000 claims description 5
- 239000003795 chemical substances by application Substances 0.000 claims description 5
- 238000001962 electrophoresis Methods 0.000 claims description 5
- 238000010828 elution Methods 0.000 claims description 5
- ONIBWKKTOPOVIA-UHFFFAOYSA-N Proline Natural products OC(=O)C1CCCN1 ONIBWKKTOPOVIA-UHFFFAOYSA-N 0.000 claims description 3
- 239000004475 Arginine Substances 0.000 claims description 2
- ONIBWKKTOPOVIA-BYPYZUCNSA-N L-Proline Chemical compound OC(=O)[C@@H]1CCCN1 ONIBWKKTOPOVIA-BYPYZUCNSA-N 0.000 claims description 2
- ODKSFYDXXFIFQN-BYPYZUCNSA-P L-argininium(2+) Chemical compound NC(=[NH2+])NCCC[C@H]([NH3+])C(O)=O ODKSFYDXXFIFQN-BYPYZUCNSA-P 0.000 claims description 2
- HNDVDQJCIGZPNO-YFKPBYRVSA-N L-histidine Chemical compound OC(=O)[C@@H](N)CC1=CN=CN1 HNDVDQJCIGZPNO-YFKPBYRVSA-N 0.000 claims description 2
- KDXKERNSBIXSRK-YFKPBYRVSA-N L-lysine Chemical compound NCCCC[C@H](N)C(O)=O KDXKERNSBIXSRK-YFKPBYRVSA-N 0.000 claims description 2
- COLNVLDHVKWLRT-QMMMGPOBSA-N L-phenylalanine Chemical compound OC(=O)[C@@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-QMMMGPOBSA-N 0.000 claims description 2
- KDXKERNSBIXSRK-UHFFFAOYSA-N Lysine Natural products NCCCCC(N)C(O)=O KDXKERNSBIXSRK-UHFFFAOYSA-N 0.000 claims description 2
- 239000004472 Lysine Substances 0.000 claims description 2
- 150000001413 amino acids Chemical class 0.000 claims description 2
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 claims description 2
- HNDVDQJCIGZPNO-UHFFFAOYSA-N histidine Natural products OC(=O)C(N)CC1=CN=CN1 HNDVDQJCIGZPNO-UHFFFAOYSA-N 0.000 claims description 2
- COLNVLDHVKWLRT-UHFFFAOYSA-N phenylalanine Natural products OC(=O)C(N)CC1=CC=CC=C1 COLNVLDHVKWLRT-UHFFFAOYSA-N 0.000 claims description 2
- 239000012535 impurity Substances 0.000 abstract description 7
- 238000011084 recovery Methods 0.000 abstract description 5
- 239000004094 surface-active agent Substances 0.000 abstract description 5
- 238000012360 testing method Methods 0.000 abstract description 4
- 238000004451 qualitative analysis Methods 0.000 abstract 1
- 238000004445 quantitative analysis Methods 0.000 abstract 1
- 230000000052 comparative effect Effects 0.000 description 8
- 238000007689 inspection Methods 0.000 description 5
- 229940023041 peptide vaccine Drugs 0.000 description 5
- 238000004128 high performance liquid chromatography Methods 0.000 description 4
- 238000012512 characterization method Methods 0.000 description 3
- 239000012141 concentrate Substances 0.000 description 3
- 230000009514 concussion Effects 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 2
- 230000000890 antigenic effect Effects 0.000 description 2
- 235000014304 histidine Nutrition 0.000 description 2
- 230000000091 immunopotentiator Effects 0.000 description 2
- 230000010354 integration Effects 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 235000008729 phenylalanine Nutrition 0.000 description 2
- 101100272788 Arabidopsis thaliana BSL3 gene Proteins 0.000 description 1
- 235000001014 amino acid Nutrition 0.000 description 1
- 238000013459 approach Methods 0.000 description 1
- 235000009697 arginine Nutrition 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 230000033228 biological regulation Effects 0.000 description 1
- 230000007969 cellular immunity Effects 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000010612 desalination reaction Methods 0.000 description 1
- 238000004945 emulsification Methods 0.000 description 1
- 150000002411 histidines Chemical class 0.000 description 1
- 230000001900 immune effect Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 235000018977 lysine Nutrition 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 230000014759 maintenance of location Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 238000001819 mass spectrum Methods 0.000 description 1
- 239000000203 mixture Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 150000002994 phenylalanines Chemical class 0.000 description 1
- 235000013930 proline Nutrition 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000012372 quality testing Methods 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 238000001179 sorption measurement Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000000108 ultra-filtration Methods 0.000 description 1
- 239000000273 veterinary drug Substances 0.000 description 1
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
- G01N33/15—Medicinal preparations ; Physical properties thereof, e.g. dissolubility
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/30—Staining; Impregnating ; Fixation; Dehydration; Multistep processes for preparing samples of tissue, cell or nucleic acid material and the like for analysis
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N1/00—Sampling; Preparing specimens for investigation
- G01N1/28—Preparing specimens for investigation including physical details of (bio-)chemical methods covered elsewhere, e.g. G01N33/50, C12Q
- G01N1/34—Purifying; Cleaning
Abstract
The invention provides a kind of fast qualitative quantitative detecting method of oil-adjuvant vaccine, it is characterised in that comprise the following steps：After oil-adjuvant vaccine is demulsified, gained water phase antigen samples carry out ZIPTIP purifying, and sample after purification carries out qualitative and quantitative detection；The method of the demulsification is：Oil-adjuvant vaccine is mixed with n-butanol, competitor is subsequently adding, after vibration is mixed, centrifugation obtains final product water phase antigen samples.The method competes antigen binding site by adding competitor with surfactant, so as to the antigen in oil-adjuvant vaccine to be discharged into water phase the rate of recovery of antigen substantially increased in water phase；The sample after demulsification is purified using ZIPTIP again, the impurity in sample is effectively removed, substantially increased the purity of sample, sample is thus carried out into qualitative and quantitative analysis again, improve the accuracy and reliability of testing result.
Description
Technical field
The present invention relates to aftosa vaccine detection technique field, and in particular to a kind of fast qualitative of oil-adjuvant vaccine is quantified
Detection method.
Background technology
Regulation efficacy test must be tested using this animal in currently available vaccines, existing vaccines quality standard, because country carries out
100% reinforced immunological policy, it is difficult to selecting susceptible inspection uses animal, and animal attacks malicious high to Experimental Establishment requirement
(BSL3 grades of laboratory), time-consuming (more than one month), capital cost is big.If selecting impressibility using serum neutralization test
Thing, is technically difficult to exclude the non-susceptible animal with cellular immunity, and inspection data often occurs in practice in inspection does not advise
The problem of rule, influences the accuracy of inspection.Therefore aftosa vaccine perplexs always in particular for the quality testing of the vaccine of ox
Veterinary drug monitoring department and associated production enterprise.Therefore need to develop tested in vitro technology as early as possible, tried instead of existing animal
Test.
Inspection of the current country to vaccine is just progressively being transitioned into the detection to vaccine endoantigen, and currently more universal side
Method is that antigen is detected after vaccine is demulsified, and demulsification treatment is carried out by by vaccine, antigen is transferred in water phase, then right
It carries out follow-up detection and analysis.It is well known that vaccine is formed with certain proportion, by specific program emulsification with adjuvant by antigen
, but adjuvant is but for vaccine demulsification detection brings huge obstacle.
Due to complicated component in the oily adjuvant used in vaccine emulsion process, contain surfactant, immunopotentiator
Exist Deng material, can not removed in industry above-mentioned containing above-mentioned impurity and demulsifier often in the water phase after demulsification
The method of impurity and demulsifier, and impurity and demulsifier etc. can also largely effect on detection process thereafter, cause signal cover or
Interference, has forced down the intensity of antigen signals, in addition cannot effective detection to antigen therein, due to being randomly assigned for wherein impurity
Property, cause the repeatability of its detection method not good, while increased instrument maintenance cost.In vaccine industry, how oil is helped
It is technological difficulties generally acknowledged in the industry that vaccinating agent is demulsified and composition in vaccine is detected and is identified, due to oil-adjuvant vaccine
Complicated component, wherein can cause to do to all detection methods containing materials such as substantial amounts of surfactant and immunopotentiators again
Disturb, or even cannot detect, it is impossible to reflect the time of day of vaccine endoantigen.
Although having breaking method in the industry, its efficiency and effect be not good, and kind is had after traditional breaking method demulsification
The problems such as can't detect antigen in the problem of kind, such as incomplete, the water phase that is demulsified and oil phase unintelligible, the water phase of boundary annoyings always
Industry.How a kind of efficient breaking method into the industry problem demanding prompt solution is looked for.
ZIPTIP methods are a kind of method of desalination before easy mass spectrum, the going for salts substances in detection is analyzed
Remove.Not yet having in the prior art, the method is used for the report of antigen purification.
The content of the invention
For defect of the prior art, the invention provides a kind of fast qualitative quantitative determination side of oil-adjuvant vaccine
Method.
The purpose of the present invention is achieved through the following technical solutions：
The invention provides a kind of fast qualitative quantitative detecting method of oil-adjuvant vaccine, comprise the following steps：
After oil-adjuvant vaccine is demulsified, gained water phase antigen samples carry out ZIPTIP purifying, and sample after purification is determined
Property quantitative determination；
The method of the demulsification is：Oil-adjuvant vaccine is mixed with n-butanol, competitor is subsequently adding, after vibration is mixed,
Centrifugation, obtains final product water phase antigen samples.
Preferably, the competitor includes at least one in amino acid and its derivative.
Preferably, the competitor is the one kind in lysine, arginine, phenylalanine, histidine and proline.
Preferably, the addition of the competitor is：1-40mg competitors are added in per 1ml oil-adjuvant vaccines, more preferably
1-20mg competitors.The excessive concentration of the competitor, the agent saturation that can constitute competition is separated out, and influences ultra-filtration process；Concentration is too low,
Can be constituted competition effect on driving birds is not good, it is impossible to the antigen of release detection enough.
Preferably, the oil-adjuvant vaccine and the volume ratio of n-butanol are 9：1~5：5.
It is highly preferred that the oil-adjuvant vaccine is 1 with the volume ratio of n-butanol:1.The volume of oil-adjuvant vaccine and n-butanol
It is identical, can preferably ensure that vaccine is demulsified completely.
Preferably, what the ZIPTIP was purified comprises the following steps that：
A1. ZIPTIP is activated using 50%ACN suctions；
A2. pipette samples suction is for several times；
A3. ZIPTIP is cleaned using the suction of 0.05%TFA water；
A4. the ACN elution samples containing 0.05%TFA are used, you can.
Preferably, the sample after purification is freezed or concentrated.
Preferably, the quantitative and qualitative for carrying out antigen samples using the method for electrophoresis poststaining is detected.
Using breaking method of the invention, because rate of recovery of antigen is high, the antigen samples for obtaining need not be further purified, i.e.,
Can be directly used for the quantitative and qualitative detection of antigen.
Prior art is compared, and the present invention has following beneficial effect：
1) present invention competes antigen binding site by adding competitor with surfactant, so as to by oil-adjuvant vaccine
Antigen discharge into water phase, compared with competitor is not added with, substantially increase the rate of recovery of antigen in water phase.
2) this method uses ZIPTIP purification process, the impurity in antigen samples can be effectively removed, by specific adsorption
Antigen, removes the impurity such as surfactant in complex sample, at the same the sample obtained after its wash-out be used directly for it is high-precision
Spend the analysis detection of instrument.
Brief description of the drawings
The detailed description made to non-limiting example with reference to the following drawings by reading, further feature of the invention,
Objects and advantages will become more apparent upon：
Fig. 1 is antigen concentration HPLC detection collection of illustrative plates in water phase after the method demulsification of comparative example 1.
Specific embodiment
With reference to specific embodiment, the present invention is described in detail.Following examples will be helpful to the technology of this area
Personnel further understand the present invention, but the invention is not limited in any way.It should be pointed out that to the ordinary skill of this area
For personnel, without departing from the inventive concept of the premise, various modifications and improvements can be made.These belong to the present invention
Protection domain.
Embodiment 1
A kind of fast qualitative quantitative detecting method of oil-adjuvant vaccine is present embodiments provided, using following steps：
1) sample demulsification will demulsification treatment as follows：
10ml vaccines to be checked (commercially available Schweineseuche synthetic peptide vaccine, concentration is 75ug/ml) are taken with n-butanol with volume
Than 1:1 mixing, adds 50mg histidines, and concussion is mixed, and under the conditions of 4 DEG C, is centrifuged 15 minutes with 3000r/min, is used after centrifugation
10ml syringes carefully extract lower floor's water phase, obtain final product water phase antigen samples.
2) the ZIPTIP purifying of sample
2.1 by step 1) obtained in water phase antigen samples it is lyophilized or concentrate, purified using ZIPTIP, its step is such as
Under：
1. ZIPTIP is activated using 50%ACN suctions；
2. pipette samples suction is for several times；
3. ZIPTIP is cleaned using the suction of 0.05%TFA water；
4. the ACN elution samples containing 0.05%TFA are used.
3) qualitative and quantitative detection
By by step 2) sample volume after purification be water phase after demulsification 1/10 (i.e. 0.5ml), i.e. concentration improves 10
Times, electrophoresis poststaining is carried out, its band is analyzed, contrasted with its theoretical molecular, antigen bands appear in theoretical molecular
Size area, can understand that its protein content is about 10-15ug to its initial characterization by band gray value, and applied sample amount is 10ul samples,
Sample sample concentration is 1-1.5mg/ml thereon, and Volume Changes (divided by 2) are original after converting its cycles of concentration (divided by 10) and demulsification
Concentration is 50-75ug/ml, and measuring concentration 69.3ug/ml results with liquid chromatogram is consistent.
The commercially available Schweineseuche synthetic peptide vaccine that the present embodiment is used is compareed with theoretical antigen concentration standard, to it
HPLC detection collection of illustrative plates samples go out peak position and are integrated, and as shown in table 1, antigenic content is integrating peak face in sample for its integration information
Long-pending form embodies, and its integrated peak areas is 2738690.To the present embodiment using antigen in water phase after Butanol+His demulsifications
The HPLC detection collection of illustrative plates samples of sample concentration go out peak position and are integrated, and its integration information is as shown in table 2, antigenic content in sample
Embodied in integrated peak areas form, its integrated peak areas is 2530549.The result for contrasting Tables 1 and 2 understands, both is integrated
After information contrast, its rate of recovery of antigen is 92.4%, that is, the efficiency that is demulsified is 92.4%.
Table 1
Table 2
Embodiment 2
A kind of fast qualitative quantitative detecting method of oil-adjuvant vaccine is present embodiments provided, using following steps：
1) sample demulsification will demulsification treatment as follows：
10ml vaccines to be checked (commercially available Schweineseuche synthetic peptide vaccine, concentration is 75ug/ml) are taken with n-butanol with volume
Than 1:1 mixing, adds 10mg phenylalanines, and concussion is mixed, and under the conditions of 4 DEG C, is centrifuged 15 minutes with 3000r/min, after centrifugation
Lower floor's water phase is carefully extracted with 10ml syringes, water phase antigen samples are obtained final product.
Aftosa vaccine is demulsified using the method for the present embodiment, demulsification efficiency is 87.6%.
2) the ZIPTIP purifying of sample
2.1 by step 1) obtained in water phase antigen samples it is lyophilized or concentrate, purified using ZIPTIP, its step is such as
Under：
1. ZIPTIP is activated using 50%ACN suctions；
2. pipette samples suction is for several times；
3. ZIPTIP is cleaned using the suction of 0.05%TFA water；
4. the ACN elution samples containing 0.05%TFA are used.
3) qualitative and quantitative detection
By by step 2) sample volume after purification be water phase after demulsification 1/10 (i.e. 0.5ml), i.e. concentration improves 10
Times, electrophoresis poststaining is carried out, its band is analyzed, contrasted with its theoretical molecular, antigen bands appear in theoretical molecular
Size area, can understand that its protein content is about 10-15ug to its initial characterization by band gray value, and applied sample amount is 10ul samples,
Sample sample concentration is 1-1.5mg/ml thereon, and Volume Changes (divided by 2) are original after converting its cycles of concentration (divided by 10) and demulsification
Concentration is 50-75ug/ml, and measuring concentration 65.7ug/ml results with liquid chromatogram is consistent.
Embodiment 3
A kind of fast qualitative quantitative detecting method of oil-adjuvant vaccine is present embodiments provided, using following steps：
1) sample demulsification will demulsification treatment as follows：
10ml vaccines to be checked (commercially available Schweineseuche synthetic peptide vaccine, concentration is 75ug/ml) are taken with n-butanol with volume
Than 1:1 mixing, often pipe addition 200mg proline, concussion is mixed, and under the conditions of 4 DEG C, is centrifuged 15 minutes with 3000r/min, centrifugation
Lower floor's water phase is carefully extracted with 10ml syringes afterwards, water phase antigen samples are obtained final product.
Aftosa vaccine is demulsified using the method for the present embodiment, demulsification efficiency is 94.8%.
2) the ZIPTIP purifying of sample
2.1 by step 1) obtained in water phase antigen samples it is lyophilized or concentrate, purified using ZIPTIP, its step is such as
Under：
1. ZIPTIP is activated using 50%ACN suctions；
2. pipette samples suction is for several times；
3. ZIPTIP is cleaned using the suction of 0.05%TFA water；
4. the ACN elution samples containing 0.05%TFA are used.
3) qualitative and quantitative detection
By by step 2) sample volume after purification be water phase after demulsification 1/10 (i.e. 0.5ml), i.e. concentration improves 10
Times, electrophoresis poststaining is carried out, its band is analyzed, contrasted with its theoretical molecular, antigen bands appear in theoretical molecular
Size area, can understand that its protein content is about 10-15ug to its initial characterization by band gray value, and applied sample amount is 10ul samples,
Sample sample concentration is 1-1.5mg/ml thereon, and Volume Changes (divided by 2) are original after converting its cycles of concentration (divided by 10) and demulsification
Concentration is 50-75ug/ml, and measuring concentration 71.7ug/ml results with liquid chromatogram is consistent.
Comparative example 1
This comparative example provides a kind of breaking method of aftosa vaccine, and the method with embodiment 1 is essentially identical, difference
Place is only that：Competitor is added without in this comparative example.
Aftosa vaccine is demulsified using the method for this comparative example, the comparative example is using in water phase after n-butanol demulsification
Antigen concentration HPLC detects collection of illustrative plates as shown in figure 1, from this figure it can be seen that in antigen theory retention time (23-28min) simultaneously
Antigen is not detected, illustrates that the amount of antigen contained in sample is extremely low, i.e., the rate of recovery of antigen in this comparative example is substantially zeroed, its demulsification
Efficiency is substantially zeroed, and demulsification efficiency is 0%.Carry out ZIPTIP after purification, carry out qualitative and quantitative detection, it is impossible to obtain target protein
Band.
The result of comparative example and embodiment illustrates, synthetic peptide vaccine uses traditional n-butanol breaking method not cause
Antigen distribution is in water phase, and the amount of antigen that can be detected in water phase is extremely low, can not meet detection and require, and after adding competitor
Amount of antigen reaches more than 85% in water phase after demulsification, subsequent purification detection is met enough and is required.
Concrete application approach of the present invention is a lot, and the above is only the preferred embodiment of the present invention.It should be pointed out that more than
Embodiment is merely to illustrate the present invention, and the protection domain being not intended to limit the invention.For the common skill of the art
For art personnel, under the premise without departing from the principles of the invention, some improvement can also be made, these improvement also should be regarded as this hair
Bright protection domain.
Claims (7)
1. the fast qualitative quantitative detecting method of a kind of oil-adjuvant vaccine, it is characterised in that comprise the following steps：
After oil-adjuvant vaccine is demulsified, gained water phase antigen samples carry out ZIPTIP purifying, and it is qualitative fixed that sample after purification is carried out
Amount detection；
The method of the demulsification is：Oil-adjuvant vaccine is mixed with n-butanol, competitor is subsequently adding, after vibration is mixed, centrifugation,
Obtain final product water phase antigen samples.
2. the fast qualitative quantitative detecting method of oil-adjuvant vaccine according to claim 1, it is characterised in that the competition
Agent includes at least one in amino acid and its derivative.
3. the fast qualitative quantitative detecting method of oil-adjuvant vaccine according to claim 2, it is characterised in that the competition
Agent is the one kind in lysine, arginine, phenylalanine, histidine and proline.
4. the fast qualitative quantitative detecting method of oil-adjuvant vaccine according to claim 1, it is characterised in that the competition
The addition of agent is：1-40mg competitors are added in per 1ml oil-adjuvant vaccines.
5. the fast qualitative quantitative detecting method of oil-adjuvant vaccine according to claim 1, it is characterised in that described
What ZIPTIP was purified comprises the following steps that：
A1. ZIPTIP is activated using 50%ACN suctions；
A2. pipette samples suction is for several times；
A3. ZIPTIP is cleaned using the suction of 0.05%TFA water；
A4. the ACN elution samples containing 0.05%TFA are used, you can.
6. the fast qualitative quantitative detecting method of oil-adjuvant vaccine according to claim 1, it is characterised in that the purifying
Sample afterwards is freezed or concentrated.
7. the fast qualitative quantitative detecting method of oil-adjuvant vaccine according to claim 1, it is characterised in that use electrophoresis
The method of poststaining carries out the quantitative and qualitative detection of antigen samples.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
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| CN113311052A (en) * | 2021-06-19 | 2021-08-27 | 河南省农业科学院动物免疫学重点实验室 | Method for evaluating content and purity of effective antigen in foot-and-mouth disease vaccine based on gel electrophoresis method |
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| 于生兰 等: "黄芪甲苷单克隆抗体的制备及鉴定", 《天然产物研究与开发》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN113311052A (en) * | 2021-06-19 | 2021-08-27 | 河南省农业科学院动物免疫学重点实验室 | Method for evaluating content and purity of effective antigen in foot-and-mouth disease vaccine based on gel electrophoresis method |
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| CN106771016B (en) | 2020-07-10 |
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