A kind of precise qualitative and quantitative detection method of oil adjuvant vaccine

A quantitative detection method and oil adjuvant technology, which is applied in the direction of material inspection products, test pharmaceutical preparations, test sample preparation, etc., can solve problems such as difficult to handle, unable to detect water phase after demulsification, and achieve improved accuracy, Increase equipment maintenance and use costs, good repeatability effect

Active Publication Date: 2018-09-11
SHANGHAI SHEN LIAN BIOMEDICAL CORP
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Substances such as surfactants in the water phase after demulsification are recognized as the most difficult impurities in the industry, and the existing technology in the industry cannot directly detect the water phase after demulsification

Method used

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  • A kind of precise qualitative and quantitative detection method of oil adjuvant vaccine
  • A kind of precise qualitative and quantitative detection method of oil adjuvant vaccine
  • A kind of precise qualitative and quantitative detection method of oil adjuvant vaccine

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] This embodiment provides an accurate qualitative and quantitative detection method for oil adjuvant vaccines, comprising the following steps:

[0054] 1) Take 10ml of the vaccine to be tested (commercially available foot-and-mouth disease synthetic peptide vaccine, the concentration is 75ug / ml) mixed with n-butanol at a volume ratio of 1:1, add 50mg of histidine, oscillate and mix, at 4°C, with Centrifuge at 3000r / min for 15 minutes, and carefully extract the lower aqueous phase with a 10ml syringe after centrifugation to obtain the aqueous phase antigen sample.

[0055] 2) Take 0.1 ml of the aqueous phase antigen sample prepared in step 1, and use HPLC to detect that the concentration of the sample is 69.3 ug / ml, and the demulsification efficiency is 92.4%.

[0056] 3) Precise and qualitative analysis of the water-phase antigen samples

[0057] 3.1 Sample treatment: put the aqueous phase antigen sample obtained in step 1 through ultrafiltration, diafiltration, and the...

Embodiment 2

[0069] This embodiment provides an accurate qualitative and quantitative detection method for oil adjuvant vaccines, comprising the following steps:

[0070] 1) Take 10ml of the vaccine to be tested (commercially available foot-and-mouth disease synthetic peptide vaccine, the concentration is 75ug / ml) mixed with n-butanol at a volume ratio of 1:1, add 10mg of phenylalanine, shake and mix well, at 4°C, Centrifuge at 3000r / min for 15 minutes, and carefully extract the lower aqueous phase with a 10ml syringe after centrifugation to obtain the aqueous phase antigen sample.

[0071] 2) Take 0.1 ml of the aqueous phase antigen sample prepared in step 1, and use HPLC to detect that the concentration of the sample is 65.7 ug / ml, and the demulsification efficiency is 87.6%.

[0072] 3) Precise and qualitative analysis of the water-phase antigen samples

[0073] 3.1 Sample processing: After the foot-and-mouth disease vaccine is demulsified, it is subjected to ultrafiltration, diafiltra...

Embodiment 3

[0085] This embodiment provides an accurate qualitative and quantitative detection method for oil adjuvant vaccines, comprising the following steps:

[0086] 1) Take 10ml of the vaccine to be tested (commercially available foot-and-mouth disease synthetic peptide vaccine, the concentration is 75ug / ml) and n-butanol at a volume ratio of 1:1, add 200mg of proline to each tube, oscillate and mix, and store at 4°C , centrifuge at 3000r / min for 15 minutes, and carefully extract the lower aqueous phase with a 10ml syringe after centrifugation to obtain the aqueous phase antigen sample.

[0087]2) Take 0.1 ml of the aqueous phase antigen sample prepared in step 1, and use HPLC to detect that the concentration of the sample is 71.1 ug / ml, and the demulsification efficiency is 94.8%.

[0088] 3) The water-phase antigen sample is subjected to precise and qualitative analysis, and the method is exactly the same as that in Example 1. The analysis result by LC / MS detection method shows th...

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Abstract

The present invention provides a method for accurate qualitative and quantitative detection of an oil adjuvant vaccine, the method comprises the following steps: demulsification of the oil adjuvant vaccine, quantitative analysis of a demulsified antigen sample, purification by a gel electrophoresis method, and qualitative detection of the purified antigen sample; the demulsification method is as follows: mixing the oil adjuvant vaccine with n-butanol, adding a competition agent, evenly mixing, centrifuging, taking a water phase solution for ultra-filtration and washing filtration, and performing vacuum freeze drying or concentrating to obtain an antigen sample. By adding of the competition agent for competition with a surface active agent for antigen binding sites, antigens in the oil adjuvant vaccine can be released to a water phase, the recovery rate of the antigens in the water phase is greatly improved; the demulsified antigen is separated and purified by electrophoresis, the purity of the antigen is improved, the impurity content of the surface active agent in the antigen is basically 0, so that the purified sample has good reproducibility during the qualitative detection, and the accuracy of test results is improved.

Description

technical field [0001] The invention relates to the technical field of foot-and-mouth disease vaccine detection, in particular to an accurate qualitative and quantitative detection method for oil adjuvant vaccines. Background technique [0002] The existing vaccine quality standards stipulate that the efficacy test must use this animal for testing. Since the country implements a 100% enhanced immunization policy, it is difficult to select susceptible animals for testing, and animal challenge has high requirements for experimental facilities (BSL3 level laboratory), time-consuming (more than one month), and large capital expenditure. If the serum neutralization test is used to select susceptible animals, it is technically difficult to exclude non-susceptible animals with cellular immunity, and the problem of irregular test data often occurs in practice, which affects the accuracy of the test. Therefore, the quality inspection of foot-and-mouth disease vaccines, especially va...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N33/15G01N1/34
CPCG01N1/34G01N33/15
Inventor 俞爱敏马贵军石海芳
Owner SHANGHAI SHEN LIAN BIOMEDICAL CORP
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