Method for detecting foot-and-mouth disease antigen 146S content with sucrose density gradient ultraviolet light quantitative method

A technology of sucrose density gradient and sucrose gradient, which is applied in the field of quantitative detection of 146S content of foot-and-mouth disease antigen, can solve the problems of detection accuracy and sensitivity constraints, inability to distinguish well, and inability to distinguish 12S and 146S well

Active Publication Date: 2010-02-24
JINYUBAOLING BIO PHARM CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Method 1 and method 2 are for the detection of viral nucleic acid, which can not only detect the nucleic acid of intact virions, but also the nucleic acid of 146S cleavage products, which cannot reflect the immunogen of the virus; method 3 is interfered by pre-complement and anti-complement factors, The accuracy and sensitivity of the detection are restric

Method used

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  • Method for detecting foot-and-mouth disease antigen 146S content with sucrose density gradient ultraviolet light quantitative method
  • Method for detecting foot-and-mouth disease antigen 146S content with sucrose density gradient ultraviolet light quantitative method
  • Method for detecting foot-and-mouth disease antigen 146S content with sucrose density gradient ultraviolet light quantitative method

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0021] The detection process is as follows:

[0022] 1. Take the adherent or suspended live virus and inactivated virus antigen in large-scale production (for example, sample a is the live virus of pig O-type OZK, sample b is the live virus of bovine O-type JMS, and sample c is the inactivated virus of bovine O-type JMS, d sample is bovine Asia-1 type JSL live virus, e sample is bovine Asia-1 type JSL inactivated virus), the samples are respectively treated with organic solvents (a, b, c use chloroform, d, e use trichloroethylene) For pretreatment, the volume of the organic solvent added is 5% of the volume of the sample, the virus liquid and the organic solvent are fully emulsified and mixed for 8 minutes, centrifuged at 3500 rpm for 10 minutes, and the supernatant virus liquid is taken;

[0023] 2. Concentration of the virus solution: Ultracentrifuge the virus solution, resuspend it in PBS to 1 / 10 of the original volume to obtain a virus concentrate, and store it in a refrig...

Embodiment 2

[0033] PD 50 The test method is described in , which is a recognized method for testing vaccine efficacy. With the method of the present invention (sample is pretreated with chloroform, other steps are identical with embodiment one) and PD 5o The same inactivated antigen solution in the following table is detected respectively by the method, and the results are as follows:

[0034] sample

The 146S measured by the method of the present invention

Content (μg / ml)

Half Protected (PD 5o ) value

Bovine Type O JMS Inactivated Antigen Solution 1

0.55

9.1

Bovine Type O JMS Inactivated Antigen Solution 2

0.87

13.2

Bovine Type O JMS Inactivated Antigen Solution 3

0.34

5.8

Bovine Asia-1 type JSL inactivated antigen solution 1

0.98

12.4

Bovine Asia-1 type JSL inactivated antigen solution 2

0.56

7.3

Bovine Asia-1 type JSL inactivated antigen solution 3

0...

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Abstract

The invention provides a method for detecting foot-and-mouth disease antigen 146S content with a sucrose density gradient ultraviolet light quantitative method; the method comprises the following steps: sucrose density gradient ultra centrifugation is carried out to virus concentration liquid to be detected, and then OD259nm value of all fractions is continuously detected by an ultraviolet light spectrophotometer; next, peak area of OD value of a sample to be detected is calculated, and the 146S content in the virus liquid is calculated out according to the formula (FR*PA*FSD*1000)/(S*PL*E*W),wherein FR is flow rate of sucrose which passes through a sample cell; PA is peak area; S is speed, FSD is an absorbance unit, PL is optical path length of the flowing sample cell, W is original weight or volume of a sample which is added on the gradient, E is correction coefficient which is optional positive number. The method has better repetitiveness and applicability, and can be used for accurate and quantitative detection of 146S relative content of various foot-and-mouth viruses.

Description

technical field [0001] The invention relates to a method for quantitatively detecting the content of the foot-and-mouth disease antigen 146S, in particular to a method for detecting the content of the foot-and-mouth disease antigen 146S by a sucrose density gradient ultraviolet light quantitative method. Background technique [0002] The FMD antigen quantitative detection techniques that have been applied or reported at present mainly include: 1. The half-lethal dose (LD50) of suckling mice is used to measure the FMD virus antigen content. Dilution, each titer subcutaneously injects 4 suckling mice by the amount neck back of every 0.2ml, finally calculates virus antigen content according to suckling mouse morbidity and death situation; For example, make a 10-fold serial dilution of the virus on a 96-well flat-bottom culture plate, the amount of virus solution in each well is 50ml, then add 100ml of cell suspension, and culture in a 37°C, 5% carbon dioxide incubator for 72 ho...

Claims

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Application Information

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IPC IPC(8): G01N21/33G01N1/34
Inventor 卢永干王伟峰刘玉梅
Owner JINYUBAOLING BIO PHARM CO LTD
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