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A kind of detection method of swine fever live vaccine potency

A swine fever live vaccine and detection method technology, applied in the field of vaccine efficacy testing, can solve the problem that the surviving swine fever virus particles and inactivated swine fever virus particles cannot be effectively distinguished, the virus is difficult to be detected by immunofluorescence technology, and it has not been widely received. Recognition and other issues, to achieve the effect of intuitive testing results, shortening the testing cycle, and avoiding long testing cycles

Active Publication Date: 2015-11-04
RINGPU (BAODING) BIOLOGICAL PHARMACEUTICAL CO LTD
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] The attenuated strain of classical swine fever virus (strain C) has no pathogenic effect on commonly used and known sensitive primary cells and passaged cell lines, so the immunofluorescence method (IFA) is more suitable for detecting the titer of classical swine fever virus (half Tissue Culture Infectious Dose, TCID 50 ) method, but due to the low rate of infection of cells by the attenuated rabbit strain of CSFV (C strain), low virus titer, monopoly of sensitive cell lines, and poor antibody specificity, it is also difficult to detect the virus by immunofluorescence technology The existence of this method is not widely used in China
[0007] The primers designed in the existing PCR detection method and the real-time fluorescent quantitative PCR detection method cannot effectively distinguish the surviving swine fever virus particles and the inactivated swine fever virions. It degrades RNA fragments), and there are many false positive detection results. These two methods only have certain reference significance when detecting fresh venom, so they have not been widely recognized

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  • A kind of detection method of swine fever live vaccine potency
  • A kind of detection method of swine fever live vaccine potency
  • A kind of detection method of swine fever live vaccine potency

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Comparison test of RT-PCR detection effect between primers of the present invention and certain reporter primers

[0052] The swine fever live vaccine used in this embodiment and the primer information are as follows:

[0053] (1) Live swine fever vaccine: Live swine fever vaccine (cell source) produced by Ringpu (Baoding) Bio-Pharmaceutical Co., Ltd., 20 heads / bottle, batch number 20120811, and the infection rate of rabbits is 7500 RID / head. The virus seed is a attenuated strain of classical swine fever (strain C), which was purchased from the China Veterinary Drug Control Institute.

[0054] (2) Control primers (a reporter primer): This pair of primers was designed in the 5'-UTR region of CSFV attenuated rabbit strain (C strain) (accession number: AF091507), and the length of the amplified product is about 239bp. The downstream primer sequences are:

[0055] F: 5'-CCTGGGTGGTCTAGTCCTG-3'

[0056] R: 5'-AGTGATCAACTCCATGTGCC-3'

[0057] The pair of primers were synth...

Embodiment 2

[0092] The detection method of the present invention detects the virus PCR titer (PD) of the semi-finished product of swine fever live vaccine

[0093] The semi-finished product information of the swine fever live vaccine used in this embodiment is as follows:

[0094] Sample A: Semi-finished virus solution of swine fever live vaccine (cell source) produced by Ringpu (Baoding) Biopharmaceutical Co., Ltd., batch number 20120710.

[0095] Sample B: Semi-finished virus solution of swine fever live vaccine (cell source) produced by Ringpu (Baoding) Biopharmaceutical Co., Ltd., batch number 20100510.

[0096] Positive control: semi-finished virus solution of swine fever live vaccine (cell source) produced by Ringpu (Baoding) Biopharmaceutical Co., Ltd., batch number 20120815, and the infection rate of rabbits was 2×10 4 RID / ml.

[0097] Negative control: DMEM medium containing 3% calf serum (cell culture medium for production).

[0098] This embodiment includes the following ste...

Embodiment 3

[0130] The detection method of the present invention is compared with the stability of repeated detection of the efficacy of the semi-finished product of swine fever live vaccine by the rabbit body stereotyped thermal reaction method

[0131] The detection method of the present invention and the rabbit body stereotyped heat reaction method were respectively used to detect the efficacy of 10 batches of semi-finished products of swine fever live vaccine, which were repeated twice, and the detection results were counted (see Table 4).

[0132] Table 4 Detection method of the present invention and rabbit body reaction heat method detection result

[0133]

[0134] The PCR method of the present invention repeats the detection result and coincidence rate is higher; The repeated detection result coincidence rate of rabbit body stereotyping heat reaction method is relatively low, because the big-eared white rabbit used for detection has individual difference to the sensitivity of ...

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Abstract

The invention relates to a method for detecting the efficacy of a swine fever live-vaccine. The method comprises the following steps: A, design of a primer; B, extraction of total RNA (Ribose Nucleic Acid); C, reverse transcription (RT); D, PCR (Polymerase Chain Reaction) reaction; E, electrophoretic analysis; and F, calculation of virus PCR titer. The method for detecting the efficacy of the swine fever live-vaccine is simple, fast and accurate, overcomes the defects of the current detection method, and meets the need of quickly detecting the efficacy in the mass production process of the swine fever live-vaccine.

Description

technical field [0001] The invention relates to a vaccine efficacy detection method, in particular to a swine fever live vaccine efficacy detection method, which belongs to the field of biotechnology. Background technique [0002] Classical Swine Fever (CSF), commonly known as "rotten intestinal fever", is called hog cholera in the United States, and classical swine fever in some parts of Europe. The disease is an acute febrile systemic septic infectious disease caused by swine fever virus. It can occur in pigs of all ages. The course of the disease varies from acute to chronic. It is popular all year round and highly contagious. It has a high morbidity and mortality rate, and once it occurs, it will be devastating, seriously threatening the development of the pig industry and affecting people's lives. Swine fever is one of the 16 infectious diseases of Class A that must be inspected by the International Veterinary Health Organization, and is listed as a Class I infectious ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
Inventor 李倬刘涛郁宏伟柳珊刘超梁武朱秀同杨保收李建丽
Owner RINGPU (BAODING) BIOLOGICAL PHARMACEUTICAL CO LTD