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Pullorum staining agglutination antigen as well as preparation method and application thereof

A technology for dyeing agglutination antigens and pullorum, applied in biochemical equipment and methods, microorganism-based methods, instruments, etc., can solve the problems of inability to achieve complete purification of breeder flocks, insufficient agglutination images, and economic losses of farmers, etc. To achieve the effect of simple and reasonable process method, good antigenicity and low cost

Active Publication Date: 2015-07-22
JIANGSU INST OF POULTRY SCI
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Problems solved by technology

Although there are plate-stained antigen products for the detection of pullorum in my country, such as "polyvalent staining plate agglutination test antigen for pullorum and typhoid fever", this antigen has low sensitivity, slow diagnosis speed, and unclear agglutination images in practical applications. Defects, resulting in the detection of breeder flocks can not achieve the goal of complete purification, causing serious economic losses to farmers

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  • Pullorum staining agglutination antigen as well as preparation method and application thereof
  • Pullorum staining agglutination antigen as well as preparation method and application thereof
  • Pullorum staining agglutination antigen as well as preparation method and application thereof

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Embodiment 1

[0018] 1) Separation, purification and staining microscopy

[0019] Salmonella pullorum SP9905 strain was isolated from the cecum contents of 11-month-old Langshan chickens in a chicken farm in Yangzhou City, Jiangsu Province in May 1999. The specific separation method is as follows: aseptically collect cecal contents, add them to selenite cystine enrichment solution for overnight culture at 37°C, then inoculate MacConkey agar at 37°C for 24 hours, pick out 0.5mm-1.0mm in diameter, Off-white transparent colonies with smooth surface, inoculated on Martin agar medium, cultured at 37°C for 24 hours. The purely cultured bacteria were subjected to Gram staining microscope inspection, and then observed with an optical microscope, showing elongated bacilli with slightly rounded ends.

[0020] Salmonella pullorum sp8441 strain was isolated from the oviduct of 6-month-old white legion laying hens in a chicken farm in Beijing in December 1984. The specific separation method is as foll...

Embodiment 2

[0042] Salmonella pullorum SP7220, SP7701, SP8441 and SP9905 freeze-dried strains of Salmonella pullorum sp7220, SP7701, SP8441 and SP9905 were respectively resuscitated and multiplied in liquid medium, and passed continuously for 50 generations indiscriminately on the artificial medium. Select 50 single colonies, observe and record the colony morphology, and use Salmonella O12 2 and O12 3 Single factor determination of serotype. The variation rate of colony type and serotype was counted after every 10 passages.

[0043] The results are shown in Table 3. After 20 consecutive subcultures of the 4 antigen-producing strains, only 2% of the colony type of SP9905 varied; after 30, 40, and 50 generations of subculture, the average variation rates of the colony types were 3.5% and 11%, respectively. and 17.5%, and the average variation rates of serotypes were 2%, 6% and 11%, respectively. The above results show that the four antigen-producing strains selected in the present invent...

Embodiment 3

[0047] Salmonella pullorum SP7220, SP7701, SP8441 and SP9905 strains were resuscitated and multiplied in liquid medium respectively, inoculated on solid plate medium, cultured at 37°C for 22 hours, and typical colonies were selected to prepare seed bacterial liquid. The seed bacteria solution was inoculated into the solid flat flask culture medium according to 4.0% of the medium volume, and after being cultivated at 37°C for 48 hours, the bacterial lawn was washed with 0.3% formalin normal saline, and the bacterial solution was collected after filtering through sterile gauze, and placed Inactivate overnight at 4°C; centrifuge the inactivated bacterial solution at 4000r / min for 15 minutes to remove the supernatant, then resuspend the bacterial sludge with 0.2% formalin saline, and adjust the turbidity of the 4 antigen-producing strains to each ml 1.2 x 10 10 After the bacterial concentration of CFU is mixed thoroughly according to the proportion. Add 2.0% crystal violet soluti...

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Abstract

The invention discloses a pullorum staining agglutination antigen as well as a preparation method and an application thereof, and belongs to the technical field of veterinarian diagnosis. Salmonella pullorum SP7220, SP7701, SP8441 and SP9905 are subjected to recovery and passage respectively to form a seed bacterium liquid, then the seed bacterium liquid is inoculated with a solid culture medium for proliferation, after inactivation with formalin, the concentration of the bacterium liquid is adjusted with turbidimetry, finally, the bacterium is fully and evenly mixed and packaged after a crystal violet solution is added for staining, and the pullorum staining agglutination antigen is prepared. The technological method is simple, reasonable, scientific, stable in production and low in cost, the selected production stains of the pullorum staining agglutination antigen are good in antigenicity and low in mutation rate, the prepared pullorum staining agglutination antigen product has the advantages of high sensitivity, high specificity, quickness in diagnosis and clearness in agglutination image, the staining agglutination antigen is relatively ideal for pullorum detection.

Description

technical field [0001] The invention belongs to the technical field of veterinary diagnosis, and in particular relates to a pullorum stained agglutination antigen and a preparation method and application thereof. Background technique [0002] Pullorum pullorum is one of the important diseases that seriously endanger the healthy development of the chicken industry. It can cause high morbidity and mortality in chicks and reduce the production performance of adult chickens. Egg transmission is the main route of the disease. Therefore, it is the key to control the disease to test the breeders and their eggs in a planned way, eliminate positive chickens, cultivate breeder flocks without pullorum, and cut off the vertical transmission route. Domestic and foreign purification methods for pullorum pullorum mainly adopt whole blood or serum glass plate agglutination test, and positive chickens are eliminated in time until the positive rate of the whole flock does not exceed 0.1%. Al...

Claims

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Application Information

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IPC IPC(8): C12N1/20G01N33/569C12R1/42
CPCC12N1/20C12N1/205C12R2001/42G01N33/569
Inventor 龚建森许明苗晋锋张笛顾蓓蓓朱春红窦新红刘学贤徐步童海兵邹剑敏
Owner JIANGSU INST OF POULTRY SCI
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