Porcine circovirus type 2 antigen capture ELISA kit
A technology of porcine circovirus and kit, applied in the field of immunology, can solve the problems of time-consuming operation, poor specificity, and high technical requirements, and achieve the effect of short time-consuming and simple operation
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Embodiment 1
[0055] Example 1 Preparation of monoclonal antibody against porcine circovirus type 2 (PCV2) Cap protein
[0056] 1. Materials and methods
[0057] 1.1 Cells, virus species, experimental animals and reagents
[0058] SP2 / 0 and PK15 cell lines, PCV1 / G strains, PCV2 strains (LG, JF, JF2, CL, YJ, SH, BDH, AH, etc.) are all preserved by the Porcine Circovirus Research Group of Harbin Institute of Veterinary Medicine, Chinese Academy of Agricultural Sciences ; 6-week-old female BALB / c mice were provided by the Experimental Animal Center of Harbin Veterinary Research Institute, Chinese Academy of Agricultural Sciences; HAT and HT mixed salt and PEG1450 were purchased from Sigma; pre-stained protein Marker was purchased from Fermentas; Mouse MonoAb-ID Kit( HRP), HRP-Goat Anti-Mouse IgG (H+L) were purchased from Invitrogen; other reagents are domestically pure.
[0059] 1.2 Purification of Porcine Circovirus Type 2 (PCV2)
[0060] The PCV2a / LG strain was purified by CsCl gradient density cent...
Embodiment 2
[0082] Example 2 Preparation of polyclonal antibodies against porcine circovirus type 2 (PCV2)
[0083] 1 Materials and methods
[0084] 1.1 Test materials
[0085] Porcine circovirus type 2 (LG strain) inactivated vaccine was provided by Harbin Veken Biotechnology Development Company. PCV2 / YJ strain (10 5.0 TCID 50 / ml) Preserved by the Porcine Circovirus Research Group of Harbin Institute of Veterinary Medicine, Chinese Academy of Agricultural Sciences, 6 piglets 30-40 days old (PCV2-IPMA serum antibody titer is less than or equal to 1:50 times) by Harbin Institute of Veterinary Medicine, Chinese Academy of Agricultural Sciences Provided by the Laboratory Animal Center.
[0086] 1.2 Test method
[0087] 1.2.1 Animal immunity test
[0088] Six piglets were randomly divided into a test group and a control group, with 3 pigs in each group and reared separately. In the first immunization, piglets were injected with 1 mL of inactivated porcine circovirus type 2 (LG strain) vaccine into th...
Embodiment 3
[0102] Example 3 Establishment of an ELISA method for capturing porcine circovirus type 2 antigen
[0103] 1. Materials and methods
[0104] 1.1 Material
[0105] ABTS was purchased from BBI, DMEM medium and fetal bovine serum (FBS) were purchased from Gibco, and other reagents were domestically pure.
[0106] Preparation of ABTS working solution: (1) Chromogenic A solution (0.1mol / L citrate buffer) preparation: weigh 29.41g sodium citrate (Na 3 C 6 H 5 O 7 ·2H 2 O) Dissolve in 1800mL deionized water, adjust the pH to 4.3 with citric acid, and then add 200μL 30% H 2 O 2 , And finally dilute the volume to 2000mL with deionized water; (2) Preparation of color developing solution B (2.19% ABTS solution): weigh 2.19g ABTS powder, fully dissolve it with 0.1mol / L citrate buffer, and dilute to 100mL; (3) Preparation of ABTS working solution: add 0.5ml of chromogenic A solution to 9.5mL of chromogenic A solution, mix well and use, ready to use.
[0107] 1.2 monoclonal antibody purification and...
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