Anti-beta-conglycinin monoclonal antibody and application thereof
A conglycinin and monoclonal antibody technology, applied in the field of monoclonal antibodies, can solve problems such as growth performance decline, death, immune dysfunction, etc., and achieve the effect of low detection limit and wide detection range
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Embodiment 1
[0048] Embodiment 1, the acquisition of hybridoma cell line and the preparation of anti-β-conglycinin monoclonal antibody
[0049] 1. Obtaining of hybridoma cell lines
[0050] 1. Animal immunity
[0051] Dissolve β-conglycinin (ie, the immunogen) in 0.5ml of normal saline, add an equal volume of adjuvant to make an emulsifier, and immunize female BALB / c mice (body weight 18-22g). Serum titers were measured 7-10 days after the third immunization. See Table 1 for the immunization steps.
[0052] Table 1. Animal immunization steps
[0053]
[0054] 2. Cell fusion and cloning
[0055] Blood was collected from the tail of the mouse after the third immunization in step 1 to determine the antibody titer, and the mouse with the highest serum titer was selected, and the splenocytes were taken and fused with SP2 / 0 myeloma cells; the cells secreting monoclonal antibodies were screened by limiting dilution method Hybridoma cell line; the stable secreted anti-β-conglycinin monoclo...
Embodiment 2
[0090]Example 2, Competitive ELISA method for detecting β-conglycinin and its special kit
[0091] 1. Checkerboard method to screen the optimal antigen coating concentration and antibody working concentration
[0092] 1. Coating antigen: take β-conglycinin, prepare a solution with a concentration of 1000ng / mL with coating buffer, and then perform 2-fold serial dilution to obtain a concentration of β-conglycinin in order of 1000ng / mL , 500ng / mL, 250ng / mL, 125ng / mL, 62.5ng / mL, 31.2ng / mL, 15.6ng / mL, 7.8ng / mL, 3.9ng / mL, 1.95ng / mL, 0.98ng / mL, 0.49ng 12 kinds of coating solutions per mL; add these 12 kinds of coating solutions in the amount of 0.1ml / well to the columns 1-12 of the microtiter plate in turn, one coating solution for each column, and coat at 4°C for 16h ; Discard liquid.
[0093] 2. Blocking: Add 200 μL / well of blocking solution, block in a 37°C wet box for 1 hour; discard the liquid, wash with washing solution 3 times for 30 seconds each time, and pat dry.
[0094]...
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