Author: Makadiya, Niraj; Brownlie, Robert; van den Hurk, Jan; Berube, Nathalie; Allan, Brenda; Gerdts, Volker; Zakhartchouk, Alexander
Title: S1 domain of the porcine epidemic diarrhea virus spike protein as a vaccine antigen Document date: 2016_4_1
ID: 1r3doeic_47
Snippet: A colostrum sample (1 mL) collected on the day of farrowing was treated with 30 μL rennet (5 mg/mL, Sigma-Aldrich). The colostrum was then incubated at 37°C for 1 h. Once solidified, the whey was separated by centrifugation at 6000 x g for 20 min. Whey samples were diluted in PBS containing 0.05 % Tween 20 and 1 % casein and applied at four-fold dilution on ELISA plate coated with the purified S1 protein. Plate was incubated at room temperature.....
Document: A colostrum sample (1 mL) collected on the day of farrowing was treated with 30 μL rennet (5 mg/mL, Sigma-Aldrich). The colostrum was then incubated at 37°C for 1 h. Once solidified, the whey was separated by centrifugation at 6000 x g for 20 min. Whey samples were diluted in PBS containing 0.05 % Tween 20 and 1 % casein and applied at four-fold dilution on ELISA plate coated with the purified S1 protein. Plate was incubated at room temperature for 2 h. The plates were washed six times with water between each step. Mouse anti-pig IgA (AbD Serotec) was applied to the plate at 1:300 dilution and incubated for 1 h. Donkey anti-mouse HRP conjugate (Jackson Immunoresearch) was applied at 1:5000 dilution and incubated for 1 h, and 1-Step Ultra TMB-ELISA substrate solution (ThermoFisher Scientific) was applied to the plates to develop the reaction. Then, the reaction was stopped and red as described above.
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