2.7. ELISA Determination of OVA-Specific Immunoglobulins
A 96-well plate was coated with OVA at 1 µg/well and then incubated with blocking solution (1% bovine serum albumin in PBS buffer) at room temperature. The wells were added with mouse serum followed by antibodies diluted with the blocking solution (1:50, 1:1000, and 1:10,000 dilutions for IgE, IgG2a, and IgG1, respectively) and incubated at 4 °C overnight. Biotin-conjugated antibodies against mouse IgE, IgG1, or IgG2a (BD Biosciences) were added after the wells were washed with 0.05% Tween-20/PBS buffer. Then, the plate was incubated at room temperature for 2 h. Next, streptavidin-conjugated horseradish peroxidase was added to the wells. 3,3′,5,5′-Tetramethylbenzidine substrate was used for color development. After the termination of the reaction by using 2 N H