Mouse Serum Antibody Screening

Mouse serum samples were screened for BLG and OVM specific IgG1, IgG2a, IgA and IgE as described recently [20 (link)]. Briefly, serum samples were diluted 1:20 for IgE and 1:100 for IgG1, IgG2a and IgA in blocking buffer (TBS containing 0.05% Tween (TBST) and 1% BSA). Detection was performed with rat anti-mouse IgG1, IgG2a, IgA and IgE (all from BD Biosciences, Heidelberg, Germany) were diluted 1:500 in TBS containing 0.05% Tween and 0.1% BSA being followed by peroxidase labeled goat anti-rat IgG (Amersham, Buckinghamshire, UK; 1:1000 in TBST/0.1%BSA). Tetramethylbenzidine (TMB; BD Bioscience) was used as substrate and reaction was stopped with 0.8mol/L H₂SO₄. Antibody titers were calculated according to a standard dilution series ranging from 100 to 0.78 ng/mL for IgE and ranging from 400 to 3.13 ng/mL for all other isotypes.