Fig. 5.

The changes in immuno-reactivity over time during the field trial evaluation of prototype poultry red mite vaccines.

Individual lanes of a western blot of soluble mite extract (SME) protein were probed with pooled egg yolks from hens from trial 1 assessing a SME prototype vaccine (panel A) or with pooled serum from trial 2 testing a recombinant antigen cocktail prototype vaccine (panel B). Pooled yolk or serum from Vaccinated hens (VAC) and the Placebo hens (PLAC) obtained at several time points throughout the trials were tested (trial 1: weeks 20, 27, 29 and 36; trial 2: weeks 12,17,19,23,27,31,35 and 38). Bound IgY was detected with rabbit anti-IgY-peroxidase conjugated antibody (Sigma, UK). PBS was substituted for the primary IgY and served as a conjugate control (conj, panel B). Additional controls were performed for trial 2 (panel C): individual western blot strips of the purified recombinant antigens: Deg-Serpin-1 (SRP), Deg-protein of unknown function-1 (PUF) and Deg-Vitellogenin (VIT) were probed with trial 2 serum pool from the Placebo group (−) and Vaccine group (+) obtained following vaccination (week 17). The bands marked with asterisk (*) indicate the detection of host IgY fragments present in SME. Bands marked with the lines (−) indicate immuno-reactive bands common to both in trial 1 and 2 Vaccine groups that developed following prolonged infestation with poultry red mite. The approximate mol. wt (kDa) of detected proteins were estimated by comparison to SeeBlue^® Plus2 pre-stained standards (GE Healthcare, UK).