Development of monoclonal antibodies for measurement of immunoglobulin G antibody responses in Blue and Gold Macaws (Ara ararauna)

Abstract

Objectives - To produce monoclonal antibodies (MAB) with specificity for the heavy chain of macaw IgG; to incorporate these MAB into an ELISA to measure IgG responses of macaws inoculated with bovine serum albumin (BSA); and to evaluate the antigenicity of BSA in Blue and Gold Macaws. Animals - Four 1-year-old Blue and Gold Macaws, 2 males and 2 females. Procedure - 1 male and 1 female 1 were randomly assigned to each of 2 study groups. Group-1 birds were inoculated with 200 μg of BSA on days 0, 14, 28, and 42. Group-2 birds were inoculated with 200 μg of BSA on days 0 and 28. Blood was collected weekly for measurement of anti-BSA titer. Hybridomas were prepared from mice immunized with Scarlet Macaw (Ara macao) IgG purified by salt precipitation and gel chromatography. Specificity for IgG of Scarlet Macaw and other macaw species was confirmed by ELISA and western blot analysis. Hybridoma HL544 was cloned and the antibody purified. Following biotinylation, MAB HL544 was incorporated into an ELISA that measured IgG responses of macaws inoculated with BSA. Results - Adult Blue and Gold Macaws developed strong primary and secondary anti-BSA antibody titers 14 days after inoculation with 200 μg of BSA. An inoculation interval of 28 days resulted in stronger secondary responses than an interval of only 14 days. Conclusions - MAB specific for macaw immunoglobulins can be used in ELISA to evaluate the humoral immune responses of macaws. 1-year-old Blue and Gold Macaws developed strong anti-BSA titer when inoculated with 200 μg of BSA. An inoculation interval of 28 days resulted in stronger secondary responses than did an interval of only 14 days. Clinical Relevance - These MAB, the first reported to have specificity for a psittacine antibody class, will be useful in the evaluation of psittacine antibody responses and in the development of psittacine diagnostic assays.