This study is the first report of the development of
monoclonal antibodies (MAbs) against
gizzerosine (GZ), one of the causative agents of black vomit, a serious
poultry disease. Balb/c mice were immunized with different GZ conjugates; the most immunogenic conjugate in experimental animals was determined by
enzyme-linked
immunoadsorbent assays (ELISA). Somatic fusions were carried out using splenic lymphocytes from GZ-immune mice and the NSO/2 myeloid cell line. Primary selection of hybridomas secreting
antibodies to GZ was done using a direct ELISA, with GZ bound to
bovine serum albumin (BSA), GZ directly bound to maleinimide preactivated plates and
histamine bound to BSA, a GZ related
biogenic amine present in fish meal. Four MAbs--3H4, 3H10, and 5B1 of the
IgG1 isotype, and 8G7 of the Ig2a isotype-were specific to GZ and did not cross-react with
histamine. Only monoclonals 3H4 and 8G7 bound GZ in
solution by means of a competitive ELISA. Finally, to determine the performance of the competitive ELISA developed with the MAbs, experiments were conducted with GZ in
solution (0 to 10 microg/ml) and with GZ labeled with
horseradish peroxidase (HRP) as the tracer; the antibody complex was captured by using rabbit anti-mouse
IgG preactivated ELISA plates. These experiments showed that monoclonal anti-GZ-3H4 generates a more sensitive assay close to linearity in the range about of 0.1 to 10 microg/ml of GZ. No cross-reaction was observed with
histamine,
histidine, or
lysine at all concentrations tested.