Development of a Microplate Enzyme Immunoassay for the Initial Screening of Monoclonal Antibodies Against Infectious Hematopoietic Necrosis Virus.

Abstract

A microplate enzyme immunoassay (MEIA) was developed for the rapid, sensitive and specific detection of monoclonal antibody-producing hybridomas against IHNV at the first screening. Ninety-six-well microplate cultures of chinook salmon embryo (CHSE-214) cells were infected with IHNV and incubated at 15 degree C to permit antigen development. The hybridoma supernatants were added to the plates containing 95% ethanol-fixed IHNV-infected CHSE-214 cells as the first antibodies. Peroxidase-conjugated antimouse Ig goat serum was applied as the second antibody and visualization was performed using diamino-benzidine. Under microscopy, positive antigen-antibody reaction was apparently observed by a brownish staining of IHNV-infected CHSE-214 cells. Thus, this method was found highly efficient and sensitive for the screening of monoclonal antibody.