Anti-immunoglobulin binding and activation of snapper (Pagrus auratus) leucocytes

Abstract

In order to perform specific immunological assays we have produced and characterised three monoclonal antibodies (MAbs) that bind snapper (Pagrus auratus, Bloch and Schneider) immunoglobulin (Ig). Hybridomas were produced and screened for anti-Ig production using ELISA, Western blot and flow cytometry. All three MAbs (designated 2C5, 4A2 and 1C6) bound specifically to the heavy (H) chain of reduced Ig in Western blot. Furthermore, 1C6 was shown to bind to reduced skin mucus Ig H chain and all three MAbs cross-reacted with the H chain of Atlantic salmon and rainbow trout Ig. In flow cytometric analyses 2C5 and 4A2 bound to B cell populations in the peripheral blood and lymphoid organs. Furthermore, cross-linked 2C5 induced an increase in intracellular protein tyrosine phosphorylation in peripheral blood lymphocytes. Phosphorylated proteins exhibited similar molecular weights to those of mammalian Igalpha and Igbeta and may represent snapper mIg accessory molecule analogues. These data exhibit the potential use of 2C5, 4A2 and 1C6 in both cellular and biochemical analyses of populations of snapper leucocytes.