Characterization of serum IgM and development of an indirect ELISA for evaluation of antigen-specific IgM in serum and skin mucus of Nile tilapia (Oreochromis niloticus)

The study aimed to characterize the serum immunoglobulin M (IgM) and develop an enzyme-linked immunosorbent assay (ELISA) for antigen-specific IgM assessment in Nile tilapia (Oreochromis niloticus). IgM was purified using ammonium sulfate precipitation and protein A chromatography. Native gradient polyacrylamide gel electrophoresis (PAGE) revealed a ~878.4 kDa band with purified IgM, while sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed heavy and light chains of ~82.2 and ~28.2 kDa, respectively. A rabbit anti-Nile tilapia IgM was produced and its specificity verified. The antiserum also cross-reacted with IgM from Indian major carps (IMCs). One indirect ELISA was developed with this antiserum to quantify IgM concentration (1.57 ± 0.07 mg/ml) in the Nile tilapia serum. Another indirect ELISA was standardized to assess antigen-specific IgM levels in serum and skin mucus over 90 days following immunization with inactivated Enterobacter cloacae. This ELISA could be useful for assessing immune responses during infection or vaccination in tilapia.