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

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 (<i>Oreochromis niloticus</i>). 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 <i>Enterobacter cloacae</i>. This ELISA could be useful for assessing immune responses during infection or vaccination in tilapia.