Expression, purification, and diagnostic evaluation of the recombinant matrix protein of peste des petits ruminants...

Peste des petits ruminants virus (PPRV) causes a highly contagious and economically devastating disease of small ruminants, particularly sheep and goats. Serological evaluation using recombinant protein-based Enzyme-linked immunosorbent assays (ELISAs) is essential for monitoring PPRV infection and assessing eradication programs. Although recombinant nucleoprotein (N) is widely used for serological evaluation, investigation of additional viral proteins may provide alternative antigenic platforms for PPRV antibody detection. The matrix (M) protein plays a central role in viral assembly by linking viral surface glycoproteins with ribonucleoprotein complexes. However, recombinant expression of full-length M protein in Escherichia coli (E. coli) is challenging because of low or truncated expression and inclusion body formation, often requiring complex refolding procedures. In this study, full-length M protein from the Nigeria 75/1 vaccine strain was successfully overexpressed as inclusion bodies (IBs) in E. coli and solubilized using N-lauroyl sarcosine (NLS) without refolding. The solubilized protein was purified by nickel-affinity chromatography, yielding 2.1 mg/L bacterial culture, and confirmed as a 38-kDa protein by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting. An indirect ELISA was developed using purified M protein and optimized with commercial control sera. Optimal conditions were achieved with 1 µg/mL antigen and 1:20 serum dilution. Using pre-confirmed field sera, the assay demonstrated 94.59% sensitivity and 92.45% specificity at a cut-off of 0.25. Comparison with the commercial ID Screen^® PPR Competition ELISA showed comparable performance and strong agreement. These findings demonstrate the recombinant PPRV M protein as a feasible diagnostic antigen for PPRV antibody detection which requires further validation for large scale application.