Enhancing expression of the pseudorabies virus glycoprotein E in yeast and its application in an indirect sandwich ELISA

Aims The purpose of this study was to produce a recombinant pseudorabies virus (PRV) glycoprotein E (gE) protein with the correct antigenicity for use as a low-cost diagnostic antigen. Methods and Results The gene fragment encoding the amino-terminal immunodominant region of PRV gE (codons 31–270) (gEN31–270) was codon-optimized and expressed constitutively and secreted using a Pichia pastoris expression system. Yeast-expressed gEN31–270 (ygEN31–270) was harvested from the culture supernatant, and ygEN31–270 was shown to exhibit N-linked glycosylation. An indirect sandwich enzyme-linked immunosorbent assay (ELISA) was developed using ygEN31–270 as a coating antigen, and the results showed that the assay had high sensitivity and specificity, as well as almost perfect concordance with a commercial gE ELISA kit. Conclusions The immunodominant region (amino acids 31–270) of gE was expressed successfully in P. pastoris using a codon optimization strategy. ygEN31–270 was secreted and N-glycosylated. The ygEN31–270-based indirect sandwich ELISA showed high sensitivity and specificity to detect gE-specific antibodies in swine serum samples. Significance and impact of Study The ygEN31–270-based indirect sandwich ELISA may provide an alternative method for developing a diagnostic kit with easy manipulation and low cost. This article is protected by copyright. All rights reserved.