Production of a fragment of glycoprotein G of herpes simplex virus type 2 and evaluation of its diagnostic potential

Abstract

Introduction: Herpes simplex virus type 2 (HSV-2) is the most common cause of genital herpes. Glycoprotein G (gG) is a prototype antigen for type-specific serodiagnosis distinguishing between HSV type 1 (HSV-1) and HSV-2 infections. As immunological diagnosis kits for accurate differentiation between HSV-1 and HSV-2 antibodies can be expensive, there is a need to develop a convenient, sensitive, specific and cost-effective serodiagnostic kit.

Methods: We successfully expressed a fragment of gG comprising residues 321-580 of HSV-2 with histidine tag (gG(321-580His)) in a Bac-to-Bac baculovirus expression system, which had an antigenicity similar to its native counterpart. An indirect enzyme-linked immunosorbent assay (ELISA) was developed using gG(321-580His) as the diagnostic antigen and evaluated by comparison with a commercial HerpeSelect 2 ELISA immunoglobulin G kit as reference.

Results: In testing 318 field serum samples, the diagnostic relative sensitivity and specificity of the developed gG(321-580His)-ELISA test in qualitative comparison with the commercial kit were 93.81% and 96.74%, respectively, and the accuracy was 94.65%.

Conclusion: The study indicates that gG(321-580His) has a high diagnostic potential for HSV-2 virus serodiagnosis in humans.

Keywords: baculovirus expression system; glycoprotein G; herpes simplex virus type 2; immunodominant fragment; indirect enzyme-linked immunosorbent assay.

Fig. 1

Agarose gel electrophoresis of the polymerase chain reaction (PCR) amplification products, for the identification of pFastBac HTc-gG_321–580His. Lane M: DNA marker; lane 1: pFastBac HTc-gG_321–580His digested by BamHI; lane 2: pFastBac HTc-gG_321–580His digested by HindIII; lane 3: pFastBac HTc-gG_321–580His digested by BamHI/HindIII; lane 4: PCR products of pFastBac HTc-gG_321–580His using gG_321–580-specific primers.

Fig. 2

Agarose gel electrophoresis of the polymerase chain reaction (PCR) amplification products, for the identification of recombinant Bacmid-gG_321–580His. Lane M: DNA marker; lane 1: PCR products of Bacmid-gG_321–580His using M13 primers; lane 2: PCR products of wild Bacmid using M13 primers; lane 3: PCR products of Bacmid-gG_{321–580 His} using gG_321–580-specific primers.

Fig. 3

(a) SDS-PAGE of the lysates of the non-infected Sf9 cells and the baculovirus-infected Sf9 cells. Lane 1: lysates of the non-infected Sf9 cells; lane 2: lysates of the baculovirus-infected Sf9 cells; lane 3: purified recombinant gG_321–580His. The lysates were separated using 12% SDS-PAGE. Additional bands (arrows) were observed in lanes 2 & 3 after Coomassie blue staining. A protein marker is shown at the left part of the Coomassie-stained gel (lane M). (b) Western blot analysis of the recombinant gG_321–580His. Lane a: protein marker; lane b: non-infected Sf9 cells culture; lane c: baculovirus-infected Sf9 cells culture; lane d: purified recombinant gG_321–580His. A single band was observed in both lanes c & d, which showed that the recombinant protein gG_321–580His was able to react with the specific mouse anti-gG-2 monoclonal antibody. No Western blot signal was observed in lanes a & b (negative controls), indicating that the recombinant protein gG_321–580His has similar functions with its natural counterpart.