Viruses, Vol. 16, Pages 393: Quantification of Hepatitis E Virus ORF2 Protein by a Novel Sandwich ELISA

Viruses, Vol. 16, Pages 393: Quantification of Hepatitis E Virus ORF2 Protein by a Novel Sandwich ELISA Viruses doi: 10.3390/v16030393 Authors: Sakthivel Subramaniam Rafaelle Fares-Gusmao David R. McGivern Hepatitis E virus (HEV) causes acute hepatitis in humans, which can progress to chronicity in immunosuppressed individuals. Almost all reported HEV infections are caused by Paslahepevirus balayani genotypes 1–4. The structural ORF2 protein is the major antigen detected in the blood of HEV-infected individuals. ELISA assays to detect IgM antibodies to HEV are the first-line diagnostic tests; however, they showed variable performance with frequently discordant results. A qualitative HEV antigen (ORF2) ELISA is currently available for research use. Here, we report a novel quantitative sandwich ELISA to measure HEV ORF2 protein in 3 matrix types. An optimal pair of capture and detection antibodies was selected among 12 unique combinations tested. A sandwich ELISA protocol was developed using these mAbs and biotin–streptavidin technology. The protocol was further optimized to quantify ORF2 antigen in different matrices by interpolating from a standard curve with a linear range of 3.17 to 50.8 femtomoles/mL. Using this method, ORF2 protein was detected in the cell culture medium of Huh7 cells as early as 2–3 days after transfection with HEV genome RNA and in a medium of human hepatocytes infected with HEV. ORF2 an...
Source: Viruses - Category: Virology Authors: Tags: Article Source Type: research