Optimization and Application of Quantitative In-house ELISA for Diagnosis of HBsAg and its Correlation with Commercial ELISA and Molecular Kits

 Several studies have indicated the role of quantitative hepatitis B surface antigen (HBsAg) evaluation in managing and prognosis of hepatitis B virus (HBV) infection. Thus, quantitative evaluation of HBsAg using cost-effective assays can be an important approach to managing HBV patients.

 This study aimed to set up and apply an in-house quantitative enzyme-linked immunosorbent assay (ELISA) to evaluate HBsAg in research and diagnosis.

 New Zealand white rabbits were immunized with HBsAg. Sera were collected 28 days after immunization, and polyclonal HBsAb (antibody to hepatitis B surface antigen) were purified and evaluated. Then, the in-house quantitative ELISA was optimized. Finally, the functional characteristics of the assay were evaluated using 200 plasma samples compared to commercial ELISA and quantitative TaqMan real-time PCR.

 The assay has a limit of detection (LOD) of 0.5 ng/mL with a specificity and sensitivity of 94% and 97%, respectively. The assay's highest coefficient of variation (CV) values for intra- and inter-assays were 7.23% and 8.59%, respectively. The correlation of the developed assay with commercial ELISA was 0.987 (P-value = 0.024). The correlations of the developed assay and commercial ELISA with quantitative TaqMan real-time PCR were 0.739 and 0.658, respectively (P-value = 0.017).

 The developed assay has a suitable sensitivity and specificity. It is also reproducible and well-correlated with commercial assays. Most importantly, it is cost-effective and, thus, can be used for detecting and quantifying HBsAg in research and diagnosis.