Background: Ultrasensitive HBsAg assays are replacing the previous versions. Unlike the sensitivity, the specificity, and its positioning to resolve weak-reactives (WR) are not studied. We investigated the ability of ARCHITECT HBsAg-Next (HBsAg-Nx) assay to resolve WR and sought its clinical validation and correlation with confirmatory/reflex testing.
Methods: Among 99,761 samples between Jan 2022 - 2023, 248 reactive samples in HBsAg-Qual-II were compared with HBsAg-Nx assay. Sufficient samples were further subjected to neutralization (n = 108) and reflex (anti-HBc total/anti-HBs antibody) testing.
Results: Out of 248 initial reactive samples in HBsAg-Qual-II, 180 (72.58%) were repeat reactive, and 68 (27.42%) were negative, whereas in HBsAg-Nx, 89 (35.89%) were reactive and 159 (64.11%) were negative (p<0.0001). Comparing the results of two assays (Qual-II/Next), 57.67% (n = 143) were concordant (++/-) and 105 (42.33%) were discordant (p = 0.0025). Testing of HBsAg-Qual-II + & HBsAg-Nx - samples revealed that 85.71% (n = 90) were anti-HBc total negative and 98.08% (n = 51) were not neutralized as well as significant proportion (89%) had no clinical correlation. The proportion of samples neutralized was significantly different between ≤5 S/Co (26.59%) and >5 S/Co (71.42%) (p = 0.0002). All samples (n = 26) with enhanced reactivity in HBsAg-Nx were effectively neutralized, while samples with no increase in reactivity, 89% (n = 72) failed neutralization (p=<0.001).
Conclusions: HBsAg-Nx assay is positioned better to resolve and refine challenging WR samples than Qual-II which correlated well with confirmatory/reflex tests and clinical disease. This superior internal benchmarking significantly reduced the cost and quantum of retesting, confirmatory/reflex testing in the diagnosis of HBV infection.
Keywords: Anti-HBc total and neutralization; False positive; HBsAg-Next; HBsAg-Qual-II; Weak reactive.
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