Hbeag reagent kit with hbeag quantitative calibrators

HBV uptake assay and infection were performed as previously described^{10 (link),32 (link)}. HBV stocks were prepared by heparin column purification and subsequent sucrose gradient centrifugation as described^{51 (link)}. DNA-containing, enveloped particles (virions) were determined for calculation of the MOI. Qualitative Hepatitis B e antigen (HBeAg) measurement was performed using a commercial immunoassay (BEP III, Siemens Molecular Diagnostics, Marburg, Germany). Sample/Cut-off was determined using internal Cut-off value where samples > 1 are considered as positive. Quantitative HBeAg analysis was obtained using the HBeAg Reagent Kit with HBeAg Quantitative Calibrators on the Architect™ platform (Abbott Laboratories, Chicago, IL, USA).

Peripheral blood was collected into Microvette 500 LH-Gel tubes (Sarstedt) and centrifuged to separate serum (10 minutes, 5000 g, RT). For measurement of ALT levels, serum was diluted 1:4 with PBS and the Reflotron ALT test was used (Roche Diagnostics). Serum HBs- and HBe-antigen levels were determined on an Architect™ platform using the quantitative HBsAg test (6C36-44; cut-off, 0.25 IU/mL) and the HBeAg Reagent Kit with HBeAg Quantitative Calibrators (7P24-01; cut-off, 0.20 PEI U/mL) (Abbott Laboratories). Serum levels of IFN-γ, TNF-α, IL-6 and MCP-1 were detected via the cytometric bead array mouse inflammation kit (BD) according to the manufacturer’s instructions and measured on a CytoFLEX S flow cytometer (Beckman Coulter). Data analysis was done with the Prism 8 software.