Hbsag and hbeag cmia kits

HBV replicative intermediates (HBV RI) from intracellular core particles, and total cellular RNA were extracted from cells or liver tissue as described previously33 (link). Samples were electrophoresed into agarose gels and blotted onto Hybond-XL membranes (GE Healthcare, Marlborough, MA, USA). Then, the membranes were probed with a DIG-labeled specific full-length HBV riboprobe, generated using the PCR DIG Probe Synthesis Kit (Roche, Basel, Switzerland), and visualized with CDP-Star using an ImageQuant LAS 2000 mini system (GE Healthcare). The densitometry of nucleic acid bands were analyzed by Image-Pro Plus software (Media Cybernetics Inc, Silver Spring, MD) and the values were presented below each of the blots as the percentage of control samples. The levels of HBsAg and HBeAg in culture supernatants were determined using the Architect system and HBsAg and HBeAg CMIA kits (Abbott Laboratories, North Chicago, IL, USA) according to the manufacturer’s instructions.

HBV replicative intermediates from intracellular core particles and HBV transcripts were extracted from hepatoma cell lines and detected by Southern and northern blotting, respectively, according to previously published protocols17 (link). HBV progeny DNA was extracted from cell culture supernatants using the QiAamp DNA Blood Mini kit (Qiagen, Hilden, Germany) and quantified by real-time PCR as described17 (link). HBV RNA in cells was also detected using quantitative real-time RT-PCR (the primers sequences are listed in Supplemental Table 2). A monoclonal antibody (clone 10E11, Santa Cruz Biotechnology, CA) was used to detect intracellular HBcAg expression and HBV capsid by western blotting, as described below. The levels of HBsAg and HBeAg in culture supernatants were determined using Architect system and HBsAg and HBeAg CMIA kits (Abbott Laboratories, Wiesbaden-Delkenheim, Germany) according to the manufacturer’s instructions.