Journal: Cellular and Molecular Gastroenterology and Hepatology
Article Title: Presence of Intact Hepatitis B Virions in Exosomes
doi: 10.1016/j.jcmgh.2022.09.012
Figure Lengend Snippet: Inoculation of HepG2- and differentiated HepaRG cells by exosomal HBV and free HBV. A , HBV genome copy number in each exosomal and viral inoculum is shown in the left . Schematic description of the infection experiment procedure is shown in the right . B , HBsAg ELISA of supernatant derived from HepG2 cells inoculated either with exosomal HBV virions or free HBV virions. Medium was changed at the indicated time points and analyzed by HBsAg ELISA. The horizontal dotted line indicated the cutoff value. C , HBsAg ELISA of supernatant derived from differentiated HepaRG cells inoculated either with exosomal HBV (fraction 8 of the iodixanol gradient) or a less pure, later exosomal fraction (fraction 10). The inoculum was preincubated with the LHBs-specific antibody MA18/7 (1 μg/mL) or an anti-hexa-His-specific monoclonal antibody (1 μg/mL) as control. Medium was changed at the indicated time points and analyzed by HBsAg-specific ELISA. S/CO, Sample to cutoff signal. The cutoff value is represented by the horizontal dotted line . D , HBsAg ELISA of supernatant derived from differentiated HepaRG cells inoculated with free HBV virions (fractions 13 and 14). The inoculum was preincubated with the LHBs-specific antibody MA18/7 (1 μg/mL) or an anti-hexa-His-specific monoclonal (1 μg/mL) as control. Medium was changed at the indicated time points and analyzed by HBsAg-specific ELISA. The horizontal dotted line delineates the cutoff value. E , HBsAg ELISA of supernatant derived from differentiated HepaRG cells inoculated either with exosomal HBV (fractions 7, 8 of the iodixanol gradient) or a less pure, later exosomal fraction (fractions 9, 10 of the iodixanol gradient). The inoculum was preincubated with the LHBs-specific antibody MA18/7 (1 μg/mL) or an anti-hexa-His-specific monoclonal (1 μg/mL) as control. Here, the HBsAg content at day 4 of the different samples is analyzed. Unpaired parametric t tests for all panels; ∗∗ P < .01; ∗∗∗ P < .001; ns, not significant. F–G , Differentiated HepaRG cells were inoculated with free HBV virions (fractions 13) ( F ) or exosomal HBV (fraction 8 of the iodixanol gradient) ( G ), and secreted HBsAg was quantified at the indicated time points by HBsAg-specific ELISA. Myrcludex B 500 nM was added during infection. The horizontal dotted line indicates the cutoff value.
Article Snippet: To the purified HBV exosomal fractions or free HBV virus fractions, the RBD-binding anti-preS1 antibody (MA18/7, 1 μg/mL) or an unrelated monoclonal anti-hexa-His antibody (sc-8036, Santa Cruz) was added and kept at 37 °C for 2 hours.
Techniques: Infection, Enzyme-linked Immunosorbent Assay, Derivative Assay, Control
