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. 2017 Nov 30;7:16616. doi: 10.1038/s41598-017-16882-5

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© The Author(s) 2017

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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HBV infection and optimization in HepG2-NTCP cells. (A) Immunofluorescence microscopy to detect NTCP in HepG2-NTCP. (B) Schematic of the experimental design. (C) Optimization of HBV infection in HepG2-NTCP cells. Cells were infected with 100 GEQ/cell and the following conditions were optimized by measuring HBeAg in cell supernatants by ELISA X days post infection: duration of infection, DMSO treatment, spinoculation, and seeding cell density (96-well plate). In all conditions, 4% PEG was present in the inoculum, then washed out one day post-infection. At least three biological replicates were performed and data are shown as means ± s.d. (one-way ANOVA for days post-infection and cell density, two-tailed unpaired t test for DMSO and spinoculation, **P < 0.001).