Reuschel, Edith and Jilg, Wolfgang and Seelbach-Goebel, Birgit and Deml, Ludwig (2019) Comparative purification and characterization of hepatitis B virus-like particles produced by recombinant vaccinia viruses in human hepatoma cells and human primary hepatocytes. PLOS ONE, 14 (2): e0212800. ISSN 1932-6203,
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This study describes the comparative expression and purification of hepatitis B surface antigen (HBsAg) particles produced upon infection of human primary hepatocytes and human hepatoma cell lines (HuH-7 and HepG2) with recombinant vaccinia viruses. The highest levels of HBsAg expression were found in HuH-7 hepatoma cells following infection with recombinant vaccinia viruses, which contain the S gene under control of a 7.5 k-promoter. Four different methods for purification of the HBsAg particles were examined: isopycnic ultracentrifugation, sucrose cushion sedimentation, isocratic column gel filtration, and binding to anti-HBs-coated microparticles. The highest degree of purity of HBsAg particles was reached by the method based on anti-HBs-coated microparticles. The resulting product was > 98% pure. Biochemical analysis and characterization of purified HBsAg particles were performed by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE), western blotting, and electron microscopy. The HBsAg, purified from human hepatoma cell lines and from human primary hepatocytes, consisted of both the non-glycosylated (p25) and the glycosylated (gp27) form and assembled into typical 22-nm particles, and thus may be of great interest and importance for research, diagnostics, and medical treatments.
| Item Type: | Article |
|---|---|
| Uncontrolled Keywords: | LONG-TERM CULTURE; SURFACE-ANTIGEN; METHYLOTROPHIC YEAST; HBSAG PARTICLES; INSECT CELLS; EXPRESSION; IMMUNOGENICITY; GLYCOPROTEIN; PROTEINS; EFFICACY; |
| Subjects: | 600 Technology > 610 Medical sciences Medicine |
| Divisions: | Medicine > Lehrstuhl für Frauenheilkunde und Geburtshilfe (Schwerpunkt Geburtshilfe) Medicine > Lehrstuhl für Medizinische Mikrobiologie und Hygiene |
| Depositing User: | Dr. Gernot Deinzer |
| Date Deposited: | 17 Apr 2020 06:24 |
| Last Modified: | 17 Apr 2020 06:24 |
| URI: | https://pred.uni-regensburg.de/id/eprint/27524 |
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