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Simultaneous Quantification of Hepatitis C Virus Envelope Glycoproteins E1 and E2 by Dual-Color Fluorescence Immunoblot Analysis

  • Fang Chen
  • Erick Giang
  • Radhika Gopal
  • Mansun LawEmail author
Protocol
Part of the Methods in Molecular Biology book series (MIMB, volume 1911)

Abstract

The hepatitis C virus (HCV) envelope glycoproteins, E1 and E2, are crucial for HCV assembly and entry, and are promising vaccine antigens. However, they are challenging to study because of technical difficulties in protein production and in quality control for protein folding and glycosylation. To study E1 and E2 in different experimental systems, e.g. infected cells, virus culture, virus-like particles, and clinical samples, a standardized method to accurately quantify the glycoproteins will be essential for most research projects. Here we outline a sensitive assay based on dual-color fluorescence immunoblot and the Odyssey imaging system to detect and quantify HCV E1 and E2 glycoproteins either using a purified E1E2 complex, or an engineered protein standard containing E1 and E2 at equal molar ratio. The method is capable of simultaneously detecting and quantifying as little as 7 ng of E1 and 5 ng of E2 in HCV pseudoparticles, and will be useful to quantify E1 and E2 from a wide variety of samples.

Key words

E1 E2 Glycoprotein Immunoblot Protein standard Antibody 

Notes

Acknowledgments

This work was supported by NIH grants AI079031, AI106005, AI123365, and AI123861.

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2019

Authors and Affiliations

  • Fang Chen
    • 1
  • Erick Giang
    • 1
  • Radhika Gopal
    • 1
  • Mansun Law
    • 1
    Email author
  1. 1.Department of Immunology and MicrobiologyThe Scripps Research InstituteLa JollaUSA

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