Next Article in Journal
The Molecular and Cellular Effect of Homocysteine Metabolism Imbalance on Human Health
Next Article in Special Issue
The Relationship between Serum Bilirubin and Elevated Fibrotic Indices among HBV Carriers: A Cross-Sectional Study of a Chinese Population
Previous Article in Journal
NR2F6 Expression Correlates with Pelvic Lymph Node Metastasis and Poor Prognosis in Early-Stage Cervical Cancer
Previous Article in Special Issue
Potential Susceptibility Mutations in C Gene for Hepatitis B-Related Hepatocellular Carcinoma Identified by a Two-Stage Study in Qidong, China
Article Menu
Issue 10 (October) cover image

Export Article

Open AccessCommunication
Int. J. Mol. Sci. 2016, 17(10), 1721; doi:10.3390/ijms17101721

Prokaryotic Expression, Purification and Immunogenicity in Rabbits of the Small Antigen of Hepatitis Delta Virus

1
Engelhardt Institute of Molecular Biology, Russian Academy of Sciences, Vavilov str. 32, Moscow 119991, Russia
2
Gamaleya Research Center of Epidemiology and Microbiology, Gamaleja str. 16, Moscow 123098, Russia
3
A Kirchenstein Institute of Microbiology and Virology, Research Department, Riga Stradins University, Dzirciema iela 16, Riga LV-1007, Latvia
*
Authors to whom correspondence should be addressed.
Academic Editor: Tatsuo Kanda
Received: 25 August 2016 / Revised: 20 September 2016 / Accepted: 27 September 2016 / Published: 20 October 2016
(This article belongs to the Special Issue Hepatitis Virus Infection and Research)
View Full-Text   |   Download PDF [1519 KB, uploaded 20 October 2016]   |  

Abstract

Hepatitis delta virus (HDV) is a viroid-like blood-borne human pathogen that accompanies hepatitis B virus infection in 5% patients. HDV has been studied for four decades; however, the knowledge on its life-cycle and pathogenesis is still sparse. The studies are hampered by the absence of the commercially-available HDV-specific antibodies. Here, we describe a set of reproducible methods for the expression in E. coli of His-tagged small antigen of HDV (S-HDAg), its purification, and production of polyclonal anti-S-HDAg antibodies in rabbits. S-HDAg was cloned into a commercial vector guiding expression of the recombinant proteins with the C-terminal His-tag. We optimized S-HDAg protein purification procedure circumventing a low affinity of the His-tagged S-HDAg to the Ni-nitrilotriacetyl agarose (Ni-NTA-agarose) resin. Optimization allowed us to obtain S-HDAg with >90% purity. S-HDAg was used to immunize Shinchilla grey rabbits which received 80 μg of S-HDAg in two subcutaneous primes in the complete, followed by four 40 μg boosts in incomplete Freunds adjuvant. Rabbits were bled two weeks post each boost. Antibody titers determined by indirect ELISA exceeded 107. Anti-S-HDAg antibodies detected the antigen on Western blots in the amounts of up-to 100 pg. They were also successfully used to characterize the expression of S-HDAg in the eukaryotic cells by immunofluorescent staining/confocal microscopy. View Full-Text
Keywords: hepatitis delta virus; prokaryotic expression; protein purification; rabbit immunization hepatitis delta virus; prokaryotic expression; protein purification; rabbit immunization
Figures

This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. (CC BY 4.0).

Supplementary material

Scifeed alert for new publications

Never miss any articles matching your research from any publisher
  • Get alerts for new papers matching your research
  • Find out the new papers from selected authors
  • Updated daily for 49'000+ journals and 6000+ publishers
  • Define your Scifeed now

SciFeed Share & Cite This Article

MDPI and ACS Style

Tunitskaya, V.L.; Eliseeva, O.V.; Valuev-Elliston, V.T.; Tyurina, D.A.; Zakirova, N.F.; Khomich, O.A.; Kalis, M.; Latyshev, O.E.; Starodubova, E.S.; Ivanova, O.N.; Kochetkov, S.N.; Isaguliants, M.G.; Ivanov, A.V. Prokaryotic Expression, Purification and Immunogenicity in Rabbits of the Small Antigen of Hepatitis Delta Virus. Int. J. Mol. Sci. 2016, 17, 1721.

Show more citation formats Show less citations formats

Note that from the first issue of 2016, MDPI journals use article numbers instead of page numbers. See further details here.

Related Articles

Article Metrics

Article Access Statistics

1

Comments

[Return to top]
Int. J. Mol. Sci. EISSN 1422-0067 Published by MDPI AG, Basel, Switzerland RSS E-Mail Table of Contents Alert
Back to Top