Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
권호 표지
DOI QR코드

DOI QR Code

Optimized Expression, Purification, and Rapid Detection of Recombinant Influenza Nucleoproteins Expressed in Sf9 Insect Cells

  • Yoon, Sung-Jin (Metabolic Regulation Research Center, Korea Research Institute of Bioscience and Biotechnology) ;
  • Park, Young-Jun (Metabolic Regulation Research Center, Korea Research Institute of Bioscience and Biotechnology) ;
  • Kim, Hyun Ju (Department of Systems Biotechnology, Chung-Ang University) ;
  • Jang, Jinwoo (R&D Center, Sugentech Inc.) ;
  • Lee, Sang Jun (Department of Systems Biotechnology, Chung-Ang University) ;
  • Koo, Sunwoo (Department of Neuroscience and Experimental Therapeutics, College of Medicine, Texas A&M University) ;
  • Lee, Moo-Seung (Department of Biomolecular Science, KRIBB School of Bioscience, Korea University of Science and Technology (UST))
  • Received : 2018.05.23
  • Accepted : 2018.08.23
  • Published : 2018.10.28
Download PDF
⟨ Previous Next ⟩

Abstract

Accurate and rapid diagnosis of influenza infection is essential to enable early antiviral treatment and reduce the mortality associated with seasonal and epidemic infections. Immunochromatography is one of the most common methods used for the diagnosis of seasonal human influenza; however, it is less effective in diagnosing pandemic influenza virus. Currently, rapid diagnostic kits for pandemic influenza virus rely on the detection of nucleoprotein (NP) or hemagglutinin (HA). NP detection shows higher specificity and is more sensitive than HA detection. In this study, we time-dependently screened expression conditions, and herein report optimal conditions for the expression of recombinant nucleoprotein (rNP), which was 48 h after infection. In addition, we report the use of the expressed rNP in a rapid influenza diagnostic test (SGT i-flex Influenza A&B Test). We constructed expression vectors that synthesized rNP (antigen) of influenza A and B in insect cells (Sf9 cells), employed the purified rNP to the immunoassay test kit, and clearly distinguished NPs of influenza A and influenza B using this rapid influenza diagnostic kit. This approach may improve the development of rapid test kits for influenza using NP.

Keywords

References

  1. Wang F, Qi J, Bi Y, Zhang W, Wang M, Zhang B, et al. 2015. Adaptation of avian influenza A (H6N1) virus from avian to human receptor-binding preference. EMBO J. 34: 1661-1673. https://doi.org/10.15252/embj.201590960
  2. Vemula SV, Zhao J, Liu J, Wang X, Biswas S, Hewlett I. 2016. Current approaches for diagnosis of influenza virus infections in humans. Viruses 8: 96. https://doi.org/10.3390/v8040096
  3. Pedersen JC. 2014. Hemagglutination-inhibition assay for influenza virus subtype identification and the detection and quantitation of serum antibodies to influenza virus. Methods Mol. Biol. 1161: 11-25.
  4. Nitsch-Osuch A, Wozniak-Kosek A, Brydak LB. 2012. Accuracy of rapid influenza diagnostic test and immunofluorescence assay compared to real time RT-PCR in children with influenza A(H1N1)pdm09 infection. Postepy Hig. Med. Dosw. (Online) 66: 752-757. https://doi.org/10.5604/17322693.1015040
  5. Wang W, Huang B, Jiang T, Wang X, Qi X, Tan W, et al. 2014. Maximal immune response and cross protection by influenza virus nucleoprotein derived from E. coli using an optimized formulation. Virology 468-470: 265-273. https://doi.org/10.1016/j.virol.2014.08.008
  6. Mizuike R, Sasaki T, Baba K, Iwamoto H, Shibai Y, Kosaka M, et al. 2011. Development of two types of rapid diagnostic test kits to detect the hemagglutinin or nucleoprotein of the swine-origin pandemic influenza A virus H1N1. Clin. Vaccine Immunol. 18: 494-499. https://doi.org/10.1128/CVI.00269-10
  7. Rota PA, Black RA, De BK, Harmon MW, Kendal AP. 1990. Expression of influenza A and B virus nucleoprotein antigens in baculovirus. J. Gen. Virol. 71(Pt 7): 1545-1554. https://doi.org/10.1099/0022-1317-71-7-1545
  8. Miyoshi-Akiyama T, Narahara K, Mori S, Kitajima H, Kase T, Morikawa S, et al. 2010. Development of an immunochromatographic assay specifically detecting pandemic H1N1 (2009) influenza virus. J. Clin. Microbiol. 48: 703-708. https://doi.org/10.1128/JCM.02262-09
  9. Choi YJ, Kim HJ, Park JS, Oh MH, Nam HS, Kim YB, et al. 2010. Evaluation of new rapid antigen test for detection of pandemic influenza A/H1N1 2009 virus. J. Clin. Microbiol. 48: 2260-2262. https://doi.org/10.1128/JCM.02392-09
  10. Kost TA, Condreay JP. 1999. Recombinant baculoviruses as expression vectors for insect and mammalian cells. Curr. Opin. Biotechnol. 10: 428-433. https://doi.org/10.1016/S0958-1669(99)00005-1
  11. Kidd IM, Emery VC. 1993. The use of baculoviruses as expression vectors. Appl. Biochem. Biotechnol. 42: 137-159. https://doi.org/10.1007/BF02788049
  12. Hoss A, Moarefi I, Scheidtmann KH, Cisek LJ, Corden JL, Dornreiter I, et al. 1990. Altered phosphorylation pattern of simian virus 40 T antigen expressed in insect cells by using a baculovirus vector. J. Virol. 64: 4799-4807.
  13. Bose ME, Sasman A, Mei H, McCaul KC, Kramp WJ, Chen LM, et al. 2014. Analytical reactivity of 13 commercially available rapid influenza diagnostic tests with H3N2v and recently circulating influenza viruses. Influenza Other Respir. Viruses 8: 474-481. https://doi.org/10.1111/irv.12246
  14. Ferguson BS, Buchsbaum SF, Wu TT, Hsieh K, Xiao Y, Sun R, et al. 2011. Genetic analysis of H1N1 influenza virus from throat swab samples in a microfluidic system for point-of-care diagnostics. J. Am. Chem. Soc. 133: 9129-9135. https://doi.org/10.1021/ja203981w
  15. Levine M, Sheu TG, Gubareva LV, Mishin VP. 2011. Detection of hemagglutinin variants of the pandemic influenza A (H1N1) 2009 virus by pyrosequencing. J. Clin. Microbiol. 49: 1307-1312. https://doi.org/10.1128/JCM.02424-10