Cloning, Sequencing, Expression, and Purification of SARS-associated Coronavirus Nucleocapsid Protein for Serodiagnosis of SARS

J Clin Virol. 2004 Aug;30(4):309-12. doi: 10.1016/j.jcv.2004.01.001.

Abstract

A novel coronavirus has been associated with a worldwide outbreak of atypical pneumonia referred to as Severe Acute Respiratory Syndrome (SARS-CoV). SARS-CoV nucleocapsid (N) protein has been cloned sequenced and expressed in Escherichia coli strain. Purified N protein was used to measure the SARS-CoV specific IgG antibodies from 16 SARS-CoV infected patients' sera and from 131 control subjects using ELISA assay. Specific antibody responses to the purified recombinant N protein after 10, 20, and 30 days of disease onset were observed in 13 of 16 (81.3%), 16 of 16 (100%) and 16 of 16 (100%) SARS patients sera, respectively. Comparison of detection results with a commercially available diagnostic kit coated with a mixture of SARS-CoV viral proteins showed 9 of 16 (56.3%), 13 of 16 (81.3%), and 15 of 16 (93.7%) positive responses, respectively. None of 131 control sera gave positive reaction in either assay. This data suggests that the N protein of SARS-CoV is immunodominant and this ELISA based test assay for detecting the SARS-CoV N antigen may hold a significant value for SARS diagnosis.

Publication types

  • Evaluation Study

MeSH terms

  • Antibodies, Viral / blood*
  • Cloning, Molecular
  • Enzyme-Linked Immunosorbent Assay
  • Escherichia coli / genetics
  • Escherichia coli / metabolism
  • Humans
  • Molecular Sequence Data
  • Nucleocapsid Proteins* / chemistry
  • Nucleocapsid Proteins* / genetics
  • Nucleocapsid Proteins* / isolation & purification
  • Nucleocapsid Proteins* / metabolism
  • Recombinant Proteins / genetics
  • Recombinant Proteins / immunology*
  • Recombinant Proteins / metabolism
  • SARS Virus / genetics
  • SARS Virus / immunology*
  • SARS Virus / metabolism
  • Sequence Analysis, DNA
  • Serologic Tests
  • Severe Acute Respiratory Syndrome / diagnosis*
  • Severe Acute Respiratory Syndrome / virology

Substances

  • Antibodies, Viral
  • Nucleocapsid Proteins
  • Recombinant Proteins
  • nucleocapsid protein, Coronavirus

Associated data

  • GENBANK/AY365036