Evaluation of a safe and sensitive Spike protein-based immunofluorescence assay for the detection of antibody responses to SARS-CoV

J Immunol Methods. 2005 Jan;296(1-2):37-44. doi: 10.1016/j.jim.2004.10.012. Epub 2004 Dec 10.

Abstract

Previously, we have identified a truncated antigenic fragment named protein C [441 to 700 amino acids (a.a.)] as the immunodominant fragment of Spike (S) protein of severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV). We have now successfully expressed protein C using the baculovirus system in S. frugiperda (Sf-9) cells. This recombinant baculovirus expressing protein C was first characterized using five SARS convalescent human sera and five normal human sera. The results showed that protein C is an authentic antigen against SARS-CoV antibody. Our Spike protein-based immunoflourescence assay (IFA) based on this recombinant baculovirus-Sf-9 system was further assessed with a panel of 163 clinical samples collected during the SARS epidemic in Singapore, which include samples from 21 clinically confirmed SARS, 42 non-SARS patient sera, and 100 normal sera. The results were compared to a commercial SARS IFA kit (EUROIMMUN, Germany) and a conventional IFA test performed in Singapore General Hospital. All of the 21 SARS-positive serum samples could be recognized by our IFA, giving a specificity and sensitivity of 100%, which was compatible with both whole virus-based IFA assays. No cross-reactivity with serum samples against infectious bronchitis virus (IBV) and transmissible gastroenteritis virus (TGEV) were detected in our assays. Thus, our Spike protein-based IFA could offer a safer procedure which can be performed in a BSL-2 laboratory as it could mimic the whole virus based-IFA without any loss of sensitivity and specificity. It is also more user-friendly and cost-effective than the whole virus-based IFA.

Publication types

  • Comparative Study
  • Evaluation Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antibodies, Viral / blood*
  • Antibodies, Viral / immunology
  • Antigens, Viral / genetics
  • Antigens, Viral / immunology*
  • Baculoviridae / genetics
  • Cross Reactions / immunology
  • Fluorescent Antibody Technique, Indirect*
  • Humans
  • Membrane Glycoproteins / immunology
  • Microscopy, Fluorescence
  • SARS Virus / immunology*
  • Sensitivity and Specificity
  • Severe Acute Respiratory Syndrome / diagnosis*
  • Spike Glycoprotein, Coronavirus
  • Spodoptera / cytology
  • Spodoptera / genetics
  • Viral Envelope Proteins / immunology
  • Viral Proteins / genetics
  • Viral Proteins / immunology*

Substances

  • Antibodies, Viral
  • Antigens, Viral
  • MHV surface projection glycoprotein
  • Membrane Glycoproteins
  • Spike Glycoprotein, Coronavirus
  • Viral Envelope Proteins
  • Viral Proteins
  • protein C, SARS virus
  • spike glycoprotein, SARS-CoV