Differential sensitivities of severe acute respiratory syndrome (SARS) coronavirus spike polypeptide enzyme-linked immunosorbent assay (ELISA) and SARS coronavirus nucleocapsid protein ELISA for serodiagnosis of SARS coronavirus pneumonia.
The use of recombinant severe acute respiratory syndrome-coronavirus (SARS-CoV) nucleocapsid protein (N) enzyme-linked immunosorbent assay (ELISA)-based antibody and antigen tests for diagnosis of SARS-CoV infections have been widely reported. However, no recombinant SARS-CoV spike protein (S)-based ELISA is currently available. In this article, we describe the problems and solutions of setting up the recombinant SARS-CoV S-based ELISA for antibody detection. The SARS-CoV S-based immunoglobulin M (IgM) and IgG ELISAs were evaluated and compared with the corresponding N-based ELISA for serodiagnosis of SARS-CoV pneumonia, using sera from 148 healthy blood donors who donated blood 3 years ago as controls and 95 SARS-CoV pneumonia patients in Hong Kong. Results obtained by the recombinant S (rS)-based IgG ELISA using the regenerated S prepared by dialysis with decreasing concentrations of urea or direct addition of different coating buffers, followed by addition of different regeneration buffer, identified 4 M urea and 1 M sarcosine for plate coating and no regeneration buffer as the most optimal conditions for antibody detection. The specificities of the S-based ELISA for IgG and IgM detection were 98.6% and 93.9%, with corresponding sensitivities of 58.9% and 74.7%, respectively. The sensitivity of the rN IgG ELISA (94.7%) is significantly higher than that of the rS IgG ELISA (P < 0.001), whereas the sensitivity of the rS IgM ELISA is significantly higher than that of the rN IgM ELISA (55.2%) (P < 0.01). An ELISA for detection of IgM against S and N could be more sensitive than one that detects IgM against N alone for serodiagnosis of SARS-CoV pneumonia.
Department of Microbiology, The University of Hong Kong, University Pathology Building, Queen Mary Hospital, Pokfulam, Hong Kong.
Patrick C Y
PC
Woo
Susanna K P
SK
Lau
Beatrice H L
BH
Wong
Hoi-Wah
HW
Tsoi
Ami M Y
AM
Fung
Richard Y T
RY
Kao
Kwok-Hung
KH
Chan
J S Malik
JS
Peiris
Kwok-Yung
KY
Yuen
D000914:Antibodies, Viral / Q000097:blood*; D000086462:Coronavirus Nucleocapsid Proteins; D004797:Enzyme-Linked Immunosorbent Assay; D006801:Humans; D008562:Membrane Glycoproteins / Q000235:genetics / Q000276:immunology*; D019590:Nucleocapsid Proteins / Q000235:genetics / Q000276:immunology*; D010455:Peptides / Q000276:immunology; D011994:Recombinant Proteins / Q000276:immunology; D045473:Severe acute respiratory syndrome-related coronavirus / Q000276:immunology; D012680:Sensitivity and Specificity; D012698:Serologic Tests; D045169:Severe Acute Respiratory Syndrome / Q000175:diagnosis* / Q000821:virology; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins / Q000235:genetics / Q000276:immunology*
D000914:Antibodies, Viral; D000086462:Coronavirus Nucleocapsid Proteins; D008562:Membrane Glycoproteins; D019590:Nucleocapsid Proteins; D010455:Peptides; D011994:Recombinant Proteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV