Recombinant nucleocapsid protein-based IgG enzyme-linked immunosorbent assay for the serological diagnosis of SARS

Abstract

The recombinant nucleocapsid protein (rNP) of severe acute respiratory syndrome (SARS) coronavirus (SARS-CoV) was expressed in a baculovirus system. The purified SARS-CoV rNP was used as an antigen for detection of SARS-CoV antibodies in IgG enzyme-linked immunosorbent assay (ELISA). The ELISA was evaluated in comparison with neutralizing antibody assay and the authentic SARS-CoV antigen-based IgG ELISA. Two-hundred and seventy-six serum samples were collected from health care workers in a hospital in which a nosocomial SARS outbreak took place and used for evaluation. The SARS-CoV rNP-based IgG ELISA has 92% of sensitivity and specificity compared with the neutralizing antibody assay and 94% sensitivity and specificity compared with the authentic SARS-CoV antigen-based IgG ELISA. The results suggest that the newly developed SARS-CoV rNP-based IgG ELISA is a valuable tool for the diagnosis and seroepidemiological study of SARS. The SARS-CoV rNP-based IgG ELISA has an advantage over the conventional IgG ELISA in that the antigen can be prepared by laboratory workers without the risk of infection.

The SARS-CoV nucleocapsid protein: A protein with multifarious activities

“…Nprotein has been produced in abundant quantity using codon optimised gene in Escherichia coli (Das and Suresh, 2006). Saijo et al (2005) have successfully expressed recombinant Nprotein using a baculovirus expression system, which was found to be 92% efficient in neutralizing antibody assay. In another study, Liu et al (2004), have expressed full length Nprotein using yeast expression system.…”

Evaluation of a novel vesicular stomatitis virus pseudotype-based assay for detection of neutralizing antibody responses to SARS-CoV

“…The conventional neutralization assay using infectious SARS-CoV (isolate HKU39849, kindly supplied by Dr. J.S. Malik Peiris) was performed as described by Saijo et al [2005]. Briefly, the heat-inactivated serum samples were diluted twofold with Eagle's minimum essential medium (MEM) containing 2% FBS from 1:10 to 1:320.…”

“…The development of early, rapid, and reliable diagnostic assay systems is a high priority for early vigilance to prevent the spread of the disease. Identification of SARScoronavirus (SARS-CoV) as the causative agent of SARS has led to the development of genome-based and serological assays for diagnosis of viral infection [Hong et al, 2004;Hourfar et al, 2004;Saijo et al, 2005]. Detection of virus neutralizing antibody (Nab) is important to assess immune responses in SARS-CoVinfected patients.…”

Development and validation of serological assays for viral hemorrhagic fevers and determination of the prevalence of Rift Valley fever in Borno State, Nigeria

“…The conventional neutralization assay using infectious RVFV (RVFV-MP12 strain) was performed as described previously. 20 Briefly, heat-inactivated serum samples were diluted three-fold (from 1:40 to 1:1080) with Eagle's minimum essential medium (MEM, Sigma, St. Louis, MO, USA) containing 2% FBS (Invitrogen, Carlsbad, CA, USA). Each test sample (50 ml) was then mixed with the same volume of RVFV-MP12 at an infectious dose of 100 plaque forming units.…”