The emergence of severe acute respiratory syndrome (SARS) resulted in several outbreaks worldwide. Early tests for diagnosis were not always conclusive in identifying a SARS suspected patient. Nucleocapsid protein (NP) is the most predominant virus derived structural protein which is shed in high amounts in serum and nasopharyngeal aspirate during the first week of infection. As part of such efforts, a simple, easy to use immunoswab method was developed by generating a panel of monoclonal antibodies (MAbs), Bispecific MAbs and chicken polyclonal IgY antibody against the SARS-CoV nucleocapsid protein (NP). Employing the MAb-based immunoswab, an NP concentration of 200 pg/mL in saline and pig nasopharyngeal aspirate, and 500 pg/mL in rabbit serum were detected. BsMAb-based immunoswabs detected an NP concentration of 20 pg/mL in saline, 500 pg/mL in rabbit serum and 20-200 pg/mL in pig nasopharyngeal aspirate. Polyclonal IgY-based immunoswabs detected an NP concentration of 10 pg/mL in pig nasopharyngeal aspirate providing the most sensitive SARS point of care assay. Results show that the robust immunoswab method of detecting SARS-CoV NP antigen can be developed into an easy and effective way of identifying SARS suspected individuals during a future SARS epidemic, thereby reducing and containing the transmission. The key feature of this simple immunoswab diagnostic assay is its ability to detect the presence of the SARS-CoV antigen within 45-60 min with the availability of the body fluid samples.

The goal of this study was to develop a quantitative detection system for severe acute respiratory syndrome-associated coronavirus (SARS-CoV), targeting the nucleocapsid protein (NP), to determine the presence and degree of infection in suspected individuals. Because the NP is the viral protein shed during infection and its template mRNA is the most abundant subgenomic RNA, it is a suitable candidate for developing antibodies for diagnostic applications. In this study, we have prepared full-length SARS-CoV NP expressed in Escherichia coli and purified. Full-length NP was used for the preparation of mouse monoclonal antibody and chicken polyclonal IgY antibodies for the development of heterosandwich ELISA for early diagnostics of SARS-suspected individuals. The sensitivity of the developed heterosandwich ELISA can detect the viral antigen at 18.5 pg/mL of recombinant NP. This study describes ultrasensitive ELISA using 19B6 monoclonal antibody as the capture antibody and IgY as the detecting antibody against the most abundant SARS-CoV NP antigens. One of the most important findings was the use of inexpensive polyclonal IgY antibody to increase the sensitivity of the detection system for SARS-CoV at the picogram level. Furthermore, the immunoassay of SARS-CoV NP antigen developed could be an effective and sensitive method of diagnosing SARS-suspected individuals during a future SARS-CoV outbreak.

Five monoclonal antibodies (MAbs) against recombinant nucleocapsid protein (NP) of severe acute respiratory syndrome (SARS)-causing coronavirus (CoV) were developed by hybridoma technology. Epitope mapping by Western blotting showed that these anti-SARS-CoV NP MAbs bind to distinct domains of NP. These anti-SARS-CoV NP MAbs, with their high specificity, are potentially ideal candidates for developing early and sensitive diagnostic assays for SARS-CoV.

An in situ dual vascular and luminal perfusion technique was used to study the effect of cholecystokinin octapeptide (CCK-8) on the transport of hexoses by the jejunum of the Sprague-Dawley rat from the lumen to the vascular bed. The lumen of the jejunum was perfused with hexoses in oxygenated Krebs buffer, while the superior mesenteric artery was infused with Krebs buffer containing Ficoll 70 as a plasma expander. CCK-8 (0.8-8 pM) in the vascular infusate selectively reduced hexose transport in a dose-dependent manner by 20-47%, although having no effect on L-glucose or L-leucine absorption. Vascular tetrodotoxin did not block CCK-8 inhibition, whereas a specific CCK-A receptor antagonist, lorglumide, did. The CCK-B receptor agonist cholecystokinin tetrapeptide had a small effect on hexose absorption, whereas somatostatin-14 and -28 had no effect. These results suggest that cholecystokinin can decrease intestinal absorption of hexoses in the small intestine, acting via CCK-A-type receptors.