Early detection and identification of SARS-CoV-infected patients and actions to prevent transmission are absolutely critical to prevent another SARS outbreak. Antibodies that specifically recognize the SARS-CoV spike and nucleocapsid proteins may provide a rapid screening method to allow accurate identification and isolation of patients with the virus early in their infection. For this reason, we raised peptide-induced polyclonal antibodies against SARS-CoV spike protein and polyclonal antibodies against SARS-CoV nucleocapsid protein using 6x His nucleocapsid recombinant protein. Western blot analysis and immunofluorescent staining showed that these antibodies specifically recognized SARS-CoV.

With the second RT-PCR assay more sensitive than the first RT-PCR assay, we are able to show that approximately 11.5% of well-protected HCWs exposed to SARS patients or specimens may have colonization without seroconversion. Only those with significant clinical symptoms or disease would have active immunity. Thus, regular NPS screening for nested RT-PCR assays in conjunction with a daily recording of body temperature in all first-line HCWs may provide an effective way of early detection.

PCR-restriction fragment length polymorphism (RFLP) analysis is a nonprobe method for the rapid identification of Mycobacterium species. We demonstrate the separation of DNA or restriction fragments digested from the mycobacterial gene encoding the 65-kDa heat shock protein (hsp65) by capillary electrophoresis (CE). By using a pair of unlabeled primers, Tb11 and Tb12, and only one restriction enzyme, HaeIII, we investigated a total of 52 reference and clinical strains encompassing 12 Mycobacterium species. The electrophoretic separation of high-resolution CE required <20 min and was capable of identifying fragments as small as 12 bp. A good agreement of measurement was observed between the sizes of restriction fragments resolved by CE, and the real sizes were deduced from the sequence analysis. Distinct differentiations were also well demonstrated between some species and subspecies by an extra HaeIII digestion site. With the advantage of the complete RFLP pattern available from CE, it appears to be more convenient to use an electropherogram rather than performing the cumbersome slab gel electrophoresis plus diagnostic algorithm to identify Mycobacterium species. Beyond the agarose and polyacrylamide gel electrophoresis, high-resolution CE provides an alternative for rapid identification of Mycobacterium species that is feasible for automation and routine use without the need for costly probes.The traditional identification method for tuberculosis used to take several weeks because of the slow growth rate of mycobacteria. In addition, antibiotic treatment varies according to the species of mycobacteria. Therefore, it is particularly important to diagnose Mycobacterium species as rapidly as possible. For many years, the rapid identification of Mycobacterium species, including both probe and nonprobe methods, had been available for clinical use. The probe method depends upon PCR and hybridization with labeled probes. The nonprobe method requires PCR of the 65-kDa heat shock protein (hsp65) gene and electrophoretic separation of the digested products to obtain the restriction pattern for each species. However, there are reasons why the rapid method cannot be widely applied. For the probe method such as the GenProbe AccuProbes test (11), its identifiable numbers of species are limited and the cost of probes remains high. For the nonprobe method (2, 15), the processes of slab gel electrophoresis are cumbersome and unable to separate the low-molecular-weight fragments. If capillary electrophoresis (CE) could solve the current problems of the nonprobe method, it could be used more commonly with the advantage of additional identifiable species, feasibility for automation, and lower cost compared to the method of Hernandez et al. (6). To achieve restriction fragment length polymorphism (RFLP) detection by automatic fluorescent fragment analysis, Hernandez et al. combined PCR-RFLP analysis of the hsp65 and 16S rRNA genes by using two labeled primer sets and four restriction enzymes. Although the unique patterns were obtai...