Validating laboratory results in a national observational cohort study without field centers: the Reasons for Geographic and Racial Differences in Stroke cohort.

OBJECTIVES: The REasons for Geographic and Racial Differences in Stroke (REGARDS) study is a prospective cohort of 30,239 Americans in the contiguous United States; the first of this scale to use home visits to obtain, process, and ship biologic samples to a core laboratory. Pre-analytical factors resulting from this study design may affect the results of some laboratory assays. We investigated the impact of REGARDS processing on a variety of analytes. DESIGN AND METHODS: In REGARDS, blood samples were processed in the field by technicians who were trained on standardized methods for phlebotomy and sample processing. Field processing included centrifugation using varying non-uniform equipment and shipping overnight on ice to the University of Vermont, where samples were re-centrifuged for 30,000 ×g-minutes and stored at -80 °C. We assessed the effects of REGARDS sample handling by processing split samples from 20 volunteers using either ideal procedures or simulated REGARDS procedures. Assays for 19 analytes for potential study in REGARDS were then run on both samples and results compared. RESULTS: Spearman correlation coefficients for analytes measured in ideal versus REGARDS processed samples ranged from 0.11 to 1.0. Thirteen of 19 analytes were highly correlated (>0.75), but platelet proteins were more variable. CONCLUSIONS: Simulation of non-optimal field processing and shipment to a central laboratory showed high variability in analytes released by platelets. The majority of other analytes produced valid results, but platelet contamination in REGARDS samples makes measurement of platelet proteins unadvisable in these samples. Future analytes considered by REGARDS or similar studies should undergo similar pilot testing.

Detection of human anti-flavivirus antibodies with a west nile virus recombinant antigen microsphere immunoassay.

We report a new, suspended-microsphere diagnostic test to detect antibodies to West Nile (WN) virus in human serum and cerebrospinal fluid (CSF). The microsphere immunofluorescence assay can be performed in less than 3 h on specimens of

Immunoassay targeting nonstructural protein 5 to differentiate West Nile virus infection from dengue and St. Louis encephalitis virus infections and from flavivirus vaccination.

West Nile virus (WNV) is an emerging flavivirus that has caused frequent epidemics since 1996. Besides natural transmission by mosquitoes, WNV can also be transmitted through blood transfusion and organ transplantation, thus heightening the urgency of development of a specific and rapid serologic assay of WNV infection. The current immunoassays lack specificity because they are based on detection of antibodies against WNV structural proteins and immune responses to structural proteins among flaviviruses cross-react to each other. Here, we describe microsphere immunoassays that detect antibodies to nonstructural proteins 3 and 5 (NS3 and NS5). In contrast to immunoassays based on viral envelope and NS3 proteins, the NS5-based assay (i) reliably discriminates between WNV infections and dengue virus or St. Louis encephalitis virus infections, (ii) differentiates between flavivirus vaccination and natural WNV infection, and (iii) indicates recent infections. These unique features of the NS5-based immunoassay will be very useful for both clinical and veterinary diagnosis of WNV infection.