DETECTION OF CRYPTOSPORIDIUM ANTIBODIES IN SERA AND ORAL FLUIDS USING MULTIPLEX BEAD ASSAY

For the first time, a multiplex bead assay (MBA) was used to assay oral fluid and serum specimens for immunoglobulin G (IgG) antibodies to specific Cryptosporidium parvum antigens that were coupled to polystyrene beads. Recombinant C. parvum 17- and 27-kDa antigens (r17 and r27, respectively) both linked with glutathione-S-transferase (GST) fusion proteins, native 17-kDa antigen, and GST alone were each coupled to microspheres that could be differentiated based on variable amounts of internally incorporated red fluorescent dye. Initial and follow-up serum and oral fluid specimens from a 1997 cryptosporidiosis outbreak in Spokane, Washington, were incubated with the coupled beads. Antibodies bound to the coupled beads were detected using biotinylated monoclonal anti-human IgG antibody and streptavidin-labeled r-phycoerythrin. Fluorescence intensity was measured by flow cytometry. For the 3 C. parvum antigens, the median of the mean fluorescence intensity (MFI) was significantly higher (P < 0.03) in the ini...