Enzyme linked immunosorbent assay for detection of anti-red blood cell(RBC) antibodies using RBC ghost protein.

概要

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A new enzyme linked immunosorbent assay (ELISA) was developed for detecting anti-red blood cell (RBC) antibodies. RBC ghost protein was immobilized on the microtiter plates with 0.05M carbonate buffer, pH 9.6. One percent bovine serum albumin (BSA) in phosphate buffered saline (PBS) was used as a blocking solution. Indirect Coombs' test positive sera and antibody eluates from patients' RBC were added to the wells, and then optimally diluted horseradish peroxidase-conjugated goat anti-human IgG was added. Absorbed peroxidase activity was detected by o-phenylenediamine, and color reactions were terminated by addition of 1 N HCl. Absorbance measured at 490nm.The reactivities of indirect Coombs' test positive sera were statistically higher (P<0.01) than that of normal sera (0.295±0.355, 0.024±0.030, respectively), and antibody eluates from patients' RBC also responded. But a few samples did not respond. This method is suitable for handling many samples and more sensitive than Coombs' test. But, because some samples positive in Coombs' test did not respond in this method, it is necessary to investigate the difference of the antigenicity between the RBC and the RBC ghost.
日本臨床免疫学会の論文