A clinically significant anti-HLA-A2 detectable by extended incubation cytotoxicity and flow cytometric techniques but not by a standard NIH lymphocytotoxicity test

A previously transfused female patient, known to have a platelet defect, was transfused with platelets prior to surgery. After the 18th unit she felt unwell, developed fever, rigor, became nauseous, and vomited. Her blood pressure decreased from 140/90 to 80/50mm Hg. Passive transfer of donor granulocytes or red cell antibodies were excluded as a cause. Therefore, a serum sample from the patient was investigated for the presence of antibodies to human leukocyte antigens (HLA) using a standard National Institutes of Health (NIH) lymphocytotoxicity test, but antibodies were not detected. However, an extended incubation cytotoxicity test demonstrated the presence of an anti-HLA-A2, and indirect immunofluorescence flow cytometry showed the presence of an IgG1 antibody reacting with 50 percent of cells in a random pool of lymphocytes. One week later, multispecific HLA antibodies were detectable by both NIH and extended incubation cytotoxicity tests. Flow cytometry showed a 16-fold increase in the amount of IgG antibodies and the appearance of an IgM component. Such clinically important HLA antibodies can be detected by extended incubation cytotoxicity and flow cytometric assays prior to becoming reactive in a standard NIH cytotoxicity technique.