P021 : DTT CAN UNCOVER HIDDEN UAS

Abstract

Aim Luminex Single Antigen (LSA) technology for antibody identification has enabled laboratories to provide highly accurate virtual crossmatch results to surgeons prior to renal transplantation. In our laboratory, unacceptable antigens (UAs) are assigned for antibodies with 1500 MFI or higher. A deceased donor offer was extended to a patient on our waitlist, and the virtual crossmatch was negative. Surprisingly, the actual flow crossmatch was strongly positive for both T cells and B cells. The organ was declined for this patient, and the discrepancy was investigated. Methods The patient was a 53 y.o. female with a prior transplant, multiple pregnancies and transfusions, and 100% cPRA. The strongest antibody this patient had to the donor (A1) had an MFI of only 1167 by LSA beads (One Lambda Inc), which was not considered a UA. In our experience, this should not have caused a positive flow crossmatch. The crossmatch was run with and without Dithiothreitol (DTT) treatment (0.05 M at 37 °C for 30 min), and the results were a Δ MESF of +123,108 (− DTT) and +86,366 (+ DTT) for T cells and +179,841 (− DTT) and +75,565 (+ DTT) for B cells. The sample had never been tested with DTT treatment on LSA beads, so the sample was run with and without DTT on LSA beads in an attempt to explain the positive crossmatch. Results The A1 bead, which had an MFI of only 1167 on LSA beads without DTT, jumped to an MFI of 17,300 with DTT for the same sample. This high MFI correlated with the strongly positive flow crossmatch, and A1 was entered as a UA for this patient. Conclusions Our laboratory had been using DTT routinely for flow crossmatches but not for antibody testing. This patient had a strong antibody to A1 that went undetected by LSA without the use of DTT. Since this discovery, our lab has begun using DTT for all LSA testing in order to more accurately predict crossmatch results.