OR3 The strength of positive linear correlations between donor-specific HLA antibodies and flow-cytometry crossmatch is locus-dependent

Abstract

To determine correlation between the mean fluorescence intensity (MFI) of donor specific antibodies (DSA) and the median channel shift (MCS) of the flow crossmatch (FXM). HLA antibodies (Ab) in DTT-treated sera were analyzed using One Lambda (OL) single antigen beads (SAB). T cell FXM (51 pronase & 120 untreated) with single class I DSA and B cell FXM (41 pronase & 101 untreated) with single class II DSA were analyzed. MCS values were plotted against the MFI of the DSA and Pearson’s correlations (R2 were assessed. Positive correlations between T-FXM and DSA to HLA-A (R2 = 0.86) or HLA-B (R2 = 0.83) were strong, but low for Cw DSA (R2 = 0.28) [Figure]. B-FXM and DSA to DR (R2 = 0.54) and DQ (R2 = 0.55) had medium degree of +ve correlations while DP (R2 = 0.48) correlation was low. Pronase enhanced MCS but did not improve XM-DSA correlations. DSA with MFI < 2K to a single HLA-A/B or to a single DR/DQ did not cause +ve T or B FXM respectively. For Cw, +ve T-FXM was not observed when DSA was <8K MFI. Similarly, +ve B-FXM was not observed for DP when DSA MFI was <10K. XM reactivity of Cw Ab can be serum dependent. Pronase T-FXM between 3 Cw4+ donors and 2 sera with comparable Ab profiles (Ab to KIR2DL1-binding C2 epitope) on both OL and Immucor SAB panels produced conflicting results. Serum 1 (Cw4 MFI = 8081) was +ve with all donors (MCS = 78, 109, 155) while serum 2 (Cw4 MFI=9610) yielded all T-FXM -ve (MCS ⩽ 10). Sera can be false +ve with OL Cw beads, i.e., −ve with Immucor SAB and −ve T-FXM. Some displayed 10–20K MFI reactivity to certain Cw beads (Cw1, 12, 15) and others reacted to all Cw beads except to self. Continuing correlation studies between DSA and XM are critical for the precise virtual XM prediction. Standalone Cw Ab require additional scrutiny including screening by another reagent and T-FXM. Compared to HLA-A/B/DR/DQ DSAs, 4 or 5 times more Ab are required to yield +ve XM by Cw and DP DSAs respectively, and thus elevating MFI cutoff for Cw and DP unacceptable antigen listing will draw an enhanced donor offer for broadly sensitized patients.Download : Download high-res image (262KB)Download : Download full-size image