P005 : LOW LEVEL ANTI-HLA ANTIBODIES: DETERMINING RELEVANT UNACCEPTABLE ANTIGENS BY SINGLE ANTIGEN MICROARRAY AND SPECIFIC/SURROGATE FLOW CROSSMATCHES

Abstract

A challenge to any HLA laboratory is to determine the cutoff or level of anti-HLA antibody that ultimately determines the unacceptable antigens (UA) of a potential transplant patient. These UA’s are placed In UNET for virtual crossmatching. Once accomplished, this level is accepted for all HLA antigens as demonstrated by the single antigen bead arrays. The microarray assay is not quantitative and may give false positive reactions at low levels. The challenge is to determine if these antibodies would cause a positive flow T and B cell Flow crossmatch (FLXM). The object of this study is to verify that a low or intermediate Mean Fluorescence Intensity (MFI) anti HLA antibody will or will not cause a positive FLXM with extra deceased donor or normal individual’s lymphocytes. We performed 159 FLXM on 31 patient sera. Only one antibody specificity that could cause a positive FLXM was demonstrated on each of these cells by molecular typing. Our MFI cutoff was 2000 MFI (+DTT). The pronase FLXM Molecules of Equivalent Soluble Fluorochrome (MESF) cutoff was 1500 (+500) for T cell and 2500 (+500) for B cell. Twenty-three patients had negative crossmatches when cross matched by at least five different cells and their serum. The false positive UA as demonstrated by a negative FLXM were removed from UNET. The MFI values for the UA’s varied from 1000 to 5000. There was no correlation of MFI levels to outcome of FLXM. One patient received a renal transplant and two were removed from the wait list. In conclusion: the use of FLXM to demonstrate relevant levels of anti-HLA antibody for each individual patient and allowed us to remove false positive UA’s. This type of analysis gives a potential kidney recipient a greater chance to receive a renal transplant.