Abstract
Introduction: Progress in assays detecting and characterizing anti-human leukocyte antigen (HLA) antibodies have significantly improved outcomes in thoracic organ transplantation. However, due to the complicated nature of humoral alloresponses practices are still evolving in laboratories to determine the best combination of assays that can be used for graft allocation in prospective heart allograft (HTx) recipients, as well as for post-transplant monitoring of sensitized patients. In the present study we analyzed the efficiency of three different laboratory methodologies for the detection of donor-specific anti-HLA antibodies (DSA), their degree of correlation and their clinical relevance. Methods: The results of a Luminex donor-specific crossmatch (DSA-LX), in which donor-isolated HLA antigens are coated onto capture microbeads (Gen-Probe Transplant Diagnostics, USA), were compared with the results of a flow-cytometric crossmatch (FCM-X) for the same donor-recipient combination and with the results of a Luminex single-antigen bead methodology (SA-LX) in which recombinant HLA class I or class II molecules are coated onto microbeads (One Lambda, USA). A total of 46 sera samples obtained pre- (27 samples) and post-transplant (19 samples) were included in the study. All 46 samples were tested with the DSA-LX and SA-LX methodology, while the 27 pre-transplant samples were also analyzed by the FCM-X methodology. Results: For the pre- transplant sera, the results of the three methods were in agreement for class I DSA in 21/27 (77.8%) samples and for class II DSA in 17/27 (63.0%) samples. When the results of the two bead-based methodologies (DSA-LX and SA-LX) were compared to the results of the FCM-X methodology, the SA-LX results showed better correlation with the flow-cytometric crossmatch results than the DSA-LX results both for class I and for class II antibodies. The results of the two bead-based assays SA-LX and DSA-LX, coincided for 40/46 (87.0%) and for 33/46 (71.7%) of sera tested for class I and class II HLA-DSA respectively. Furthermore, in the group of transplanted patients the SA-LX results had more clinical relevance than the DSA-LX results, particularly when samples were tested positive by SA-LX and negative by DSA-LX. The methodology reproducibility score for both the SA-LX and the FCM-X was 100% whereas for DSA-LX that score was 88.9%. Conclusions: The results of the DSA-LX methodology do not have a good correlation with the results of the more established FCM-X and SA-LX techniques for the detection of DSA, particularly as far as class II antibodies. Furthermore, in comparison to FCM-X and SA-LX, the DSA-LX methodology has a lower reproducibility performance and a lower clinical relevance which renders it unsuitable for reliable graft allocation and patient monitoring.
Published Version
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