技术资料

Efficient allocation of deceased donor organs depends upon effective prediction of immunologic compatibility based on donor HLA genotype and recipient alloantibody profile,referred to as virtual crossmatching (VCXM). VCXM has demonstrated utility in predicting compatibility,though there is reduced efficacy for patients highly sensitized against allogeneic HLA antigens. The recently revised deceased donor kidney allocation system (KAS) has increased transplantation for this group,but with an increased burden for histocompatibility testing and organ sharing. Given the limitations of VCXM,we hypothesized that increased organ offers for highly-sensitized patients could result in a concomitant increase in offers rejected due to unexpectedly positive crossmatch. Review of 645 crossmatches performed for deceased donor kidney transplantation at our center did not reveal a significant increase in positive crossmatches following KAS implementation. Positive crossmatches not predicted by VCXM were concentrated among highly-sensitized patients. Root cause analysis of VCXM failures identified technical limitations of anti-HLA antibody testing as the most significant contributor to VCXM error. Contributions of technical limitations including additive/synergistic antibody effects,prozone phenomenon,and antigens not represented in standard testing panels,were evaluated by retrospective testing. These data provide insight into the limitations of VCXM,particularly those affecting allocation of kidneys to highly-sensitized patients. View Publication

Recognition of tumor-associated Ags (TAAs) on tumor cells by CTLs and the subsequent tumor cell death are assumed to be dependent on TAA protein expression and to correlate directly with the level of peptide displayed in the binding site of the HLA class I molecule. In this study we evaluated whether the levels of Her-2/neu protein expression on human tumor cell lines directly correlate with HLA-A*0201/Her2/neu peptide presentation and CTL recognition. We developed a TCR mimic (TCRm) mAb designated 1B8 that specifically recognizes the HLA-A2.1/Her2/neu peptide (369-377) (Her2(369)-A2) complex. TCRm mAb staining intensity varied for the five human tumor cell lines analyzed,suggesting quantitative differences in levels of the Her2(369)-A2 complex on these cells. Analysis of tumor cell lines pretreated with IFN-gamma and TNF-alpha for Her2/neu protein and HLA-A2 molecule expression did not reveal a direct correlation between the levels of Her2/neu Ag,HLA-A2 molecule,and Her2(369)-A2 complex expression. However,compared with untreated cells,cytokine-treated cell lines showed an increase in Her2(369)-A2 epitope density that directly correlated with enhanced tumor cell death (p = 0.05). Although a trend was observed between tumor cell lysis and the level of the Her2(369)-A2 complex for untreated cells,the association was not significant. These findings suggest that tumor cell susceptibility to CTL-mediated lysis may be predicted based on the level of specific peptide-MHC class I expression rather than on the total level of TAA expression. Further,these studies demonstrate the potential of the TCRm mAb for validation of endogenous HLA-peptide epitopes on tumor cells. View Publication