1014-LBP: Positive flow cytometry crossmatch results with pronase-treated t-cells induced by non-HLA autoantibodies in HIV-infected patients

Aim Binding of IgG to FcR on B-cells decreases the sensitivity & specificity of the B-cell FCXM. To prevent this, pronase treatment (PT) of un-separated T- & B-cells is widely used. However, the effect of PT on the T-cell FCXM is not well-documented. We have observed a high rate of positive FCXM with PT T-cells in the absence of DSA in HIV+ patients. Methods 26 HIV+ and 30 HIV- 0% PRA patients were tested. 348 sera from HIV+ patients were crossmatched with PT cells, and 81 sera were crossmatched with non-PT (NT) cells from 196 deceased donors. 60 sera from HIV- patients were crossmatched with PT & NT cells from 48 deceased donors. Selected HIV+ sera were pre-treated with reducing agents. Selected HIV+ sera were pre-absorbed with PT & NT T-cells or PT & NT B-cells and then crossmatched with PT cells. Results 97% of HIV+ patients exhibited strongly positive FCXM with PT T-cells while 17% exhibited weakly positive FCXM with NT T-cells. Only 9% & 16% of HIV+ patients exhibited weakly positive FCXM with PT & NT B-cells, respectively. Positive FCXM were observed with PT but not with NT CD4+ & CD8+ T-cells. Pre-treatment of HIV+ sera with reducing agents did not have any effect on the FCXM strength. Pre-absorption of HIV+ sera with PT T-cells significantly lowered the T-cell FCXM strength and had no effect on the B-cell FCXM strength. In contrast, pre-absorption of HIV+ sera with PT B-cells or with NT T- & B-cells did not have any effect on the FCXM strength. 12 HIV+ patients have been transplanted with kidney allografts with positive PT T-cell/negative NT T-cell FCXM. 100% of these allografts are functioning today (MST  = 991 ± 571 days, range: 253–1966 days). Conclusions Non-specific IgG binding is not the cause of positive T-cell FCXM in HIV+ patients. Reduction of the T-cell FCXM strength observed on HIV+ sera pre-absorbed with PT T-cells indicates that these positive results are due to autoantibodies recognizing cryptic epitopes exposed by PT on T-cells.