Single Antigen Beat Assay (Luminex) Allows Detection of Immunised Patients with an Increased Risk for Long Term Graft Survival Prior to Transplantation Rudolf Richter1, Lisa Holschuh1, Anna-Alicia Schödel1, Stefanie Feuchtinger1,2, Sara Qidan1,2, Martin Brzoska 2, Aida Asbe-Vollkopf 2, Stefan Büttner2, Christoph Betz2, Stefan Gauer2, Erhard Seifried1, Helmut Geiger2, Ingeborg Anna Hauser2, Christian Seidl1 1Institute of Transfusion Medicine, Department of Transplantation Immunology, 2Department of Nephrology, Medical Clinic III, Johann Wolfgang Goethe-University Conclusions Introduction Recently, the Single Antigen Bead (SAB) assay (Luminex), a highly sensitive assay for detection of HLA-class I and HLA-class II specific antibodies (Ab) has been established in routine testing. In addition, a modified assay for identification of complement C1q fixing HLA Ab was developed (C1q-SAB). The single antigen format of the SAB and the C1q-SAB facilitates antibody differentiation with a sensitivity, which exceeds the sensitivity of the ELISA and the complement dependent cytotoxicity (CDC) assay. However, there is ongoing discussion concerning the clinical relevance of the detected Ab specificities. The increased sensitivity leads to detection of an increased number of HLA Ab which increases virtual (v)PRA values, which finally might end up with a prolonged patient´s waiting time for a kidney transplant. In the present study we compared the transplant survival in patients positive for Ab detected with the SAB and the C1q-SAB assay. For definition of HLA Ab (normalized fluorescence intensity [nMFI]) cut off values of ≥1000 and ≥3000 were used. 183 patients were included in this study. In the SAB assay, Kaplan–Meier analyses revealed a significantly reduced death-censored long-term graft survival in patients with exclusive HLA class I Ab, and with both HLA class I and class II Ab. Contrary to the results of the SAB assay, the Kaplan- Meyer analyses of the C1q-SAB results revealed a trend for a reduced death-censored long-term graft survival in patients positive for both HLA class I- and II C1q fixing Ab only. Cutoff values of nMFI ≥ 1000 for definition of HLA Ab allowed best detection of immunised patients with an increased risk for long term graft loss. The graft survival in patients negative for SAB or C1q- SAB detected HLA-class I + II Ab differed significantly (Figure A). In conclusion, our results point to a significantly increased risk for long-term graft survival in patients which are positive for SAB detected HLA class I or both HLA class I and II Ab prior to transplantation. However, in our study the C1q SAB assay did not allow a clear detection of immunised patients with an increased risk for death censored long-term graft survival. Results Impact of SAB and C1q-SAB detected HLA class I and II Ab on transplant survival nMFI cutoff values ≥1000 and ≥3000 HLA class I and II Ab and DSA on transplant survival (1.A) HLA class I – Ab positive patients (A) (B) Transplant survival (%) Ab neg /pos Hazard Ratio (95% Cl) P* N neg/pos ELISA 88/100 3.12 (0.51 – 19.04) 0.21 142/20 Luminex (nMFI cutoff) 1000 100/80 0.07 (0.02 - 0.30) 0.0003 82/53 3000 95/79 0.17 (0.04 - 0.81) 0.025 120/35 (1.B) HLA class I – C1q-SAB positive patients Transplant survival (%) Ab neg /pos Hazard Ratio (95% Cl) P* N neg/pos C1q-SAB (nMFI cutoff) 1000 84 / 100 2,9 (0.32 - 27.17) 0.34 154/11 3000 87 / 100 0,34 (0.06 – 3.36) 0.36 161/9 (C) (D) (1.C) HLA class I + II – Ab positive patients Transplant survival (%) Ab neg /pos Hazard Ratio (95% Cl) P* N neg/pos ELISA 88/64 0.48 (0.01 – 2.39) 0.38 142/14 Luminex (nMFI cutoff) 1000 100/63 0.06 (0.01 – 0.28) 0.0002 82/42 3000 95/68 0.21 (0.05 – 0.96) 0.044 120/26 Legend Figure A to D: Impact of SAB and C1q detected HLA class I and HLA class II Ab. For definition of AB positive patients nMFI cutoff values of 1000 and 3000 were used. Kaplan–Meier graft survival for patients (A) exclusively positive HLA class I Ab (cutoff ≥1000 nMFI) (B) exclusively positive for HLA class I Ab (cutoff ≥3000 nMFI) (C) positive for HLA Ab class I+II Ab (cutoff ≥ 1000 nMFI) (D) positive for HLA Ab class I+II Ab (cutoff ≥ 3000 nMFI) (1.D) HLA class I + II – C1q-SAB positive patients Transplant survival (%) Ab neg /pos Hazard Ratio (95% Cl) P* N neg/pos C1q-SAB (nMFI cutoff) 1000 88 / 54 4,29 (0.69 - 26.72) 0.12 154/10 3000 88 / 43 14.24 (0.99 – 204.8) 0.05 157/7