Out-of-the-black-box: Standardisation of the EliSpot Method for immunomonitoring in personalized Immunosupression after solid organ tranplantation

Abstract

Abstract Individual Immunotherapy for transplantation patients has been moved more and more into the focus of transplant centers, as individualized therapy approaches enable to minimize long-term side effects and increase the quality of life for the patients. The Enzyme-Linked ImmunoSpot assay (EliSpot) has become a powerful tool in solid organ and stem cell transplantation. In an European multicenter study, the Bio-DrIM project c, the use of the EliSpot has been extensively evaluated with regard to monitoring of viral (Epstein-Barr Virus, EBV; Human Cytomegalovirus, HCMV and BK Virus, BKV) reactivation during immunosuppressive therapy after kidney transplantation. The reference institute established SOPs and trained the attending sites. Additionally, EliSpot Reader Systems and EliSpot kits were extensively technical validated to guarantee that all centers reach the same results. A total of 21 plates with 1983 wells were counted. Linear regression of all 1983 pairs of values lead to a linear regression line with the equation y=1.026*x+0,06888 and a Pearson correlation coefficient R2=9988, demonstrating the high concordance of each reader system. To further demonstrate the extraordinary high standardization of the procedure, Inter- and Intra-Assay evaluation were performed. In the set of analyzed results the CV % varied under 20 %, which is far below cellular assays specified by the FDA with 25 %. In conclusion, the combination of the EliSpot method with the EliSpot Reader System is a high-performance tool for diagnostics and monitoring of immunosuppression. Based on the high standardization potential, this state-of the-art technology will play an important role in diagnostic validation of next generation personalized therapy.