Journal
AMERICAN JOURNAL OF TRANSPLANTATION
Volume 13, Issue 7, Pages 1871-1879Publisher
WILEY
DOI: 10.1111/ajt.12286
Keywords
Alloreactive T cells; biomarker; cytokine secretion assay; ELISPOT; flow cytometry; guidelines
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Funding
- National Institute of Allergy and Infectious Diseases [U0163594]
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Emerging evidence indicates memory donor-reactive T cells are detrimental to transplant outcome and that quantifying the frequency of IFN-producing, donor-reactive PBMCs by ELISPOT has potential utility as an immune monitoring tool. Nonetheless, differences in assay performance among laboratories limit the ability to compare results. In an effort to standardize assays, we prepared a panel of common cellular reagent standards, developed and cross validated a standard operating procedure (SOP) for alloreactive IFN ELISPOT assays in several research laboratories supported by the NIH-funded Clinical Trials in Organ Transplantation (CTOT) Consortium. We demonstrate that strict adherence to the SOP and centralized data analysis results in high reproducibility with a coefficient of variance (CV) of approximate to 30%. This standardization of IFN ELISPOTassay will facilitate interpretation of data from multicenter transplantation research studies and provide the foundation for developing clinical laboratory testing strategies to guide therapeutic decision-making in transplant patients.
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