Routine surveillance of kidney allograft rejection using a fully automated urinary CXCL9 and CXCL10 immunoassay

Abstract

Background For kidney transplant recipients, assessing non-invasively the individualized risk of acute rejection is one of the most unmet need. Urinary chemokines are one of the short-term most promising biomarkers, because of their simple and low-cost analytical method in easily accessible samples, and their high diagnostic performance consistently assessed over the last decade. In this study, we aimed at confronting all practical issues of routine implementation of kidney allograft rejection monitoring using urine CXCL9 and CXCL10. Methods The next-generation immunoassay Ella® was investigated as feasible quantification platform, from sample collection to render of the results. Urine CXCL9/10 levels were measured using Ella® microfluidic cartridges, across preanalytical (sample preparation, storage conditions, freeze-thaw cycles) and analytical (linearity, ranges, intra/inter-assay variability) performances studies, and diagnostic accuracy was assessed in comparison to the ELISA reference method, in urine samples from previously published cohorts. Results Upon assay preparation, Ella® appeared very efficient with a minimal workflow (urine sample thaw> centrifugation>1:2 dilution>loading) and a time to result of 90 minutes. Preanalytical studies showed high stability of urine CXCL9/10 levels across various temperatures (4° vs 25°C) and time (24/48/72H) before storage and over 5 freeze-thaw cycles. How complex urine matrix, analytical studies confirmed excellent linearity of dosage, as well as intra-assay (≤5%) and inter-assay precision. Across 1024 samples, Ella® results were highly correlated to those quantified by ELISA (P<0.0001), and further entered into our previously validated urine chemokine model. 268 out of the 1024 samples were collected at time of acute rejection (26.2%). In this cohort, accuracy was 0.85 for acute rejection diagnosis. Conclusion Ella® fulfils all prerequisites for clinical implementation of urinary chemokines monitoring. It has proven a robust, easy-to-use platform with unprecedented validation to quantify urine chemokines.