B-243 Validating a highly automated, rapid turnaround urine biomarker assay for diagnosis of acute interstitial nephritis

Abstract

Abstract Background Urinary tumor necrosis factor alpha (TNF-α) and chemokine C-X-C motif ligand 9 (CXCL9) have been identified as potential diagnostic biomarkers for acute interstitial nephritis (AIN). Although validated on standard methods like ELISA and mesoscale discovery (MSD), these platforms have a turnaround time of greater than 4hr. Ella, is a new platform which takes around 90 minutes and is more suited to be used for diagnosis of AIN in hospital setting. However, validation data for urine TNF-α and CXCL-9 in Ella assays are currently unavailable. Methods Five samples were selected for dilution-linearity and spike-recovery analysis. At the time of collection, samples were centrifuged at 2,000xg and 4°C for 10 min and the supernatant was aliquoted and stored at -80°C until assayed. We investigated dilution linearity by performing serial dilutions at three levels (Neat, 1:2, 1:4) assayed in duplicate. We performed spike and recovery experiment at three different concentration levels (high, medium, and assay diluent) by adding 5% spike volume of mixed TNF-α and CXCL-9 calibrator or assay diluent to the samples. We defined acceptance when recovery was between 80% - 120%. Intraassay %CV was calculated by averaging %CV of duplicate runs across all experiments. Inter-assay %CV was calculated by averaging %CV three levels of QC over three days. Samples were analyzed on MSD for method comparison and correlation coefficients (Pearson and Spearman) were calculated between the two measurements. Results Average recoveries and MSD comparison for TNF-α and CXCL9 are in the table below. Diluting the urine samples 4-fold yielded acceptable recoveries between 80% - 120%. Comparison between MSD and Ella yielded strong correlation for CXCL9 and moderate correlation for TNF-α. Conclusions The Ella platform can be used to measure urinary TNF-α and CXCL9 with good precision and acceptable recoveries at 1:4 dilution. Ella measurements compare favorably with those of MSD.