Validation of suPAR turbidimetric assay on Cobas® (c502 and c702) and comparison to suPAR ELISA

Abstract

suPAR is a plasma marker of chronic inflammation, and an elevated suPAR is consistently associated with worse outcome in a variety of clinical conditions. Quantification of suPAR is useful for determining patient risk in triage, but there is no fast automatized method for quick determination of suPAR. We developed and validated a rapid latex particle-enhanced turbidimetric immunoassay for quantification of plasma suPAR on the c502 and the c702 Roche Cobas® 8000 measurment systems. The turbidimetric assay was validated against the suPARnostic® ELISA (ViroGates, Denmark). This validation demonstrates suPAR can be analysed by turbidimetry giving very similar results (<15% difference) compared to the ELISA method and the observed correlations (n = 103) were strong, r > 0.95. Roche Cobas® 8000 instruments demonstrated repeatability and repoducibility, CV % at 3.4–4.1 and 5.7–11.4, respectively. The estimated limit of detection was 1.30 µg/L and 1.31 µg/L for the Cobas® c502 and c702, respectively. Dilution tests showed linearity of suPAR from 1.8 to 26.5 μg/L. The acceptable concentrations of Bilirubin, Intralipid and Hemoglobin, were 350 µmol/L, 3.3 g/L and 1.4 g/L, respectively. suPAR can be quantified reproducibly within 10 min using a turbidimetry assay. This assay is faster than ELISA with similar results, making it suitable for clinical routine analysis.

Keywords:

• Receptors
• soluble urokinase plasminogen activator
• validation studies
• immunoturbidimetry assay
• ELISA
• biomarker

Acknowledgements

We would acknowledge Hanne Mathiesen, Bioanalytical Specialist Technician at Department of Biochemistry and Immunology, Hospital of Southern Jutland, for skilful laboratory work and analysis of the samples in this assay validation study. Lisbeth Søndergaard, Application Specialist, Roche Diagnostics A/S, Hvidovre, Denmark is acknowledged for the support in the development of the suPAR assay application program for Cobas® 8000, model c502 and c702.

Author contributions

Thor A. Skovsted developed the Cobas® c502 and c702 methods based on key information of a similar assay developed at Cobas® c111, determined the validation design and managed the analysis of samples and reported the validation results in this paper. Eva Rabing Brix Petersen contributed with the clinical application of suPAR with discussions, layout, and writing this manuscript. Maj-Britt Fruekilde contributed with discussions, layout, and writing this manuscript. Andreas Kristian Pedersen performed the statistical analysis of the validation result. Tomasz Pielak delivered key information of a similar assay developed at Cobas® c111, and contributed in the discussion of the validation design and supplied turbidimetric reagents, calibrators, ELISA analysis and plasma samples for comparison. Jesper Eugen-Olsen suggested to develop a turbidimetric assay and contributed with clinical application of suPAR with discussions, layout, writing this manuscript. All authors approved the final version of the paper.

Disclosure Statement

Jesper Eugen-Olsen is shareholder, co-founder and the CSO of ViroGates (NASDAQ:VIRO), the company that produce the suPARnostic technologies. Tomasz Pielak is CEO in NUTOPI Sp. a subcontractor to ViroGates A/S.