Endogenous alkaline phosphatase interference in cardiac troponin I and other sensitive chemilumihescence immunoassays that use alkaline phosphatase activity for signal amplification

Background: False positive cardiac troponin results can lead to inappropriate diagnosis. Our laboratory workflow includes systematic quality practices to identify false positive cardiac troponin I (cTnl) results reported by the Dxl AccuTnl+3 assay, which uses alkaline phosphatase (ALP) for signal amplification. Recently, a sample with elevated cTnI failed our quality standards and was found to have extremely elevated endogenous ALP activity. The objective of this study was to determine the true cTnI concentration and evaluate whether ALP was the source of interference. Methods: The suspicious cTnl result was evaluated by repeat analyses, dilution, heterophile blocking treatment, alternative methodology (Vista), and heat treatment Purified ALP was added to reference serum and we quantified Dxl cTnl and human chorionic gonadotropin (hCG). Next, cTnl and/or hCG was measured in specimens with normal (N = 20) or elevated (N = 26) ALP using Dxl and Vista assays. Finally, cTnI was quantified using a prototype, ALP-dependent high-sensitivity assay. Results: The sentinel sample's Dxl-cTnl results were imprecise on repeat, linear on dilution, unaffected by heterophile blocking antibodies, and correlated with ALP lability following heat treatment. The Vista-cTnl concentrations were similar to 7-fold lower. Addition of purified ALP to reference serum linearly increased the Dxl-cTnl results. Dxl-hCG results also appeared affected by ALP. Several independent patients' specimens with elevated ALP appeared to have falsely elevated Dxl-cTnl and Dxl-hCG. Conclusions: Elevated ALP can interfere with contemporary, ALP-dependent immunoassays, including Dxl-cTnl and Dxl-hCG. The validation of such methods should include evaluations for endogenous ALP interference. Specimens with ALP >1000 U/L and elevated Dxl-cTnl should be evaluated for ALP interference. (C) 2016 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.