Daniel S Herman1, Pratistha Ranjitkar1, Diane Yamaguchi1, David G Grenache2, Dina N Greene3. 1. Department of Laboratory Medicine, University of Washington, Seattle, WA, USA. 2. Department of Pathology, University of Utah School of Medicine, Salt Lake City, UT, USA. 3. Department of Laboratory Medicine, University of Washington, Seattle, WA, USA. Electronic address: dngreene@uw.edu.
Abstract
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 (cTnI) results reported by the DxI AccuTnI+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 cTnI 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 DxI cTnI and human chorionic gonadotropin (hCG). Next, cTnI and/or hCG was measured in specimens with normal (N=20) or elevated (N=26) ALP using DxI and Vista assays. Finally, cTnI was quantified using a prototype, ALP-dependent high-sensitivity assay. RESULTS: The sentinel sample's DxI-cTnI results were imprecise on repeat, linear on dilution, unaffected by heterophile blocking antibodies, and correlated with ALP lability following heat treatment. The Vista-cTnI concentrations were ~7-fold lower. Addition of purified ALP to reference serum linearly increased the DxI-cTnI results. DxI-hCG results also appeared affected by ALP. Several independent patients' specimens with elevated ALP appeared to have falsely elevated DxI-cTnI and DxI-hCG. CONCLUSIONS: Elevated ALP can interfere with contemporary, ALP-dependent immunoassays, including DxI-cTnI and DxI-hCG. The validation of such methods should include evaluations for endogenous ALP interference. Specimens with ALP >1000U/L and elevated DxI-cTnI should be evaluated for ALP interference.
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 (cTnI) results reported by the DxI AccuTnI+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 cTnI 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 DxI cTnI and human chorionic gonadotropin (hCG). Next, cTnI and/or hCG was measured in specimens with normal (N=20) or elevated (N=26) ALP using DxI and Vista assays. Finally, cTnI was quantified using a prototype, ALP-dependent high-sensitivity assay. RESULTS: The sentinel sample's DxI-cTnI results were imprecise on repeat, linear on dilution, unaffected by heterophile blocking antibodies, and correlated with ALP lability following heat treatment. The Vista-cTnI concentrations were ~7-fold lower. Addition of purified ALP to reference serum linearly increased the DxI-cTnI results. DxI-hCG results also appeared affected by ALP. Several independent patients' specimens with elevated ALP appeared to have falsely elevated DxI-cTnI and DxI-hCG. CONCLUSIONS: Elevated ALP can interfere with contemporary, ALP-dependent immunoassays, including DxI-cTnI and DxI-hCG. The validation of such methods should include evaluations for endogenous ALP interference. Specimens with ALP >1000U/L and elevated DxI-cTnI should be evaluated for ALP interference.