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Quantification of cardiac troponin I in patient plasma by magnetic particle immunoenrichment and targeted mass spectrometry



Nicole A. Schneck, Karen W. Phinney, Sang Bok Lee, Mark Lowenthal


Absolute quantification of the protein biomarker cardiac troponin I (cTnI) has been achieved in patients' plasma based on a selective and sensitive isotope dilution (ID) liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. Here, an immunoaffinity enrichment strategy coupled with LC-MS/MS was developed to measure cTnI in plasma samples obtained from patients who had undergone moderate-to-severe myocardial infarctions. The key steps in this approach involved isolating cTnI from plasma using anti-cTnI antibody coupled to magnetic nanoparticles, followed by an enzymatic digestion with trypsin. Three tryptic peptides from cTnI were monitored and used for quantification by ID-LC-MS/MS via multiple reaction monitoring (MRM). A full-length isotopically labeled protein analog of cTnI was pre-spiked into patient plasma samples, allowing for absolute quantification of the diagnostic biomarker. The method was applied to determine the concentration of cTnI in five patient plasma samples, with concentrations measuring between 4.83 ng/mL and 11.3 ng/mL. The LC-MS/MS measurement precision was based on three unique peptides from cTnI using two MRM transitions per peptide. Average relative standard deviation (CV) from the five patient plasma samples was determined to be 14.3 %. Overall, this study is the first to achieve quantification of multiple peptides from cTnI in native (not spiked) plasma based on an ID-LC-MS/MS method, using matrix-matched calibrants and a protein internal standard. A precise and robust method is the first step in developing a reference measurement procedure for cTnI in plasma, and for the certification of quantitative reference materials that can be used to harmonize cTnI clinical assays.
Analytical Chemistry


troponin I, mass spectrometry, clinical biomarker, clinical assay, LC-MS


Schneck, N. , Phinney, K. , Lee, S. and Lowenthal, M. (2018), Quantification of cardiac troponin I in patient plasma by magnetic particle immunoenrichment and targeted mass spectrometry, Analytical Chemistry (Accessed July 5, 2022)
Created April 17, 2018, Updated October 12, 2021