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Development of an ID-LC-MS/MS method using targeted proteomics for quantifying cardiac troponin I in human serum.
Asicioglu, Meltem; Oztug, Merve; Karaguler, Nevin Gul.
Afiliación
  • Asicioglu M; TUBITAK National Metrology Institute (TUBITAK UME), Gebze, 41400, Kocaeli, Turkey.
  • Oztug M; Department of Molecular Biology and Genetics, Faculty of Science and Letters, Istanbul Technical University, Istanbul, Turkey.
  • Karaguler NG; Dr. Orhan Ocalgiray Molecular Biology-Biotechnology and Genetics Research Center, Istanbul Technical University, Istanbul, Turkey.
Clin Proteomics ; 20(1): 40, 2023 Sep 27.
Article en En | MEDLINE | ID: mdl-37759177
BACKGROUND: Cardiac troponin is a complex protein consisting of the three subunits I, T and C located in heart muscle cells. When the heart muscle is damaged, it is released into the blood and can be detected. Cardiac troponin I (cTnI) is considered the most reliable and widely accepted test for detecting and confirming acute myocardial infarction. However, there is no current standardization between the commercial assays for cTnI quantification. Our work aims to create a measurement procedure that is traceable to the International System of Units for accurately measuring cardiac cTnI levels in serum samples from patients. METHODS: The workflow begins with immobilizing anti-cTnI antibodies onto magnetic nanoparticles to form complexes. These complexes are used to isolate cTnI from serum. Next, trypsin is used to enzymatically digest the isolated cTnI. Finally, the measurement of multiple cTnI peptides is done simultaneously using isotope dilution liquid chromatography-tandem mass spectrometry (ID-LC-MS/MS). RESULTS: The maximum antibody immobilization was achieved by combining 1 mg of nanoparticles with 100 µg of antibody, resulting in an average of 59.2 ± 5.7 µg/mg of immobilized antibody. Subsequently, the anti-cTnI-magnetic nanoparticle complex was utilized to develop and validate a method for quantifying cTnI in human serum using ID-LC-MS/MS and a protein calibration approach. The analytical method was assessed regarding linearity and recovery. The developed method enables the quantification of cTnI from 0.7 to 24 µg/L (R > 0.996). The limit of quantification was 1.8 µg/L and the limit of detection was 0.6 µg/L. Intermediate precision was ≤ 9.6% and repeatability was 2.0-8.7% for all quality control materials. The accuracy of the analyzed quality control materials was between 90 and 110%. Total measurement uncertainties for target value assignment (n = 6) were found to be ≤ 12.5% for all levels. CONCLUSIONS: The analytical method demonstrated high analytical performance in accurately quantifying cardiac troponin I levels in human serum. The proposed analytical method has the potential to facilitate the harmonization of cTnI results between clinical laboratories, assign target values to secondary certified reference materials and support reliable measurement of cTnI.
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Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Idioma: En Revista: Clin Proteomics Año: 2023 Tipo del documento: Article País de afiliación: Turquía Pais de publicación: Reino Unido

Texto completo: 1 Colección: 01-internacional Base de datos: MEDLINE Idioma: En Revista: Clin Proteomics Año: 2023 Tipo del documento: Article País de afiliación: Turquía Pais de publicación: Reino Unido