Multiplexed quantification of insulin and C-peptide by LC-MS/MS without the use of antibodies.

Autor: Foulon N; Department of Laboratory Medicine & Pathology, University of Washington, Seattle, WA, USA., Goonatilleke E; Department of Laboratory Medicine & Pathology, University of Washington, Seattle, WA, USA., MacCoss MJ; Department of Genome Sciences, University of Washington, Seattle, WA, USA., Emrick MA; Department of Laboratory Medicine & Pathology, University of Washington, Seattle, WA, USA., Hoofnagle AN; Department of Laboratory Medicine & Pathology, University of Washington, Seattle, WA, USA.; Department of Medicine, University of Washington, Seattle, WA, USA.
Jazyk: angličtina
Zdroj: Journal of mass spectrometry and advances in the clinical lab [J Mass Spectrom Adv Clin Lab] 2022 Jun 10; Vol. 25, pp. 19-26. Date of Electronic Publication: 2022 Jun 10 (Print Publication: 2022).
DOI: 10.1016/j.jmsacl.2022.06.003
Abstrakt: Introduction: The measurement of insulin and C-peptide provides a valuable tool for the clinical evaluation of hypoglycemia. In research, these biomarkers are used together to better understand hyperinsulinemia, hepatic insulin clearance, and beta cell function. Liquid chromatography-tandem mass spectrometry (LC-MS/MS) is an attractive approach for the analysis of insulin and C-peptide because the platform is specific, can avoid certain limitations of immunoassays, and can be multiplexed. Previously described LC-MS/MS methods for the simultaneous quantification of insulin and C-peptide measure the intact analytes and most have relied on immunoaffinity enrichment. These approaches can be limited in terms of sensitivity and interference from auto-antibodies, respectively. We have developed a novel method that does not require antibodies and uses proteolytic digestion to yield readily ionizable proteotypic peptides that enables the sensitive, specific, and simultaneous quantitation of insulin and C-peptide.
Methods: Serum samples were precipitated with acetonitrile. Analytes were enriched using solid phase extraction and then digested with endoproteinase Glu-C. Surrogate peptides for insulin and C-peptide were analyzed using targeted LC-MS/MS.
Results: Inter-day imprecision was below 20 %CV and linearity was observed down to the lower limit of quantitation for both analytes (insulin = 0.09 ng/mL, C-peptide = 0.06 ng/mL). Comparison to a commercially available insulin immunoassay (Beckman Coulter UniCel DxI 600 Access) revealed a 30% bias between methods.
Conclusion: A novel LC-MS/MS method for the simultaneous analysis of insulin and C-peptide using Glu-C digestion was developed and evaluated. A detailed standard operating procedure is provided to help facilitate implementation in other laboratories.
Competing Interests: The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
(© 2022 THE AUTHORS.)
Databáze: MEDLINE
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