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Accepted on ,

Technical Briefs

Feasibility of Standardization of Serum C-Peptide Immunoassays with Isotope-Dilution Liquid Chromatography-Tandem Mass Spectrometry

¹ Laboratory for Analytical Chemistry, Faculty of Pharmaceutical Sciences, Ghent University, Gent, Belgium
² Laboratory for Hormonology and Department of Endocrinology, Ghent University Hospital, Gent, Belgium

^* To whom correspondence should be addressed. E-mail: linda.thienpont{at}ugent.be.

Background: Serum C-peptide concentrations reflect pancreatic function in different clinical and diagnostic settings; however, the utility of C-peptide testing is limited by the lack of standardized commercial immunoassays. Standardization can best be done by split-sample comparison with a hierarchically higher reference measurement procedure with a set of native sera. For serum peptides, isotope-dilution liquid chromatography\Nmass spectrometry (ID-LC/MS) is recommended as a reference measurement procedure.

Methods: We evaluated the analytical performance characteristics of an ID-LC/tandem MS procedure for measurement of serum C-peptide after a 2-step solid-phase extraction. To investigate the feasibility of this procedure for use in standardization, we also performed a method comparison with 3 representative commercial assays.

Results: The ID-LC/tandem MS procedure showed maximum within-run, between-run, and total CVs on dedicated sera (C-peptide concentrations, 1.6 and 4.0 µg/L) of 2.1%, 2.5%, and 2.9%, respectively; an accuracy of 94.6%-104.1%; a mean minimum trueness of 98.1% (95% confidence interval, 96.2%-100.0%), and limits of quantification and detection of 0.15 and 0.03 µg/L, respectively. Deming linear regression analysis of the method-comparison data showed that the immunoassays correlated well with ID-MS and were specific, but lacked intercomparability and trueness. We propose that the deficiencies can be resolved by recalibration on the basis of the method comparison.

Conclusions: The ID-LC/tandem MS procedure is suitable for specific and accurate measurement of basal and stimulated serum concentrations of proinsulin C-peptide fragment 33-63 and is suitable for use in standardization of C-peptide immunoassays.