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Clinical Chemistry and Laboratory Medicine (CCLM)

Published in Association with the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM)

Editor-in-Chief: Plebani, Mario

Ed. by Gillery, Philippe / Greaves, Ronda / Lackner, Karl J. / Lippi, Giuseppe / Melichar, Bohuslav / Payne, Deborah A. / Schlattmann, Peter


IMPACT FACTOR 2018: 3.638

CiteScore 2018: 2.44

SCImago Journal Rank (SJR) 2018: 1.191
Source Normalized Impact per Paper (SNIP) 2018: 1.205

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1437-4331
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Volume 56, Issue 11

Issues

Development and validation of a mass spectrometry-based assay for quantification of insulin-like factor 3 in human serum

Jakob AlbrethsenORCID iD: http://orcid.org/0000-0002-2842-1128
  • Corresponding author
  • Department of Growth and Reproduction, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark; and Reprounion and International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health (EDMaRC), Rigshospitalet, University of Copenhagen, Section 5064, Rigshospitalet, Blegdamsvej 9, 2100, Copenhagen, Denmark, Phone: (+45) 3545 5085
  • orcid.org/0000-0002-2842-1128
  • Email
  • Other articles by this author:
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/ Hanne Frederiksen
  • Department of Growth and Reproduction, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark; and Reprounion and International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health (EDMaRC), Rigshospitalet, University of Copenhagen, Copenhagen, Denmark
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/ Anna-Maria Andersson
  • Department of Growth and Reproduction, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark; and Reprounion and International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health (EDMaRC), Rigshospitalet, University of Copenhagen, Copenhagen, Denmark
  • Other articles by this author:
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/ Ravinder Anand-Ivell / Loa Nordkap
  • Department of Growth and Reproduction, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark; and Reprounion and International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health (EDMaRC), Rigshospitalet, University of Copenhagen, Copenhagen, Denmark
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/ Anne Kirstine Bang
  • Department of Growth and Reproduction, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark; and Reprounion and International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health (EDMaRC), Rigshospitalet, University of Copenhagen, Copenhagen, Denmark
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/ Niels Jørgensen
  • Department of Growth and Reproduction, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark; and Reprounion and International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health (EDMaRC), Rigshospitalet, University of Copenhagen, Copenhagen, Denmark
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  • De Gruyter OnlineGoogle Scholar
/ Anders Juul
  • Department of Growth and Reproduction, Rigshospitalet, University of Copenhagen, Copenhagen, Denmark; and Reprounion and International Center for Research and Research Training in Endocrine Disruption of Male Reproduction and Child Health (EDMaRC), Rigshospitalet, University of Copenhagen, Copenhagen, Denmark
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Published Online: 2018-05-30 | DOI: https://doi.org/10.1515/cclm-2018-0171

Abstract

Background:

The circulating level of the peptide hormone insulin-like factor 3 (INSL3) is a promising diagnostic marker reflecting Leydig cell function in the male. Few commercial immunoassays of varying quality exist. Therefore, we decided to develop and validate a precise method for quantification of INSL3 by mass spectrometry.

Methods:

We developed an assay in which the INSL3 A-chain is released from the INSL3 A-B heterodimer by chemical reduction and alkylation. The alkylated INSL3 A-chain is quantitated by liquid chromatography-tandem mass spectrometry (LC-MS/MS), as substitute for serum INSL3. The method was compared to a validated and sensitive in-house serum INSL3 immunoassay using 97 serum samples from 12 healthy boys during pubertal transition. Adult levels were determined based on sera from 72 adult healthy males aged 18–40 years.

Results:

An LC-MS/MS assay with limit of detection and limit of quantification (LOQ) of 0.06 and 0.15 ng/mL, respectively, and intra-assay CVs <9% in the relevant ranges was obtained. The LC-MS/MS compared well with the in-house immunoassay (Deming regression slope: 1.28; Pearson correlation: R=0.86). INSL3 concentrations increased with pubertal maturation in healthy boys. INSL3 concentrations were above the LOQ in all samples from the adult men. The mean (±2 SD range)for serum INSL3 concentrations in the adult men was 2.2 (0.5–3.9) ng/mL.

Conclusions:

We have developed a robust and sensitive method suitable for quantitation of serum INSL3 in a clinical setting using LC-MS/MS instrumentation available in modern clinical laboratories. The method paves the way for future studies into the clinical role of serum INSL3 measurements.

This article offers supplementary material which is provided at the end of the article.

Keywords: assay development; insulin-like factor 3 (INSL3); mass spectrometry

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About the article

Received: 2018-02-14

Accepted: 2018-05-03

Published Online: 2018-05-30

Published in Print: 2018-10-25


Author contributions: All the authors have accepted responsibility for the entire content of this submitted manuscript and approved submission.

Research funding: Innovation Fund Denmark 14-2013-4; International Center for Research, and Research Training in Endocrine Disruption of Male Reproduction and Child Health (EDMaRC); ReproUnion.

Employment or leadership: None declared.

Honorarium: None declared.

Competing interests: The funding organization(s) played no role in the study design; in the collection, analysis, and interpretation of data; in the writing of the report; or in the decision to submit the report for publication.


Citation Information: Clinical Chemistry and Laboratory Medicine (CCLM), Volume 56, Issue 11, Pages 1913–1920, ISSN (Online) 1437-4331, ISSN (Print) 1434-6621, DOI: https://doi.org/10.1515/cclm-2018-0171.

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