MSACL 2016 EU Abstract

Investigation of Insulin-Mediated Hypoglycemia in the Living and the Dead: The Role of LC-MS/MS

Daniel Holmes (Presenter)
St. Paul's Hospital

Bio: Daniel Holmes did his undergraduate degree in Chemical Physics from the University of Toronto with a focus on Quantum Mechanics. He went to medical school at the University of British Columbia (UBC) where he also did his residency in Medical Biochemistry. He is a Clinical Associate Professor of Pathology and Laboratory Medicine at UBC and Division Head of Clinical Chemistry at St. Paul's Hospital in Vancouver. Interests include laboratory medicine statistics, clinical endocrinology with a focus on secondary hypertension, clinical lipidology and clinical mass spectrometry. Assay development efforts in the last five years have focused on novel use of mass spectrometry for assays directed at specialized endocrine testing.

Authorship: J Grace Van Der Gugten (1), Daniel T. Holmes (1,2)
(1) St. Paul's Hospital, Dept of Pathology and Laboratory Medicine, Vancouver, BC, Canada (2) University of British Columbia Department of Pathology and Laboratory Medicine, Vancouver, BC, Canada.

Short Abstract

Spontaneous adult hypoglycemia can be caused by insulinoma, autoantibodies against insulin or its receptor, over-administration of insulin secretagogues, and over-administration of exogenous insulin. Immunoassays for exogenous insulin do not uniformly detect synthetic analogues if one or more of the antibodies are directed against the C-terminal of the B-chain. For example the Roche assay detects none of the synthetic analogues except those with natural insulin peptide sequence (Insulin R and Insulin NPH). We have developed an immunoprecipitation LC-MS/MS method for insulin analogues and will discuss case experience in cases of accidental and intentional over-administration of insulin and the role of LC-MS/MS in case elucidation.

Long Abstract


Automated immunoassays for insulin demonstrate heterogenous analytical cross-reactivity with synthetic analogue insulins. In cases of spontaneous adult hypoglycemia,clinicians frequently request analysis for insulin secretagogues in the serum or urine and understand the need to interact with clinical laboratorians for this purpose. However, the same clinicians request insulin analysis without an understanding that immunoassays may yield confusing and/or misleading results and that care needs to be paid to which specimen is selected for analysis and how it is stored. In cases having legal ramifications, attention to detail in these matters becomes particularly important.


Insulin was measured as previously published [1] in 14 cases of suspected insulin overadministration as requested by forensic pathologist or clinician on heterogeneous sample types of unpredictable quality and integrity.


Numerous challenges were faced with regard to the specimen and the interpretation of the results. While some specimens were deteriorated to the point that there was no insulin signal, there were a number of cases where high insulin concentration could be confirmed and insulin-mediated hypoglycemia caused by synthetic analogue could be confidently identified by piecing together the clinical story, plasma and/or vitreous glucose, beta-hydroxybutyrate, C peptide and insulin concentration. Some modifications to the method were undertaken to detect glargine metabolite as will be discussed. We will discuss the collective experience of these cases and provide summary advice on specimen handling, analysis and reporting.


Immunoprecipitation LC-MS/MS provides a means to confidently identify the presence of insulin analogues providing clinical clarification to cases of accidental or intentional insulin overadministration.

References & Acknowledgements:

1. Van Der Gugten JG, Wong S, Holmes DT. Quantitation of Insulin Analogues in Serum Using Immunoaffinity Extraction, Liquid Chromatography, and Tandem Mass Spectrometry. Clinical Applications of Mass Spectrometry in Biomolecular Analysis: Methods and Protocols. 2016:119-30.

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