Analytical and Bioanalytical Chemistry

, Volume 409, Issue 14, pp 3677–3684 | Cite as

Immunoassay quantification of human insulin added to ternary parenteral nutrition containers: comparison of two methods

  • Héloïse Henry
  • Damien Lannoy
  • Nicolas Simon
  • David Seguy
  • Michèle D’Herbomez
  • Christine Barthélémy
  • Bertrand Décaudin
  • Thierry Dine
  • Pascal Odou
Research Paper


Adding insulin directly into infusion bags seems to be a useful method for controlling hyperglycemia in patients under ternary parenteral nutrition (TPN). Its efficacy is assessed by glycemic monitoring but few data are available on insulin stability in this situation. Among the various methods for quantifying insulin levels in human serum, the immunoassay ones seemed potentially appropriate for a TPN admixture containing high lipid concentrations. We sought to identify and validate which of two immunoassay methods was the better to quantify human insulin and consequently be adapted to studying its stability in a TPN admixture. Two immunoassay methods to quantify recombinant human insulin were assessed in industrial TPN: an immunoradiometric assay (IRMA) and an immunoelectrochemiluminometric assay (IECMA). Validation trials for both methods were based on the accuracy profile method. Interference with immunometric assays due to the high lipidic content of TPN was eliminated through an improved preparation protocol using a bovine serum albumin (BSA) diluted in phosphate buffer saline (PBS). The relative total error of IECMA varied from 1.74 to 4.52% while it varied from −0.32 to 8.37% with IRMA. Only IECMA provided an accuracy profile with a 95% confidence interval of calculated-tolerance limits falling between the chosen acceptance limits (i.e., total error ≤±10%). IECMA combined with a BSA dilution is a simple and semi-automatic method that provides an accurate quantification of human insulin in a TPN admixture without any interference from lipids.


Human insulin Parenteral nutrition solutions Immunoassay Validation Accuracy profile 



The authors wish to thank Mrs. Alexandra Tavernier, M.A. University of Glasgow, U.K.; Professeur Agrégée, University of Lille, for her extensive revision of the final manuscript.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. 1.
    McMahon MM, Nystrom E, Braunschweig C, Miles J, Compher C, American Society for Parenteral and Enteral Nutrition (A.S.P.E.N.) Board of Directors, et al. A.S.P.E.N. Clinical guidelines: nutrition support of adult patients with hyperglycemia. JPEN J Parenter Enteral Nutr. 2013;37:23–36.CrossRefGoogle Scholar
  2. 2.
    Edakkanambeth Varayil J, Yadav S, Miles JM, Okano A, Kelly DG, Hurt RT, et al. Hyperglycemia during home parenteral nutrition administration in patients without diabetes. JPEN J Parenter Enteral Nutr. 2015.Google Scholar
  3. 3.
    Olveira G, Tapia MJ, Ocón J, Cabrejas-Gómez C, Ballesteros-Pomar MD, Vidal-Casariego A, et al. Parenteral nutrition-associated hyperglycemia in non-critically ill inpatients increases the risk of in-hospital mortality (multicenter study). Diabetes Care. 2013;36:1061–6.CrossRefGoogle Scholar
  4. 4.
    Mirtallo J, Canada T, Johnson D, Kumpf V, Petersen C, Sacks G, et al. Safe practices for parenteral nutrition. JPEN J Parenter Enteral Nutr. 2004;28:S39–70.Google Scholar
  5. 5.
    Bassons T, Maria Sanchez J, Bassas L, Cardona D, Ordonez J, Bonal J. Insulina recuperada según los componentes de la nutrición parenteral. Rev SENPE. 1985;4:93–9.Google Scholar
  6. 6.
    Rusavý Z, Sramek V, Suchat R, Lacigova S, Topolcan O. Effects of carrier solution on insulin bioavailability. JPEN J Parenter Enteral Nutr. 2004;28:439–41.CrossRefGoogle Scholar
  7. 7.
    Yilmaz B, Kadioglu Y, Capoglu I. Determination of insulin in humans with insulin-dependent diabetes mellitus patients by HPLC with diode array detection. J Chromatogr Sci. 2012;50:586–90.CrossRefGoogle Scholar
  8. 8.
    Hoyer GL, Nolan PE, LeDoux JH, Moore LA. Selective stability-indicating high-performance liquid chromatographic assay for recombinant human regular insulin. J Chromatogr A. 1995;699:383–8.CrossRefGoogle Scholar
  9. 9.
    Yu K-H, Tsao H-L, Lin S-J, Chen C-Y. Quantitative analysis of insulin in total parenteral nutrition bag in Taiwan. J Food Drug Anal. 2016;24:214–9.CrossRefGoogle Scholar
  10. 10.
    Oliva A, Fariña J, Llabrés M. Development of two high-performance liquid chromatographic methods for the analysis and characterization of insulin and its degradation products in pharmaceutical preparations. J Chromatogr B Biomed Sci App. 2000;749:25–34.CrossRefGoogle Scholar
  11. 11.
    Hess C, Thomas A, Thevis M, Stratmann B, Quester W, Tschoepe D, et al. Simultaneous determination and validated quantification of human insulin and its synthetic analogues in human blood serum by immunoaffinity purification and liquid chromatography-mass spectrometry. Anal Bioanal Chem. 2012;404:1813–22.CrossRefGoogle Scholar
  12. 12.
    Marcuard SP, Dunham B, Hobbs A, Caro JF. Availability of insulin from total parenteral nutrition solutions. JPEN J Parenter Enteral Nutr. 1990;14:262–4.CrossRefGoogle Scholar
  13. 13.
    Martínez-Subiela S, Cerón JJ. Effects of hemolysis, lipemia, hyperbilirrubinemia, and anticoagulants in canine C-reactive protein, serum amyloid A, and ceruloplasmin assays. Can Vet J. 2005;46:625–9.Google Scholar
  14. 14.
    Owen WE, Roberts WL. Cross-reactivity of three recombinant insulin analogs with five commercial insulin immunoassays. Clin Chem. 2004;50:257–9.CrossRefGoogle Scholar
  15. 15.
    Heurtault B, Reix N, Meyer N, Gasser F, Wendling M-J, Ratomponirina C, et al. Extensive study of human insulin immunoassays: promises and pitfalls for insulin analogue detection and quantification. Clin Chem Lab Med. 2014;52:355–62.CrossRefGoogle Scholar
  16. 16.
    Manley SE, Stratton IM, Clark PM, Luzio SD. Comparison of 11 human insulin assays: implications for clinical investigation and research. Clin Chem. 2007;53:922–32.CrossRefGoogle Scholar
  17. 17.
    Agin A, Jeandidier N, Gasser F, Grucker D, Sapin R. Use of insulin immunoassays in clinical studies involving rapid-acting insulin analogues: bi-insulin IRMA preliminary assessment. Clin Chem Lab Med. 2006;44:1379–82.CrossRefGoogle Scholar
  18. 18.
    Sapin R, Le Galudec V, Gasser F, Pinget M, Grucker D. Elecsys insulin assay: free insulin determination and the absence of cross-reactivity with insulin lispro. Clin Chem. 2001;47:602–5.Google Scholar
  19. 19.
    Cassidy JP, Luzio SD, Marino MT, Baughman RA. Quantification of human serum insulin concentrations in clinical pharmacokinetic or bioequivalence studies: what defines the “best method”? Clin Chem Lab Med. 2012;50:663–6.CrossRefGoogle Scholar
  20. 20.
    Levy-Marchal C, Bridel MP, Sodoyez-Goffaux F, Koch M, Tichet J, Czernichow P, et al. Superiority of radiobinding assay over ELISA for detection of IAAs in newly diagnosed type I diabetic children. Diabetes Care. 1991;14:61–3.CrossRefGoogle Scholar
  21. 21.
    Hubert P, Nguyen-Huu J-J, Boulanger B, Chapuzet E, Chiap P, Cohen N, et al. Harmonization of strategies for the validation of quantitative analytical procedures. A SFSTP proposal—part II. J. Pharm. Biomed. Anal. 2007;45:70–81.Google Scholar
  22. 22.
    COFRAC. SH GTA 04 Guide technique d’accréditation de vérification (portée A)/validation (portée B) des méthodes en biologie médicale [Internet]. 2011 [cited 2017 Jan 10]. Available from:
  23. 23.
    Kelley M, DeSilva B. Key elements of bioanalytical method validation for macromolecules. AAPS J. 2007;9:E156–63.CrossRefGoogle Scholar
  24. 24.
    Christianson MA, Schwartz MW, Suzuki N. Determinants of insulin availability in parenteral nutrition solutions. JPEN J Parenter Enteral Nutr. 2006;30:6–9.CrossRefGoogle Scholar
  25. 25.
    Ciszewska M, Knyt A, Kopec B, Pertkiewicz M. Insulin availability from all in one and fat-free nutrients admixtures. Clin Nutr. 1994;13(Supplement 1):56.CrossRefGoogle Scholar
  26. 26.
    Sapin R. Interferences in immunoassays: mechanisms and outcomes in endocrinology. Ann Endocrinol. 2008;69:415–25.CrossRefGoogle Scholar
  27. 27.
    Ichikawa E, Kimura M, Mori H, Yamazaki F, Hirano K. Apparent increase of insulin peak area in HPLC analysis of a preparation consisting of a mixture of insulin and total parenteral nutrition. Chem Pharm Bull (Tokyo). 2006;54:1196–9.CrossRefGoogle Scholar
  28. 28.
    Saracevic A, Nikolac N, Simundic A-M. The evaluation and comparison of consecutive high speed centrifugation and LipoClear® reagent for lipemia removal. Clin Biochem. 2014;47:309–14.CrossRefGoogle Scholar
  29. 29.
    Livesey JH, Donald RA. Prevention of adsorption losses during radioimmunoassay of polypeptide hormones: effectiveness of albumins, gelatin, caseins, Tween 20 and plasma. Clin Chim Acta Int J Clin Chem. 1982;123:193–8.CrossRefGoogle Scholar
  30. 30.
    Andersen L, Jørgensen PN, Jensen LB, Walsh D. A new insulin immunoassay specific for the rapid-acting insulin analog, insulin aspart, suitable for bioavailability, bioequivalence, and pharmacokinetic studies. Clin Biochem. 2000;33:627–33.CrossRefGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2017

Authors and Affiliations

  • Héloïse Henry
    • 1
    • 2
  • Damien Lannoy
    • 1
    • 2
  • Nicolas Simon
    • 1
    • 2
  • David Seguy
    • 3
    • 4
  • Michèle D’Herbomez
    • 5
  • Christine Barthélémy
    • 1
  • Bertrand Décaudin
    • 1
    • 2
  • Thierry Dine
    • 1
  • Pascal Odou
    • 1
    • 2
  1. 1.Univ. Lille, EA 7365 - GRITA - Groupe de Recherche sur les formes Injectables et les Technologies AssociéesLilleFrance
  2. 2.Institut de PharmacieCHU LilleLilleFrance
  3. 3.Univ. Lille, U995 – LIRIC – Lille Inflammation Research International CenterLilleFrance
  4. 4.Department of NutritionCHU LilleLilleFrance
  5. 5.Immunoanalysis Center, Biology and Pathology CenterCHU LilleLilleFrance

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