Insulin-like growth factor I (IGF-I, a.k.a. somatomedin C) is a 7.6 kDa, 70 amino acid residue peptide, which mediates the actions of growth hormone (GH).1 IGF-I is synthesized as a prohormone, a polypeptide consisting of A, C, B, D, and E domains.1, 2 After post-translational modification, the mature IGF-I consist of the A, C, B and D domains, and is structurally homologous to IGF-II and insulin. In vivo, IGF-I is secreted by the liver and several other tissues and is postulated to have mitogenic and metabolic actions at or near the sites of synthesis; i.e. paracrine effects.1 IGF-I also appears in the peripheral circulation where it circulates primarily in a high molecular weight tertiary complex with IGF-binding protein-3 (IGFBP-3) and acid-labile subunit (ALS).2,3 A smaller proportion of IGF-I circulates in association with other IGF-binding proteins.3

Recently, there has been research interest in the measurement of serum/plasma “unbound” IGF-I which, theoretically, is the biologically active fraction. Although the existence of a true unbound IGF serum/plasma compartment is controversial, pharmacokinetic studies indicate that a small percentage of plasma IGF-I is not associated with IGF-binding proteins.4,5 Unbound IGF-I has also been observed in saliva.6 In addition, it appears that IGF-I may exert a tonic hypoglycemic effect under normal conditions that is inhibited by exogenous IGFBP-1 administration.7

It is likely that the measured unbound IGF-I fraction is a combination of the true unbound and the fraction of IGF-I that can be readily dissociated from IGFBP’s under the specific assay conditions. In this respect, it has been shown that exogenously administered IGF-I almost immediately associates with low MW IGFBP’s, then quickly moves into the high MW tertiary complex.5,8,9 The tertiary complex does not appear to be easily dissociated and does not re-equilibrate with exogenously added IGF-I or IGFBP-3 to a significant degree.8 On the other hand, the low MW complexes have a rapid turnover, and may be the source for much of the measured unbound IGF-I.

Various methods have been used to estimate the unbound (or freely dissociated) IGF-I fraction.4,9,10 Size-exclusion chromatography and filtration methods.4,9 have the theoretical disadvantage of altering the sample matrix and the equilibrium between IGF-I and IGFBP’s. A direct detection unbound IGF-I assay using immobilized IGFBP-3 for capture and anti-IGF-I antibody for detection has been reported.11

The Ansh Labs Free IGF-I kit uses a highly sensitive two-site antibody method which allows detection of unbound IGF-I. The kit may be used as a “direct” assay to measure the “dissociable” fraction of IGF-I.8

References:
1. Daughaday E, Rotwein P: Insulin like growth factors I and II. Peptide, messenger ribonucleic acid and gene structures, serum and tissue concentrations. Endocrin Rev 10:68-91, 1989.
2. Baxter RC, Martin JL, Beniac VA: High molecular weight insulin-like growth factor binding protein complex. J Biol Chem 264:11843-11848, 1989.
3. Rechler M: Insulin-like growth factor binding proteins. Vit Horm 47:1-114, 1993.
4. Guler HP, Zapf J, Froesch ER: Short-term metabolic effects of recombinant human insulin-like growth factor-I in healthy adults. New Engl J Med 317:137-140, 1987.
5. Zapf J, Hauri C, Waldvogel M, Froesch ER: Acute metabolic effects and half-lives of intravenously administered insulinlike growth factors I and II in normal and hypophysectomized rats. J Clin Invest 77:1768-1775, 1986.
6. Costigan DC, Guyda HJ, Posner BI: Free insulin-like growth factor I (IGF-I) and IGF-II in human saliva. J Clin Endocrinol Metab 66:1014-1018, 1988.
7. Lewitt MS, Denyer GS, Cooney GJ, Baxter RC: Insulin-like growth factor-binding protein-1 modulates blood glucose levels. Endocrinology 129:2254-2256, 1991.
8. Lewitt MS, Saunders H, Baxter RC: Bioavailability of insulin-like growth factors (IGFs) in rats determined by the molecular distribution of human IGF-binding protein-3. Endocrinology 133:1797-1802, 1993.
9. Lieberman SA, Bukar J, Chen SA, Celniker AC, Compton PG, Cook J, Albu J, Perlman AJ, Hoffman AR: Effects of recombinant human insulin-like growth factor-I (rhIGF-I) on total and free IGF-I concentrations, IGF-binding proteins, and glycemic response in humans. J Clin Endocrinol Metab 75:30-36, 1992.
10. Lee PDK, Powell D, Baker B, Liu F, Mathew G, Levitsky I, Gutierrez OD, Hintz RL: Characterization of a direct, non-extraction immunoradiometric assay for free IGF-I. Presented at the 76th annual meeting of the Endocrine Society, Anaheim, 1994.
11. Mukku VR: A 96-well microtiter plate assay for free IGF-I in plasma using immobilized IGFBP-3. Presented at the 73rd annual meeting of The Endocrine Society, Washington, D.C., 1991. (abstract #462).
12. HHS Publication, 5th ed., 2007. Biosafety in Microbiological and Biomedical Laboratories. Available http://www.cdc.gov/biosafety/publications/bmbl5/BMBL5
13. DHHS (NIOSH) Publication No. 78–127, August 1976. Current Intelligence Bulletin 13 – Explosive Azide Hazard. Available http:// www.cdc.gov/niosh.
14. Kricka L. Interferences in immunoassays – still a threat. Clin Chem 2000; 46: 1037–1038.

Anastasilakis AD, Koulaxis D, Upadhyay J, Pagkalidou E, Kefala N, Perakakis N, Polyzos SA, Economou F, Mantzoros CS. Free IGF-1, Intact IGFBP-4, and PicoPAPP-A are Altered in Acute Myocardial Infarction Compared to Stable Coronary Artery Disease and Healthy Controls. Horm Metab Res. 2019 Feb;51(2):112-119.

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Becker M, Haluska P, Bale L, Oxvig C, Conover C. A Novel Neutralizing Antibody Targeting Pregnancy-Associated Plasma Protein-A Inhibits Ovarian Cancer Growth and Ascites Accumulation in Patient Mouse Tumorgrafts. Mol Cancer Ther. 2015 Apr;14(4):973-81. Epub 2015 Feb 18.

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Becker M, Haluska P, Bale L, Oxvig C, Conover C. A Novel Neutralizing Antibody Targeting Pregnancy-Associated Plasma Protein-A Inhibits Ovarian Cancer Growth and Ascites Accumulation in Patient Mouse Tumorgrafts. Mol Cancer Ther. 2015 Apr;14(4):973-81. Epub 2015 Feb 18.

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Cabrera-Salcedo C, Mizuno T, Tyzinski L, Andrew M, Vinks AA, Frystyk J, Wasserman H, Gordon CM, Hwa V, Backeljauw P, Dauber A. Pharmacokinetics of IGF-1 in PAPP-A2-Deficient Patients, Growth Response, and Effects on Glucose and Bone Density. J Clin Endocrinol Metab. 2017 Dec 1;102(12):4568-4577.

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Damen, L.; Elizabeth, M.S.M.; Donze, S.H.; van den Berg, S.A.A.; de Graaff, L.C.G.; Hokken-Koelega, A.C.S. Free Insulin-like Growth Factor (IGF)-I in Children with PWS. J. Clin. Med. 2022, 11, 1280. https://doi.org/10.3390/jcm11051280

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Dauber A, Munoz-Calvo MT, Barrios V, Domene HM, Kloverpris S, Serra-Juhe C, Desikan V, Pozo J, Muzumdar R, Martos-Moreno GA, Hawkins F, Jasper HG, Conover CA, Frystyk J, Yakar S, Hwa V, Chowen JA, Oxvig C, Rosenfeld RG, Perez-Jurado LA, Argente J. Mutations in pregnancy-associated plasma protein A2 cause short stature due to low IGF-I availability. EMBO Mol Med. 2016 Mar 31;8(4):363-74.

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Fujimoto M, Khoury JC, Khoury PR, Kalra B, Kumar A, Sluss P, Oxvig C, Hwa V, Dauber A. Anthropometric and biochemical correlates of PAPP-A2, free IGF-I, and IGFBP-3 in childhood. European journal of endocrinology, 182(3), 363–374. https://doi.org/10.1530/EJE-19-0859

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Guerra-Cantera S, Frago LM, Díaz F, Ros P, Jiménez-Hernaiz M, Freire-Regatillo A, Barrios V, Argente J, Chowen JA. Short-Term Diet Induced Changes in the Central and Circulating IGF Systems Are Sex Specific. Front Endocrinol (Lausanne). 2020 Aug 11;11:513. doi: 10.3389/fendo.2020.00513. PMID: 32849298; PMCID: PMC7431666.

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Heitzeneder S, Shem J, Khan J, Mackall C. Pregnancy Associated Plasma Protein A (PAPP-A) Is A Novel Therapeutic Target In Ewing Sarcoma. Presented at 8th Ovarian Cancer Research; 2016 May 15-17; Niagra Falls, ON, Canada.

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Kumar A, Kalra B, Chowdavarapu K, Shah S, Savjani G, Oxvig C. Development of IGF-1 ELISA Assays to Measure Free and Total Circulating IGF-1. Poster session presented at the 96th Annual Endocrine Society Meeting; 2014 Jun 21-24; Chicago, IL.

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Kumar A, Kalra B, Chowdavarapu K, Shah S, Savjani G, Oxvig C. Development of IGF-1 ELISA Assays to Measure Free and Total Circulating IGF-1. Presented at 98th Annual Endocrine Society Meeting; 2016 Apr 1-3; Boston, MA

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Kumar A, Kalra B, Kommareddy V, Chowdavarapu K, Mistry S, Savjani G, Oxvig C. Development of Well Characterized ELISAs for Bound and Unbound Insulin-Like Growth Factors and their Binding Proteins. Poster presented at 98th Annual Endocrine Society Meeting; 2016 Apr 1-3; Boston, MA.

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Martín-Rivada Á, Guerra Cantera S, Campillo-Calatayud A, Andrés-Esteban EM, Sánchez Holgado M, Martos-Moreno GÁ, Pozo J, Güemes M, Soriano-Guillén L, Pellicer A, Oxvig C, Frystyk J, Chowen JA, Barrios V, Argente J. Pappalysins and Stanniocalcin and Their Relationship With the Peripheral IGF Axis in Newborns and During Development. J Clin Endocrinol Metab. 2022 Sep 28;107(10):2912-2924. doi: 10.1210/clinem/dgac453. PMID: 35902207.

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Panagiotou G, Ghaly W, Upadhyay J, Pazaitou-Panayiotou K, Mantzoros CS. Serum Follistatin Is Increased in Thyroid Cancer and Is Associated With Adverse Tumor Characteristics in Humans. J Clin Endocrinol Metab. 2021 Apr 23;106(5):e2137-e2150. doi: 10.1210/clinem/dgab041. PMID: 33493282.

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Pellikaan K, Elizabeth M, Donze S, van den Berg S, van Doorn J, Peeters RP, Hokken-Koelega A, Graaff L. Normal IGF-bioactivity and low free IGF-I in patients with Prader-Willi syndrome with high total serum IGF-I: immunoreactive IGF-I concentrations poorly reflect IGF bio-activity and bio-availability. Endocrine Abstracts (2019) 63 GP175 | DOI: 10.1530/endoabs.63.GP175

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Valenzuela-Vallejo L, Chrysafi P, Agraz M, Connelly M, Papatheodoridis G, Verrastro O, Mingrone G, Mantzoros CS. GROWTH DIFFERENTIATION FACTOR 15 (GDF-15) LEVELS IS ASSOCIATED WITH THE PRESENCE AND SEVERITY OF NAFLD. Nutr Diabetes. 2022 Feb 22;12(1):9. doi: 10.1038/s41387-022-00187-2. PMID: 35194014; PMCID: PMC8863897

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