Since its discovery over 50 years ago, the biological aspects of insulin-like growth factor-1 (IGF-1) have been determined to be extensive. Ranging from cell proliferation, cell differentiation and apoptosis, tissue growth, and organ-specific functions, the biological functions of IGF-1 and its measurement have been deemed significant. As a biomarker, it is used in the diagnosis and treatment of growth disorders, has been implicated in the prognosis and diagnosis of several cancers, and is used to aid in the identification of athletic enhancement from growth hormone use. Its therapeutic forms have been implicated in athletic doping as well.
On account of the significance of its measurement, immunoassays have been developed over the years in attempts to accurately and precisely measure IGF-1. Since the first IGF-1 radioimmunoassay in 1977, many methods for detecting IGF-1 have been introduced, including radioimmunoassay, ELISA, immuno-chemoluminescence, and mass spectrometry-based assays. However, these described methods are not without insufficiencies that result from the complexities in IGF-1 as a peptide.
Measurement of IGF-1 is complicated, by both the biology of IGF-1 and the recent need to differentiate the endogenous form of IGF-1, and the synthetic therapeutic forms developed to treat growth disorders.
IGF-1 is a small peptide that circulates in plasma with the majority (98%) bound to the IGF-binding proteins (IGFBP). There are six binding proteins, each with a different affinity for IGF-1. Therefore, direct measurement requires that the IGF-1/IGFBP complex be disrupted. Due to the differences in the native IGF-1/IGFBP complexes, extensive efforts have been focused on their disruption.
Over the years, methods have been refined into the most commonly used protocol today, which uses an acid/ethanol precipitation (to disrupt the IGF-1/IGFBP complex) followed by the addition of insulin-like growth factor-2 (IGF-2) to prevent the reformation of the IGF/IGFBP complex. However, this liberation methodology is not without faults.
First, in the process of disrupting the IGF-1/IGFBP complex by acid/ethanol precipitation, there is the potential IGF-1 loss, which would directly affect the measurement’s accuracy. Secondly, the post-disruption addition of IGF-2 is not a 100% prophylactic treatment in preventing IGF-1/IGFBP re-complexation. In fact, this method has little effect on preventing the reformation of the IGF-1/IGFBP complex with the smaller IGFBPs (IGFBP-1 and IGFBP-4). Hence, these contribute to the reasons for the current inconsistencies in IGF-1 measurements.
In addition to having to overcome the IGF-1/IGFBP complex issues, the IGF-1 analytical techniques have recently had to contend with the existence of therapeutic forms of IGF-1. As a result of these synthetic forms, immunoassays that are able to differentiate endogenous IGF-1 from the therapeutic forms have become highly sought after.
For conventional immunoassays, this is costly in both money and time. As these approaches require antibodies to be developed that enable this degree of differentiation. However, for mass spectrometric immunoassays (MSIA), IGF-1 is detected by the measurement of its intrinsic molecular mass and, therefore, alternate forms of IGF-1 that differ in mass can be readily differentiated.
One such MSIA, termed IGF-1 MSIA-SRM, has been developed for IGF-1 with improved protocols for disrupting the IGF-1/IGFBP complex and for monitoring the immunoassays’ efficiency from sample preparation to mass spectrometric detection.