Quantification of human insulin-like growth factor-1 and qualitative detection of its analogues in plasma using liquid chromatography/electrospray ionisation tandem mass spectrometry.
Human insulin-like growth factor-1 (IGF-1) is a peptide hormone that acts as a mediator of most of the somatotropic effects of growth hormone (GH). Therefore, it is supposed to be a biomarker indicating GH abuse in sports as well as diseases associated with a change in IGF-1 plasma concentration. It can be applied locally by injection to increase total protein and DNA content in tissues such as skeletal muscle--a highly desirable effect in various sports disciplines. In order to improve its growth-promoting properties, the primary structure of IGF-1 has been modified, yielding analogues such as des(1-3)IGF-1 and LONGR3IGF-1, which show a considerably reduced affinity to the respective binding proteins in plasma and, thus, an increased bioavailability at target tissues. Due to their capability to enhance performance, IGF-1 and its analogues belong to the prohibited list of the World Anti-Doping Agency. Hence, it was necessary to develop a reliable assay for the quantification of human IGF-1 as well as the detection of its derivatives. Immunoaffinity isolation and purification from 60 microL of plasma followed by liquid chromatography/electrospray ionisation tandem mass spectrometry enabled the unequivocal determination of all target analytes. Diagnostic product ions were characterised utilising an Orbitrap mass spectrometer with high resolution/high accuracy properties and employed for triple quadrupole MS/MS analysis. The described assay provided lower limits of detection (LLODs) between 20 and 50 ng/mL, recovery rates between 34-43% and a precision <15% at the LLOD as well as higher concentration levels. In order to prove the applicability of the developed assay, human plasma samples were analysed and the results were compared with the values obtained from a commercially available immunoradiometric assay (IRMA). Four of six samples resulted in concentration ratios with good correlation between both assays, whereas the absolute concentrations were lower for the presented procedure.
Bredehöft
Michael
M
Center for Preventive Doping Research - Institute of Biochemistry, German Sport University Cologne, Carl-Diem-Weg 6, 50933 Cologne, Germany. m.bredehoeft@biochem.dshs-koeln.de
Schänzer
Wilhelm
W
Thevis
Mario
M
D001774:Blood Chemical Analysis / Q000379:methods*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D004300:Doping in Sports / Q000517:prevention & control*; D013006:Growth Hormone / Q000097:blood*; D007334:Insulin-Like Growth Factor I / Q000032:analysis*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D015813:Substance Abuse Detection / Q000379:methods*
D007334:Insulin-Like Growth Factor I; D013006:Growth Hormone