Development of a rapid and sensitive detection of hGH-dependent serum markers based on fluorescence

Code: 12B11GK

Recombinant human growth hormone (rhGH) has been on the list of forbidden substances since availability of its recombinant form in the early 1990s; however adequate routine doping tests are lacking. The project aims to develop a fast and highly sensitive drug test for detecting two or more hGH-dependent markers in the serum of elite adolescent athletes. In our approach proteomic markers for hGH action such as insulin-like growth factor 1 (IGF-1) and pro-collagen type III N-terminal peptide (P-III-P) will be identified in just a single immunoassay. Considering the desirable reduction of time, costs and workload using FCS instead of other currently available IGF-1 and P-III-NP assays the presented methodology will be an important contribution for a functional doping test for proper use of rhGH. 

Main Findings: 

Two approaches have been developed to detect recombinant hGH in blood in order to control its misuse with the intention of improving athletic performance. Adequate non-invasive tests for human growth hormone (hGH)-dependent markers such as insulin-like growth factor 1 (IGF-1) and pro-collagen type Ill N-terminal peptide (PIIINP) are still lacking. In this one-year project a fast and sensitive bead-based immunoassay for IGF-1 and PIIINP detection in serum was developed based on Fluorescence Correlation Spectroscopy (FCS). FCS was used as a reliable technology for measuring absolute concentrations in the nano-molar range. Three FCS immunoassays - a sandwich and a competitive immunoassay for IGF-1 as well as a competitive PIIINP assay - were established and validated against commercially available ELISA. We were able to detect molarities between 0.5 and lOnM of IGF-1 and between 0.5 and 2.5nM of PIIINP with high accuracy in serum samples.  
Considering the desirable reduction of time, costs and workload using FCS instead of other currently available IGF-1 and PIIINP assays the presented methodology might be an important contribution for proper use of rhGH in sports.