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Myostatin inhibitors in sports drug testing: Detection of myostatin‐neutralizing antibodies in plasma/serum by affinity purification and Western blotting
Author(s) -
Walpurgis Katja,
Thomas Andreas,
Schänzer Wilhelm,
Thevis Mario
Publication year - 2016
Publication title -
proteomics – clinical applications
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.948
H-Index - 54
eISSN - 1862-8354
pISSN - 1862-8346
DOI - 10.1002/prca.201500043
Subject(s) - myostatin , antibody , blot , affinity chromatography , microbiology and biotechnology , biology , chemistry , skeletal muscle , immunology , biochemistry , endocrinology , enzyme , gene
Purpose Myostatin is a key regulator of skeletal muscle growth and inhibition of its signaling pathway results in an increased muscle mass and function. The aim of this study was to develop a qualitative detection assay for myostatin‐neutralizing antibodies for doping control purposes by using immunological approaches. Experimental design To detect different types of myostatin‐neutralizing antibodies irrespective of their amino acid sequence, an immunological assay specific for antibodies directed against myostatin and having a human Fc domain was established. Affinity purification and Western blotting strategies were combined to allow extracting and identifying relevant analytes from 200 μL of plasma/serum in a non‐targeted approach. The assay was characterized regarding specificity, linearity, precision, robustness, and recovery. Results The assay was found to be highly specific, robust, and linear from 0.1 to 1 μg/mL. The precision was successfully specified at three different concentrations and the recovery of the affinity purification was 58%. Conclusions and clinical relevance Within this study, an immunological detection assay for myostatin‐neutralizing antibodies present in plasma/serum specimens was developed and successfully characterized. The presented approach can easily be modified to include other therapeutic antibodies and serves as proof‐of‐concept for the detection of antibody‐based myostatin inhibitors in doping control samples.