Recent developments in doping testing for erythropoietin.

The constant development of new erythropoiesis-stimulating agents (ESAs), since the first introduction of recombinant erythropoietin (rhEpo) for clinical use, has also necessitated constant development of methods for detecting the abuse of these substances. Doping with ESAs is prohibited according to the World Anti-Doping Code and its prohibited list of substances and methods. Since the first publication of a direct and urine-based detection method in 2000, which uses changes in the Epo isoform profile as detected by isoelectric focusing in polyacrylamide slab gels (IEF-PAGE), the method has been constantly adapted to the appearance of new ESAs (e.g., Dynepo, Mircera). Blood had to be introduced as an additional matrix, because Mircera (a PEGylated Epo) is best confirmed in serum or plasma after immunoaffinity purification. A Mircera ELISA was developed for fast screening of sera. With the appearance of Dynepo and copy epoetins, the additional application of sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE or equivalent) became necessary. The haematological module of the Athlete Biological Passport is the latest development in multivariable indirect testing for ESA doping. The article summarizes the main strategies currently used in Epo anti-doping testing with special focus on new developments made between 2009 and 2010.