Abstract

Recombinant erythropoietin (rhEPO) has been misused for over two decades by athletes, mainly but not only in endurance sports. A direct rhEPO detection method in urine by isoelectric focusing (IEF) was introduced in 2000, but the emergence of “third generation” of erythropoiesis-stimulating agents and so-called “biosimilars” rhEPOs, together with the sensitivity of human endogenous EPO (huEPO) pattern to enzymatic activities and its modification following short strenuous exercise, prompted the development of a complementary test based on SDS-PAGE analysis. While Mircera and NESP can be detected with the existing IEF and SDS-PAGE methods, some samples containing both epoetin-α/β and huEPO present profiles that may still be difficult to interpret. As doping practices having moved to microdosing, these mixed patterns are more frequently observed. We have investigated the impact of enzymatic desialylation on the urinary and serum EPO profiles obtained by SDS-PAGE with the aim of improving the separation of the bands in these mixed EPO populations. We have observed that the removal with neuraminidase of the sialic acid moieties from the different EPOs studied reduced their apparent molecular weight (MW) and increased the migration distance between huEPO and rhEPO centroids, therefore eliminating the size overlaps between them and improving the detection of rhEPO. Verf.-Referat