Detection capabilities of a single dried blood spot for recombinant erythropoietin micro-dose testing in sport

Abstract

Objectives: Dried blood spot (DBS) micro-sampling is gaining prominence in anti-doping testing, particularly for the detection of erythropoietin-receptor agonists (ERAs) such as recombinant erythropoietin (rEPO). However, detecting micro-doses of rEPO remains challenging due to the low concentrations in blood. This study explores the detection capabilities of micro-doses of rEPO using a single collection pad of a DBS device. Design and methods: A single subcutaneous micro-dose of rEPO (Procrit®, 15 IU/kg) was administered to four healthy participants. DBS samples (4 × 17.5 μL) were collected using the polymeric Tasso-M20® device at multiple time points post-administration, and compared with 15 mL urine samples. All samples were analyzed using SAR-PAGE and Western-Blot techniques. In line with the new WADA regulations, two MAIIA immunopurification methods with different capture antibodies were validated for the initial testing and confirmation of rEPO in DBS. Results: All ERAs were detectable in polymeric DBS matrix at the limits of detection required by WADA. In polymeric DBS, analytes remained stable for at least six months at room temperature and at −20 °C. rEPO micro-dose was detectable in DBS up to 72–96 h post-dose, and in urine up to 84–96 h. Conclusions: These findings support the suitability of DBS as a complementary matrix to urine for the detection of rEPO, reinforcing confidence in its effectiveness for the detection of ERAs.