Tailor-Made Immunochromatographic Test for the Detection of Multiple 17α-Methylated Anabolics in Dietary Supplements

Abstract

In recent years, the undeclared presence of various anabolic androgenic steroids (AAS) in commercial supplements has been confirmed. This fact can be a potential threat to all athletes using these supplements, and therefore, there is of increased interest in the implementation of rapid methods for the detection of AAS. The presented study describes the development of an immunostrip test for the detection of multiple 17α-methylated AAS based on direct and indirect competitive principle using gold nanoparticles as a label. As a capture reagent on test lines conjugated stanazolol to rabbit serum albumin (RSA/ST-3) was used, the intensity of color formed in the test line of the AAS-positive sample was visually distinguishable from that of negative sample within 10 min. The optimized closed direct and indirect format of the test provided a similar visual detection limit (0.7 and 0.9 ng/mL, respectively). The most commonly orally abused AAS (17α-methyltestosterone, methandienone, methyldihydrotestosterone, oxandrolone and oxymetholone) showed a strong cross-reaction. Developed immunostrips were successfully applied to analysis of artificially contaminated dietary supplements with 17α-methylated AASs. The developed immunostrips offer potential as a useful user-friendly method for capturing suspicious dietary supplement samples with different contents of AAS at levels far below the usually used concentrations of AAS.

Advances in the Determination of Anabolic-Androgenic Steroids: From Standard Practices to Tailor-Designed Multidisciplinary Approaches

“…The presented LFIAs in Table 3 differ in the analyte of interest and the origin of the antibodies used; however, they all use gold labeling. The lowest achieved LOD for AASs that can be detected by a naked eye is 0.7 ng per mL in the case of 17α-methylated AASs such as stanozolol [ 41 ]. Compared to the currently developed ELISAs, LFIAs are generally less sensitive; on the other hand, for example, an ethanol extract of food supplements can be diluted to a lower extent for LFIA than for ELISA [ 89 ].…”

“…A stark contrast to the intentional misuse of AASs is their undeclared occurrence in dietary supplements (DSs) [ 33 , 34 , 35 ], which is a topic we have dealt with for a long time at the University of Chemistry and Technology, Prague [ 36 , 37 , 38 , 39 , 40 , 41 ]. Despite the apparent threat and criminal nature of undeclared enrichment of DSs with AASs, this phenomenon has been detected worldwide from time to time during research [ 42 ], random inspections of overseeing authorities [ 43 , 44 ], or as a result of revealing the cause of health problems [ 45 ].…”

“…17α-Methylated AASs Gold-labeled rabbit polyclonal 0.7 ng•mL−1 Dietary supplements[41] Dehydroepiandrosterone LOD = 500 µg•kg −1Slimming products (herbal teas, capsules, pills)[88] …”