Evaluating the Testosterone‐to‐Luteinizing Hormone Ratio in Male Anti‐Doping Serum Samples: Results From a Transdermal Testosterone Administration Trial and Field Collected Samples

The serum testosterone‐to‐luteinizing hormone (T/LH) ratio has previously been suggested as a sensitive marker of testosterone use in an anti‐doping setting. When measured with an automated immunoassay platform, this ratio represents a quick and cost‐effective screening biomarker to further direct isotope ratio mass spectrometry (IRMS) testing efforts in concurrently collected urine samples to confirm testosterone abuse. To evaluate the practicality of implementing the serum T/LH ratio for routine use, testosterone values in 356 serum samples were compared between a “gold‐standard” LC–MS/MS method and an automated immunoassay method. Excellent correlation and minimal bias between the two methods were observed, highlighting the validity of the immunoassay method. Next, a testosterone administration study utilizing a transdermal delivery route was conducted to compare the effectiveness of the serum T/LH ratio to the currently used serum testosterone to androstenedione (T/A4) and urinary testosterone to epitestosterone (T/E) ratios. The serum T/LH ratio was more sensitive than the T/A4 ratio and showed similar sensitivity to the urinary T/E ratio with high interindividual variability. Finally, T/LH analysis was conducted on 626 capillary serum samples collected in the field from male Ultimate Fighting Championship athletes. Of the three samples that showed elevated T/LH ratios in this pool, two of the corresponding urine samples were IRMS positive, one of which showed an unremarkable urinary T/E ratio and relatively normal steroid profile. These results indicate serum T/LH ratio monitoring provides a beneficial addition to the anti‐doping tool kit and can improve urinary IRMS testing recommendations.