VA Medical Center, Gainesville, FL 32608, United States; Department of Applied Physiology & Kinesiology, University of Florida, Gainesville, FL 32611, United States.
Steroids. 2013 Dec 11;78(12-13):1220-5. doi: 10.1016/j.steroids.2013.08.013. Epub 2013 Sep 6.
Enzyme immunoassays (EIA) are commonly utilized for the evaluation of androgens in biological fluids; however, careful consideration must be given to cross-reactivity with other endogenous sex-steroid hormones. Our purpose was to determine the validity of a commonly-utilized commercially-available dihydrotestosterone (DHT) EIA. Serum samples obtained from older hypogonadal men who participated in a 12-month randomized controlled trial evaluating the effects of testosterone-enanthate (125 mg/week) or vehicle in combination with finasteride (5mg/day) or placebo were assayed for DHT via EIA and using a validated gold-standard LC-MS/MS approach. Additionally, commercially-available (DHT-free) buffer containing graded testosterone doses was evaluated by DHT immunoassay. DHT concentrations measured via EIA were 79% to >1000% higher than values obtained by LC-MS/MS (p<0.05), with the largest differences (415-1128%) occuring in groups receiving finasteride. Both LC-MS/MS and EIA indicated that testosterone-enanthate increased serum DHT to a similar magnitude. In contrast, finasteride-induced reductions in DHT were detected by LC-MS/MS, but not EIA (p<0.05). No significant associations were present for DHT concentrations between measurement techniques. Cross-reactivity of testosterone with the immunoassay ranged from 18% to 99% and DHT concentrations measured by EIA were highly associated with the spiked testosterone concentrations in DHT-free buffer (r=0.885, p<0.001). In conclusion, we provide evidence invalidating a commonly-utilized commercially-available DHT immunoassay because significant cross-reactivity exists between testosterone and the EIA and because the changes in DHT observed via EIA were not associated with a validated gold-standard measurement technique. The cross-reactivity of testosterone is particularly concerning because testsoterone is present in 100-fold greater concentrations than is DHT within the circulation.
酶免疫分析（EIA）常用于评估生物体液中的雄激素；然而，必须仔细考虑与其他内源性性激素的交叉反应。我们的目的是确定一种常用的商业上可利用的二氢睾酮（DHT）EIA 的有效性。从参加为期 12 个月的随机对照试验的老年性腺功能减退男性的血清样本中，该试验评估了睾酮-庚酸（125mg/周）或载体与非那雄胺（5mg/天）或安慰剂联合治疗的效果，通过 EIA 和经过验证的金标准 LC-MS/MS 方法测定 DHT。此外，还通过 DHT 免疫分析评估了含有分级睾酮剂量的市售（无 DHT）缓冲液。通过 EIA 测量的 DHT 浓度比通过 LC-MS/MS 获得的值高 79%至>1000%（p<0.05），在接受非那雄胺的组中差异最大（415-1128%）。LC-MS/MS 和 EIA 均表明，睾酮-庚酸使血清 DHT 增加到相似的程度。相比之下，非那雄胺诱导的 DHT 减少仅通过 LC-MS/MS 检测到，而通过 EIA 未检测到（p<0.05）。两种测量技术之间的 DHT 浓度没有显著关联。睾酮与免疫测定的交叉反应率为 18%至 99%，EIA 测量的 DHT 浓度与无 DHT 缓冲液中添加的睾酮浓度高度相关（r=0.885，p<0.001）。总之，我们提供了证据，证明一种常用的商业上可利用的 DHT EIA 是无效的，因为睾酮与 EIA 之间存在显著的交叉反应，并且通过 EIA 观察到的 DHT 变化与经过验证的金标准测量技术无关。睾酮的交叉反应性特别令人担忧，因为在循环中，睾酮的浓度比 DHT 高 100 倍。
