Wolken Jill K, Peterson Meghan M, Cao Wenjing, Challoner Keith, Jin Zhicheng
University of Wisconsin Hospital and Clinics, Madison, WI 53792, USA.
Department of Pathology and Laboratory Medicine, University of Wisconsin-Madison, 600 Highland Avenue, Madison, WI 53792, USA.
J Clin Med. 2024 Nov 22;13(23):7056. doi: 10.3390/jcm13237056.
: Testosterone is an androgenic hormone that plays important roles in both males and females. The circulating levels of total testosterone vary from 1 to 1480 ng/dL. High-throughput immunoassays often lack accuracy in lower concentration ranges (below 100 ng/dL), particularly when used for females or children. To address this limitation, we developed a total testosterone LC-MS/MS assay on three instruments. : Sample preparation began with the dilution and conditioning of 200 µL of serum. A supported liquid extraction cartridge was used to extract the analyte from biological matrices. Chromatographic separation was achieved using a C18 column with a runtime of 5 min per sample. This assay was validated on a Triple Quad 6500 and an API 4500 instrument. : Method validation was carried out according to the CLSI C62-ED2 guideline and our hospital protocol. The within-day coefficient of variation (CV) was less than 10% and the between-day CV was less than 15%. The assay had a limit of quantitation of 0.5 ng/dL with an analyte measure range of 2-1200 ng/dL. A comparison using Deming regression and Bland-Altman plots showed that this assay correlated well with a reference method. The results from the API 4500 and an Orbitrap were consistent with those from the TQ 6500. Both serum-separator tubes (BD) and serum-activator tubes were found to be suitable. : We successfully developed and validated a robust total testosterone LC-MS/MS assay for routine clinical testing. This assay was harmonized across two triple quadrupole instruments and one high-resolution mass spectrometer.
睾酮是一种雄激素，在男性和女性体内都发挥着重要作用。总睾酮的循环水平在1至1480 ng/dL之间变化。高通量免疫测定法在较低浓度范围（低于100 ng/dL）时往往缺乏准确性，尤其用于女性或儿童时。为解决这一局限性，我们在三种仪器上开发了总睾酮液相色谱-串联质谱测定法。：样品制备始于200 μL血清的稀释和预处理。使用固相萃取柱从生物基质中提取分析物。使用C18柱实现色谱分离，每个样品的运行时间为5分钟。该测定法在三重四极杆6500和API 4500仪器上进行了验证。：方法验证按照CLSI C62-ED2指南和我院方案进行。日内变异系数（CV）小于10%，日间CV小于15%。该测定法的定量限为0.5 ng/dL，分析物测量范围为2至1200 ng/dL。使用Deming回归和Bland-Altman图进行的比较表明，该测定法与参考方法相关性良好。API 4500和轨道阱的结果与TQ 6500的结果一致。发现血清分离管（BD）和血清激活管均适用。：我们成功开发并验证了一种用于常规临床检测的可靠的总睾酮液相色谱-串联质谱测定法。该测定法在两台三重四极杆仪器和一台高分辨率质谱仪上实现了统一。
