However, chromatography and mass spectrometry require specialized technical skill along with dedicated and often expensive instrumentation. LC/MS/MS provides a high degree of specificity for steroid hormone measurement. An increasing number of clinical laboratories, particularly reference laboratories, utilize liquid chromatography-tandem mass spectrometry (LC/MS/MS) for steroid hormone measurement. The most common alternative approach for measurement of steroid hormones is chromatography (especially high-performance liquid chromatography, HPLC), either alone or in combination with mass spectrometry (MS) (HPLC/MS or simply LC/MS). A variety of immunoassay methods are available, ranging from enzyme-linked immunosorbent assays (ELISAs) to homogeneous immunoassays that can be run on high-throughput analyzers commonly found in hospital clinical laboratories. Immunoassays are frequently used in laboratory medicine for quantitation of plasma/serum concentrations of steroid hormones such as cortisol, dehydroepiandrosterone (DHEA) sulfate, estradiol, progesterone, or testosterone. Simple similarity calculations can help triage compounds for future testing of assay cross-reactivity. Clinically significant interactions can occur with structurally similar drugs (e.g., prednisolone and cortisol immunoassays methyltestosterone and testosterone immunoassays) or with endogenous compounds such as 21-deoxycortisol that can accumulate to very high concentrations in certain disease conditions. ConclusionsĬompounds producing cross-reactivity in steroid hormone immunoassays generally have a high degree of structural similarity to the target hormone. Using two-dimensional similarity calculations, all compounds with high cross-reactivity also showed a high degree of similarity to the target molecule of the immunoassay. Norethindrone therapy may impact immunoassay measurement of testosterone in women.

Several anabolic steroids may produce clinically significant false positives on the testosterone assay, although interpretation is limited by sparse pharmacokinetic data for some of these drugs. In addition, 21-deoxycortisol likely produces clinically relevant cross-reactivity for cortisol in patients with 21-hydroxylase deficiency, while 11-deoxycortisol may produce clinically relevant cross-reactivity in 11β-hydroxylase deficiency or following metyrapone challenge. 6-Methylprednisolone and prednisolone showed high cross-reactivity for the cortisol assay, with high likelihood of clinically significant effect for patients administered these drugs. The Roche Elecsys Cortisol and Testosterone II assays showed a wider range of cross-reactivity than the DHEA sulfate, Estradiol II, and Progesterone II assays. Cross-reactivity was computationally predicted using the technique of two-dimensional molecular similarity. These data were compared and contrasted to package insert data and published cross-reactivity studies for other marketed steroid hormone immunoassays. MethodsĬross-reactivity of a structurally diverse set of compounds were determined for the Roche Diagnostics Elecsys assays for cortisol, dehydroepiandrosterone (DHEA) sulfate, estradiol, progesterone, and testosterone. Interfering molecules include structurally related endogenous compounds and their metabolites as well as drugs such as anabolic steroids and synthetic glucocorticoids.

One limitation of steroid hormone immunoassays is interference caused by compounds with structural similarity to the target steroid of the assay. Immunoassays can be performed on a variety of standard clinical chemistry analyzers, thus allowing even small clinical laboratories to do analysis on-site. Immunoassays are widely used in clinical laboratories for measurement of plasma/serum concentrations of steroid hormones such as cortisol and testosterone.

Antibody reactivity and specificity