Short communicationCross reactions elicited by serum 17-OH progesterone and 11-desoxycortisol in cortisol assays
Abstract
Background
Different pathophysiological situations such as congenital adrenal hyperplasia, adrenocortical carcinoma, metyrapone treatment, etc. elicit specificity problems with serum cortisol assay.
Methods
We assayed cortisol using 2 kits and performed cross reaction studies as well as multiple regression analysis using 2 other steroids: 11-desoxycortisol and 17-OH progesterone.
Results
Analysis showed the existence of an analytical bias. Importantly, significantly different biases were demonstrated in newborns or patients taking metyrapone. Multiple regression analysis and cross reaction studies showed that 11-desoxycortisol level significantly influenced cortisol determination. Moreover, despite using the normal ranges provided by manufacturers discrepant results occurred such as 17% discordance in the diagnosis of hypocorticism in infants.
Conclusion
We wish to raise awareness about the consequences of the (lack of) specificity of cortisol assays with regard to the evaluation of hypocorticism in infants or when “unusual” steroids may be increased.
Introduction
Cross reaction problems in steroids immunoassay are well described as these very similar molecules are assayed by competitive techniques that potentially overestimate true concentrations [1], [2], [3]. Overestimating cortisol concentrations can have serious consequences as it leads to under diagnosing hypocortisolism. Mass spectrometry provides gold standards but its common use is limited to standardize and compare kits [4], [5]. The routine use of these immunoassay commercial kits usually rests upon reference ranges obtained in healthy adult (usually obtained in the morning). In such a physiological condition, cortisol concentrations are clearly above those of potentially interfering steroids thus minimizing the impact of cross reactions. Hence, a good agreement is obtained between different kits in these normal conditions.
Obviously, in a university hospital, the situations requesting cortisol assay depart from physiology. Indeed, in some pathological situations, the concentration of steroids closely related to cortisol dramatically increases (e.g. cortisol precursors in patients with congenital adrenal hyperplasia (CAH) [6] or with adrenocortical carcinoma [7] or under metyrapone treatment or undergoing a metyrapone test, or in premature infants [8]). Manufacturers usually provide some data about the cross reactivity of their assays although there is no mandatory list of compounds to be tested.
We investigated 2 cortisol assay kits to assess the clinical impact of possible differences of cross reactivity in real-life conditions in a university hospital, in which some physiological or pathological situations require the assessment of cortisol concentrations.
Section snippets
Materials
Cortisol concentration was assayed by Gamma Coat CA 1529, kit A (Clinical Assays, Stillwater, USA) and by Spectria cortisol RIA, kit B (Orion Diagnostic, Espoo, Finland). Inter-assay coefficients of variation (CV) were: kit A 7.7% and 3.5% (81 and 571 nmol/L) and kit B 7.4% and 9.7% (73.5 and 446 nmol/L). Normal ranges provided by the manufacturers were: [200–700] and [131–642] nmol/L for kit A and B, respectively. Both kit leaflets contained information about cross reactions: for 17-OH
Results
Two RIA kits were used to evaluate the degree of agreement between cortisol concentrations: concentrations obtained with the kit A were higher than those obtained with the kit B (n = 114, p < 0.0001).
In a first approach, the ratios of cortisol concentrations obtained in both kits were calculated ([A]/[B]). We found differences of [A]/[B] ratios between healthy control adults (ratio 1.2) and infants < 1 yr (ratio 1.8, p < 0.001), patients with CAH (ratio 6.4, p < 0.005), patients under metyrapone
Discussion
Assay accuracy is a major point when a diagnosis rests upon hormonal threshold concentrations. Adrenal status for instance can be established by cortisol concentrations at 8 h. An overestimation of cortisol concentrations leading to a subsequent failure to diagnose hypocorticism may have dramatic consequences. In the current study, we showed an example of 2 kits failing to identically assess adrenal status when an excess of “unusual” steroids is present in the serum of patients despite
Acknowledgment
The authors wish to thank Dr A Georges for her help in reading an early version of the manuscript.
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