Highly Responsive Bioassay for Quantification of Glucocorticoids.

Abstract

Measurement of total cortisol levels in serum samples is currently based on immunoassays or liquid chromatography-mass spectrometry (LC-MS/MS). However, measurement of bioavailable cortisol is laborious, unreliable, and inconvenient for the patient. Therefore, a new versatile assay with the ability to measure both total and bioavailable cortisol from serum represents an important supplement to the current methods. We have generated a cell-based glucocorticoid reporter assay (HEK293F-GRE). The assay was validated for cell line stability, accuracy by dilution, precision, repeatability, reproducibility, and specificity. Additionally, the assay was tested for measuring both total and bioavailable cortisol in serum. The assay showed linearity at five dilution levels with R2 = 0.98 and an accuracy between 0.8 and 1.2. Precision (CV < 20%) was validated down to 3-6 nM dexamethasone, and estimation of the total cortisol concentration was comparable to cortisol immunoassay and LC-MS/MS in most serum samples. Moreover, the assay estimated the bioavailable cortisol fraction in serum samples to a level that agreed with the literature. The HEK293F-GRE assay holds the potential to be a complementary method for estimating cortisol in clinical practice. The ability to quantify bioavailable cortisol directly from serum samples is alluring and provides an opportunity for monitored and personal dose regimens of exogenous glucocorticoids.