Accurate cortisol measurement is essential in diagnosing adrenal insufficiency and adjusting stress dosing.  While immunoassays are commonly used in clinical settings, their limited molecular specificity makes them less reliable for assessing adrenocortical function, especially since total cortisol levels are influenced by binding proteins like corticoid-binding globulin and albumin.  Mass spectrometry, combined with chromatographic separation, offers superior specificity and sensitivity, enabling precise quantification of free cortisol and multiple steroid hormones in a single analytical run.