An enzyme immunoassay for the quantitative in vitro diagnostic measurement of active free cortisol (hydrocortisone and hydroxycorticosterone) in saliva. Measurements of cortisol are used in the diagnosis and treatment of disorders of the adrenal gland.
Summary and Explanation
The hormone Cortisol is vital for several functions of the human body. A strong correlation exists between stress related conditions and Cortisol levels (13) Cortisol is a steroid hormone made in the adrenal glands. Among its important functions in the body include roles in the regulation of blood pressure and cardiovascular function as well as regulation of the body's use of proteins, carbohydrates, and fats. Cortisol secretion increases in response to any stress in the body, whether physical (such as illness, trauma, surgery, or temperature extremes) or psychological. When cortisol is secreted, it causes a breakdown of muscle protein, leading to release of amino acids into the bloodstream. These amino acids are then used by the liver to synthesize glucose for energy, in a process called gluconeogenesis. This process raises the blood sugar level so the brain will have more glucose for energy. Cortisol also leads to the release of so-called fatty acids, an energy source from fat cells, for use by the muscles. Taken together, these energy-directing processes prepare the individual to deal with stressors and ensure that the brain receives adequate energy sources (4).
Cortisol is the most potent glucocorticoid produced by the human adrenal (5-7). It is synthesized from cholesterol and its production is stimulated by pituitary adrenocorticotropic hormone (ACTH) which is regulated by corticotropin releasing factor (CRF). ACTH and CRF secretions are inhibited by high cortisol levels in a negative feedback loop. Cortisol acts through specific intracellular receptors and affects numerous physiologic systems including immune function, glucose counter regulation, vascular tone, and bone metabolism.
Elevated cortisol levels and lack of diurnal variation have been identified with Cushings disease (ACTH hypersecretion). Elevated circulating cortisol levels have also been identified in patients with adrenal tumors. Low cortisol levels are found in primary adrenal insufficiency (e.g. adrenal hypoplasia, Addisons disease) and in ACTH deficiency. Due to the normal circadian variation in cortisol levels (8), distinguishing normal from abnormally low cortisol levels can be difficult, therefore several daily collections are recommended.
Saliva is an excellent medium to measure steroids because it is a natural ultra-filtrate of blood, and steroids not bound by carrier proteins in the blood freely diffuse into saliva. Only about 1-10% of the steroids in blood are in the unbound or free form, and it is this fraction that diffuses into target tissues of the body, and into saliva (9, 10). The majority (90-99%) of steroid hormones in the blood are bound to carrier proteins (cortisol binding globulin, sex-hormone binding globulin and albumin) and are unavailable to target tissues. The process of passive diffusion of non-bound (free) steroid hormones occurs because these small molecules are of a low molecular weight (less than 400 daltons) and are relatively nonpolar, thus enabling them to freely diffuse from blood to saliva. Bound steroids are too large to diffuse freely through the salivary cells into the salivary gland lumen. (11-14)
The DRG Salivary Cortisol ELISA Kit is a solid phase enzyme-linked immunosorbent assay (ELISA), based on the principle of competitive binding.
The microtiter wells are coated with a monoclonal (mouse) antibody directed towards an antigenic site on the cortisol molecule.
Endogenous cortisol of a patient sample competes with a cortisol-horseradish peroxidase conjugate for binding to the coated antibody. After incubation the unbound conjugate is washed off.
The amount of bound peroxidase conjugate is inversely proportional to the concentration of cortisol in the sample. After addition of the substrate solution, the intensity of colour developed is inversely proportional to the concentration of cortisol in the patient sample.