The Calbiotech ACTH ELISA Kit is intended for the quantitative determination of ACTH Adrenocorticotropic Hormone) in human plasma.
SUMMARY AND EXPLANATION
ACTH is a 39-amino acid peptide hormone (MW=4500) secreted by the pituitary to regulate the production of steroid hormones by the adrenal cortex. ACTH increases the synthesis and release of all adrenal sterioids, aldosterone, cortisol and adrenal androgens. It is the principal modulator of cortisol, the most important glucocorticoid in man. As the cortisol level in blood increases, release of ACTH is inhibited directly at the pituitary level. Through this same mechanism, decreasing cortisol levels lead to elevated ACTH levels. In healthy individuals, ACTH reaches a peak in the early morning (6:00 – 8:00 hour) and levels become lowest late in the day and near the beginning of the sleep period. Stress may also override the diurnal variation. Plasma ACTH assays are useful in the differential diagnosis of pituitary Cushing’s disease, Addison’s disease, autonomous ACTH producing pituitary tumors (e.g. Nelson’s syndrome), hypopituitarism with ACTH deficiency and ectopic ACTH syndrome. Primary adrenocortical insufficiencies, Addison’s disease. Hypopituitarism with ACTH deficiency, which is secondary adrenocortical insufficiency, is characterized by low plasma ACTH and cortisol concentrations, and a subnormal, but usually distinct adrenal response to stimulation with synthetic ACTH (Cortrosyn).
PRINCIPLE OF THE TEST
The CBIACTH Immunoassay is a two-site ELISA for the measurement of the biologically active 39 amino acid chain of ACTH. One antibody is prepared to bind only the C-terminal ACTH 34-39 and this antibody is biotinylated. The other antibody is prepared to bind only the mid-region and N-terminal ACTH 1-24 and this antibody is labeled with HRP for detection. In this assay, calibrators, controls, or patient samples are simultaneously incubated with the enzyme labeled antibody and a biotin coupled antibody in a streptavidin-coated microplate well. At the end of the assay incubation, the microwell is washed to remove unbound components and the enzyme bound to the solid phase is incubated with the TMB substrate. Stop solution is then added to stop the reaction and converts the color to yellow. The intensity of the yellow color is directly proportional to the concentration of ACTH in the sample. A dose response curve of absorbance unit vs. concentration is generated using results obtained from the calibrators. Concentrations of ACTH present in the controls and patient samples are determined directly from this curve.