Detection of the Diurnal Rhythm of Cortisol Secretion - Essay Example

Irregular high levels of cortisol indicate stress in a patient’s case. Enzyme linked immunosorbent assay (ELISA) utilises antibodies coupled to colour changes in order to qualitatively and quantitatively evaluate substances in biological organisms’ fluid streams such as blood and saliva (Lequin, 2005) (De La Rica & Stevens, 2012). A competitive ELISA operates by competitive binding where the competition occurs between already bound antigens and standard added sample antigens. Competitive ELISA allows for strict quantification of results unlike other ELISA methods such as basic ELISA which is slightly quantitative (National Diagnostics, 2013).

Cortisol detection requires the use of competitive ELISA since cortisol molecules are too small to be used with sandwich ELISA methods and need to be bound to carrier proteins. A competitive ELISA requires that the purified antigen be attached to the surface after which it is probed in the presence of similar free antigens labelled as samples. The free antigens are treated as the standard for comparison and tend to compete with the bound antigens which in turn lead to lower levels of bound antibodies.

The samples are standard yields are then evaluated in order to discern quantitative comparisons (National Diagnostics, 2013). This is illustrated in the diagram provided below. Cortisol tends to pass into saliva relatively quickly so saliva samples from the same person are obtained at regular intervals over 24 hours. Saliva samples are labelled with the time when they are obtained. A microtitre plate is coated with just enough anti-cortisol antibody that can bind around half of the labelled cortisol added.

Row eight is left uncoated in order to measure the non-specific binding of labelled cortisol for tubes without any antibody. In order to set up a calibration graph, a set of