Biochemical laboratory

In the Biochemical Laboratory of the Department of Biological and Clinical Psychology, analytes can be determined from the body fluids blood, saliva, and urine, as well as from cell culture supernatants, using a wide variety of methods.

Free cortisol in saliva - DELFIA

More than 30 years ago, the development of a highly specific method for the analysis of cortisol in saliva (Dressendörfer et al., 1992) laid the foundation for a large number of studies on the responsiveness of the hypothalamic-pituitary-adrenocortical (HPA) axis. In many psychobiological studies of HPA axis activity, cortisol is measured as its end product. Most of the cortisol in serum is bound to transport proteins and thus is not biologically active. Only the smaller fraction of free cortisol finds passage into saliva; correlations between concentrations in saliva and serum are very high ~r=.90.

We use a highly specific DELFIA immunoassay to determine the concentration of free cortisol in saliva. DELFIA stands for "Dissociation-Enhanced Lanthanide Fluorescent Immunoassay" and is a highly sensitive and specific analytical method used in various bioanalytical and diagnostic tests. DELFIA uses lanthanide chelates (rare earths) as a fluorescent marker to enhance the detection of specific molecules or markers such as antibodies, antigens or other biological molecules, in our case cortisol. Key features of DELFIA include the ability to provide a low background signal, high sensitivity and a wide dynamic range.

Alpha amylase in saliva

Alpha amylase is an enzyme found in various biological fluids, including saliva. It plays an important role in the digestive system by breaking down starch and glycogen into simpler sugars, especially maltose. Alpha-amylase is therefore involved in the digestion of carbohydrates. There are two main sources of alpha-amylase in the human body: the pancreas (pancreatic amylase) and the salivary glands (salivary amylase). The concentration of alpha-amylase in saliva can serve as an indirect marker of stress responses. During acute stress, the release of alpha-amylase in saliva increases. Therefore, this measurement is often used in psychology and stress research to study the physiological response to stressful situations.

A colorimetric assay using 2-chloro-4-nitrophenyl-aD maltrotrioside (CNP-G3) as substrate is used to determine a-amylase activity in saliva samples. Amylase activity cleaves the color-producing CNP group and displays it as an increase in absorbance. The color intensity of the formed p-nitrophenol (CNG) is directly proportional to the amylase activity and is measured photometrically.

Sex steroids in saliva

Estradiol, progesterone, and testosterone can generally be measured in saliva. Reliability is high to very high, depending on the assay. The correlations between saliva and plasma are as follows, depending on the analyte and assay: Testosterone: r ~ 0.6–0.9 Progesterone: r ~ 0.5–0. Estradiol: r ~ 0.5–0.9, depending on the phase of the cycle and the assay. Whether saliva values are suitable for determining or validating the phase of the cycle is currently the subject of research.

The measurement is carried out in our lab using commercial ELISAs (Demeditec, Salimetrics, IBL, etc.). The steroid hormones are quantified using solid-phase enzyme immunoassays (ELISA), which are based on the principle of competitive binding. The unknown amount of free steroid hormone (antigen) contained in the sample and an enzyme-labeled steroid hormone (enzyme conjugate) compete for the limited binding sites of specific antibodies bound to the surface of the reaction vessels. After a defined incubation period, unbound components are removed by washing steps. The bound enzyme conjugate contains horseradish peroxidase (HRP), which reacts with the chromogenic substrate tetramethylbenzidine (TMB). This reaction leads to the development of a blue color, the intensity of which changes to yellow after the addition of a stop solution (acid) and is quantified photometrically as optical density (OD) at the corresponding wavelength. The color intensity is inversely proportional to the concentration of the steroid hormone in the sample. To determine the hormone concentrations, a standard curve is created by plotting the measured OD values against the known concentrations of the standards. The concentrations of the unknown samples are then calculated by interpolation using this standard curve.

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Numerous research groups in Germany and abroad use the competence of our laboratory.
If you have any questions regarding possible analyses and conditions, please do not hesitate to contact our team at cortlabuni-trierde