Hormone determination
Most hormones are present in the blood in very low concentrations. This requires special methods that are very complex and prone to failure.
Even before a blood sample is taken it is important to consider the diurnal rhythm, pulsatility and interference (e.g. use of the contraceptive pill). The rapid breakdown of some hormones in the blood makes rapid processing and targeted preparation of the samples necessary, depending on the hormone tested.
Another important feature of hormone determination is the hormone-binding proteins! Only the “free hormones” – i.e. only the hormones that are not bound to proteins – are available for the effect! This requires special knowledge to select the correct method for determining the effective hormone concentration – a problem that has by no means been solved for all hormones!
The chapter on error analysis will address the issue of accuracy and precision. In particular, it should be made clear why the values obtained with the same methodology must fluctuate greatly, especially at high and low concentrations!
In the interpretation of the results, age, gender, etc. must also be taken into account.
Development and methods
Almost all of the hormones produced by endocrine glands were discovered by endocrinologists. The method of determining the concentration of messenger substances in the blood was almost always first described in endocrinology laboratories.
The RIA (Radio-Immuno-Assay) was developed as an essential tool for the routine determination of hormones, in which antibodies specially cultivated against the corresponding hormone (usually in rabbits) are used with hormones labeled with radioisotopes. The RIA is based on the law of mass action with the associated, unavoidable, large variances in the range of high or low hormone concentrations.
In the IRMA– the Immun Radiometric Assay – two antibodies directed against the hormone are used, one firmly bound in the tube, the other radioactively labeled and in dissolved form. As the hormone concentration in the incubate increases, more hormone is bound to the solid antibody. However, the hormone also binds the labeled antibody, so that the radioactivity increases with the concentration of the substance to be measured after decanting the tube.
In addition to the use of radioactive substances, the same examination principles are applied only using other endpoint determinations (EIA, FIA, LIA, ILMA)
Preanalytics
Daily rhythm
Pulsatility
Interference
The problem of "free" hormones
Thyroid hormones
Cortisol
IGF1
Error analysis
The limits of “correctness”