Therapeutic Drug Monitoring (TDM) of immunosuppressant drugs plays an important role in transplantation medicine but also in patients treated with these drugs suffering from autoimmune diseases. There are two analytical methods which are used for the determination of these compounds, immunoassays and chromatographic procedures mainly high-pressure liquid chromatography coupled to mass spectrometry (LC-MS/MS). The quantification of the active metabolites of the purine antagonists is most often done in patients with autoimmune diseases in order to prevent severe side effects. Ciclosporin is the only drug where also peak levels are regularly determined (so called C2 levels) because it has been shown that these concentrations correlate better with the pharmacological effect. TDM of mycophenolic acid is nowadays also most often done by the quantification of through levels despite the fact that the area under concentration-time curve (AUC) has a much better correlation with the outcome. But the determination of also an abbreviated AUC asks for different blood samples over at least a 4 hours period of time. All immunoassays have cross-reactivities of the antibodies either to the metabolites of the drugs or in the case of everolimus and sirolimus both compounds are also detected by the assays quantifying the other drug. The different levels of cross reactivities have to be taken into account for the interpretation of patient results.