Ultra-high-performance liquid chromatography tandem mass spectrometry (UHPLC-MS/MS) is an analytical technique that combines the separating abilities of liquid chromatography with the highly sensitivity and selective mass analysis of mass spectrometry. The sample solution containing analytes of interest are pumped through a liquid chromatography column (LC column) also known as the stationary phase by a mobile phase flowing through at high pressure. The LC column are chosen to match the specific physiochemical profile of the analyte of interest. Chemical interactions between the components of the sample, the LC column and the mobile phase affects migration rates through the LC column. These all affect the separation of compounds of the sample solution. Therefore, the components of the LC column and the mobile phase allows for customizing separation to suit many complex solutions.

After elution from the LC column, the effluent undergoes an evaporation and ionization process and gets transferred directed into the mass spectrometer. The ionization process is essential for the mass spectrometry (MS) method as it is only the charged particles that will be quantified by the MS system. After ionization a number steps through electromagnetic fields with high specificity for the structure of the compound of interest is performed to provide a high degree of selectivity. 

UHPLC is a suitable method to overcome the challenges of low sample volumes and low concentrations obtained in microdialysis studies.