Design of experiments for development and optimization of a LC‐MS/MS bioanalytical assay

Design of experiments (DoE) is a valuable tool for optimization of a quantitative bioanalytical method utilizing liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Liquid chromatography mass spectrometry (LC-MS) is composed of several processes, including, liquid introduction and analyte ionization. The goal is to transfer analytes from atmospheric pressure to vacuum and maintain conditions that are compatible for both the LC and the MS. These processes involve many experimental factors which need to be simultaneously optimized to obtain maximum sensitivity and resolution at minimum retention time. In this tutorial the basic concepts of DoE will be explained with focus on practical use of DoE, and then three case studies about utilization of DoE for optimization of liquid chromatography tandem mass spectrometry (LC-MS/MS) quantitative assays will be presented.