Nonspecific Binding in LC–MS Bioanalysis

Nonspecific binding (NSB) is an undesirable yet very common phenomenon in LC-MS bioanalysis. This chapter explores how people would even know NSB exists; ways to evaluate its severity; why is NSB particularly problematic for biomarker quantitation; and how it can be differentiated from a seemingly similar stability issue. The questions are addressed using representative application examples from the LC-MS bioanalysis of both small and large molecules. To confirm NSB or evaluate the severity, many different approaches can be taken, such as multiple sequential transfers, the deliberate preparation of small and large volumes, and frequent comparisons with fresh spiking. The key behind these various approaches is to amplify the impact of NSB by exposing the compounds of interest to as large of a surface area as possible and by exposing as many times as possible, so that it will not go unnoticed.