A novel MS3 experiment for quantifying ions with a linear ion trap

Liquid chromatography coupled with tandem mass spectrometry has long been employed for the quantitation of molecules. With judicious selection of precursor and fragment ions, multiple-reaction monitoring assays can be developed rapidly for these experiments. However, there are cases where analyses struggle due to high background signals caused by matrix effects that interfere with the analytical signal. An alternative to MRMs involves using two stages of tandem mass spectrometry — an MS3 experiment. Although this technique can provide greater selectivity than MS/MS experiments, cycle times for MS3 experiments are typically longer than MRM-type experiments. Here, we present a quantitation technique employing an MS3 method with shorter cycle times than traditional linear ion trap MS3 scans. Termed “scan-free” MS3, this technique performs “mass analysis” by isolating the ions of interest in the linear ion trap and then emptying the trap of these ions. The signal will be due only to those preselected ions, resulting in an MS3 experiment with up to a ∼35% reduction in cycle times relative to standard MS3 experiments without loss of sensitivity. We compare the analytical performance of this method with MRMs, as well as standard MS3 experiments, finding equivalent or better performance from the scan-free MS3 method.