on 05-01-201210:55 AM - edited on 10-15-202111:23 AM by Closed Account
Huhmer AFR, Hao Z, Zhang Y, Biringer RG, Fonteh NF, Kauffman S, Harrington MG. Scientific Poster Traditionally, targeted peptide quantification studies use a triple quadrupole (QQQ) mass spectrometer because of the high throughput and high sensitivity, allowing for large numbers of proteins to be quantified in a single LC/MS experiment.However, the transition from discovery to target verification and quantification can be challenging due to the transition between mass spectrometer platforms. The newly developed Thermo Scientific Q Exactive hybrid quadrupole-OrbitrapTMmass spectrometer (Figure 1) enables the seamless transition from discovery to target quantification and verification on a single platform. We describe two high-resolution and accurate mass (HR/AM) targeted peptide quantification approaches. The quadrupole-based high-resolution SIM scan ensures accurate target selection and high sensitivity. Spectrum multiplexing (msx) and concurrent ion injection and detection greatly improve duty cycle (Figure 2). Targeted quantification can also be done using an HCD-based SRM like approach where product ion peak area is integrated for quantification. This approach provides further selectivity from second level of MS (Figure 3). Table 1 lists method parameters for both approaches.
In this study, we evaluated the performance of Q Exactive mass spectrometer on the quantification of peptide targets in complex biological samples. Peptides derived from eicosanoid pathway enzymes in human cerebrospinal fluid (CSF) were quantified using the targeted HCD approach. Eicosanoids exert complex control of inflammation or immunity, and are messengers in the central nervous system. The targeted MSX SIM approach was evaluated using standard peptides spiked into a yeast tryptic digest. Detection limits and linear dynamic range were obtained for these approaches .