Introduction
Targeted LC-MS assays are increasingly applied in the post-discovery omics area with an emphasis on validation. A novel DIA mode of operation, which provides both qualitative and quantitative information, has been developed for a tandem quadrupole/oa-time-of-flight (ToF) MS and applied to targeted lipidomics experiments using transition extraction lists and compound library based approaches.
Method
LC separation was performed using a Waters Acquity UPLC equipped with a 2.1mm x 100mm column. The m/z isolation range of the quadrupole was continuously scanned with MS data acquired up to a frequency of 2000 spectra/s. Alternate MS scans comprised precursor and CID product ions. The resulting 2D data, m/z (ToF) vs. m/z (quadrupole) were processed using Progenesis and Skyline informatics.
Results
Quantitative proof-of-principle data were acquired by serially diluting lipid standards in protein precipitated plasma. Analysis indicates that scanning quad DIA enables over an order of magnitude more specificity than a static quad operated with the same resolution and it was found that a quadrupole transmission window of approx 10 Da provided optimum identifications. Qualitative information was obtained by extracting lipid class information based on neutral loss or product ion extraction. Control, diabetic and obese human plasma samples were treated with isopropanol and centrifuged to precipitate proteins and the lipid-containing layer collected. 2DMS data were collected and differentially expressed lipid classes across the three conditions were quantified. The obtained results agreed with previous discovery studies and the expected changes in relation to disease and/or phenotype.
Novel Aspect
Targeted quantitation in lipidomics using a novel precursor quadrupole scanning based DIA method