SWATH- mass spectrometry coupled with isotope labelled standard peptides for absolute quantification (AQUA) of proteins is a relatively new technique for the characterization of proteins. In this study, a mass spectrometry-based method for absolute quantification of selected influenza A virus proteins (hemagglutinin, neuraminidase, nucleoprotein, matrix protein 1) is presented.

Infection of Madine-Darby kidney cells (MDCK) with influenza A/PR/8/34 at an MOI of 10. Sampling for a time course of 13 hours post-infection. Determination of absolute protein copy numbers of matrix protein 1, hemagglutinin, neuraminidase and nucleoprotein by spike-in of isotopically labelled peptides before tryptic digest. TimsTofpro measurements were carried out with the SWATH method by defining m/z windows of 25 m/z with an overlap of 1 m/z

Cells: MDCK, suspension, ECACC, #84121903 Virus: influenza A/PR/8/34; MOI of 10 Cell pellet and supernatant samples Medium: Driving M medium Volume: 100 mL working volume in 250 mL baffled shaker flasks Cell pellet sampled were washed twice with PBS; both samples were heat inactivated before MS preparation