Masshunter workstation qualitative analysis b 06

Lysates of new and old mitochondria from 4 individual FACS sorts were pooled. Mitochondrial samples, MEGM media sample and whole cell sample were frozen on dry ice, lipids extracted according to the method of Folch^{52 (link)}
and stored at -80°C. The solvent was evaporated, lipid residue dissolved in methanol/chloroform (2:1, v/v), spiked with a cocktail of 13 internal standards (including representatives for each PL class detected), and supplemented with 1% NH₄OH before direct infusion of the sample solution into the ESI source of a triple quadrupole mass spectrometer, Agilent 6490 Triple Quad LC/MS (with iFunnel technology; Agilent Technologies). In addition to positive and negative ion mode MS detections, PL class specific MS/MS detection modes were used with information on equipment and ion source settings^{53 (link)}
. Spectra were processed by MassHunter Workstation Qualitative Analysis B.06.00 software (Agilent Technologies) and the concentrations of individual PL species quantified using internal standards and software Lipid Mass Spectrum Analysis (LIMSA, v.1.0)^{54 (link)}
. Concentration data were converted to PL class composition (as molar %).