Launchpad 2

Dried lipid A samples were dissolved with 10 μl of 50% (v/v) methanol/chloroform. Then, 1 μl of the lipid A sample was mixed with 1 μl of 50 mg 2,5-dihydroxybenzoic acid (DHB) solution in 1 mL 70% (v/v) acetonitrile/water. Subsequently, 2 μl of the mixture was loaded on a MSP 96 target polished steel MALDI plate and dried at room temperature. The lipid A samples were analyzed via a Bruker Daltonics Microflex LRF MALDI-TOF MS instrument and controlled by FlexControl 3.0 software (Bruker, Bremen, Germany). The operating conditions were as follows: negative-ion and reflectron mode, detector gain = 6.9, laser frequency = 60.0 Hz and laser power = 70%. Data acquisition and processing were performed by Flexanalysis 3.3 software (Bruker, Bremen, Germany). MS/MS analysis was performed using an Axima Resonance MALDI-quadrupole ion trap-TOF instrument (Shimadzu, Manchester, UK). The instrument was operated in negative ion and reflectron mode. Fragment ions were analyzed by collision-induced dissociation (CID) of the precursor ions and argon gas was used as a collision gas. Operating conditions were as follows: intro endcap = −5.5 kV, extr endcap = 10.0 kV, flight tube = 10.0 kV, reflectron center = −0.2 kV, reflectron back = −0.2 kV, and Detector = 2.2 kV. The data acquisition and processing were performed using the Launchpad 2.9.3 software (Kratos Analytical, Manchester, UK).