Super dhb solution

Lipid A was precipitated from GMMA using mild-acid hydrolysis and was analyzed with an Ultraflex matrix-assisted laser desorption ionization–time of flight (MALDI-TOF) mass spectrometer (Bruker Daltonics) in negative-ion reflectron mode as reported previously (20 (link)). A peptide calibration standard (Bruker Daltonics) was included in each analysis. The samples and the standard were mixed with Super DHB solution (Sigma-Aldrich), a matrix substance for MALDI-TOF analyses. The m/z ratios were determined by flexAnalysis software in comparison to the peptide standard.

Lipid A was precipitated from GMMA using mild acid hydrolysis with 1% acetic acid for 2 h at 100 °C (35 ). Samples were centrifuged at 14,000 × g for 15 min; the pellets were resuspended in water and washed twice with water. The pellets were dried overnight using a SpeedVac and resuspended in chloroform/methanol 4:1 and mixed with an equal volume of Super DHB solution (Sigma). 2 μl of the mixture were loaded to the target plate (MTP 384 target plate ground steel BC, Bruker Daltonics) and analyzed by Ultraflex MALDI-TOF (Bruker Daltonics) in reflectron ion-negative mode. A peptide calibration standard (Bruker Daltonics), mixed with the Super DHB solution, was included in each analysis. For MS/MS analysis of lipid A, main peaks from the linear mode analysis were selected for collision-induced dissociation, and the resulting fragments were detected by MALDI TOF-TOF in ion negative mode. For each sample, spectra represent the integration of the analysis of 20 different areas of the spot by 50 single laser shots. The m/z rations were determined by Flex Analysis software in comparison with the peptide standard.