Lipid A Extraction and Analysis by MALDI-TOF MS

Lipid A analysis was conducted using the methods described in Sorensen et al. (65 (link)). Briefly, pelleted bacteria were smeared onto a MALDI plate with a sterile inoculation loop. A total of 1 μL of buffer, consisting of 0.2 M anhydrous citric acid and 0.1 M trisodium citrate dihydrate, was spotted atop the spotted colony. The plate was then incubated at 110°C for 30 minutes to extract membrane lipids. The plate was then rinsed with endotoxin-free water to wash cell debris. One microliter of 10 mg/mL norharmane matrix suspended in 2:1 chloroform-methanol was spotted onto extracted sample on the MALDI plate. Mass spectra was collected using a Bruker Microflex LRF MALDI-TOF MS instrument in negative ion and reflectron mode.