Acid hydrolysis of LPS.

In our previous work (5), we tested LPS from Escherichia coli, Salmonella enterica, and Klebsiella pneumoniae. We found that each of these LPS types can stabilize poliovirus. In this study, we chose LPS from E. coli for detailed study. LPS from E. coli O127:B8 (L3129; Sigma-Aldrich) was dissolved in 2% acetic acid at 5 mg/ml. After incubation at 100°C for 2 h with occasional shaking, chloroform and methanol were added to yield a final chloroform/methanol/water ratio of 2:1:3. The mixture was vortexed and centrifuged at 5,000 × g for 10 min. The resulting upper phase containing detoxified LPS and the lower phase containing lipid A were collected separately and dried in a Speed-Vac. For monosaccharide analysis, detoxified LPS was dissolved in 1 M HCl and heated at 100°C for 30 min. Samples were dried in a Speed-Vac.