| Title | PROTOCOL] Applications of different solvents and conditions for differential extraction of lipopolysaccharide in Gram-negative bacteria |
|---|---|
| Author | Mai Phuong Nguyen1, Le Viet Ha Tran1, Hyun Namgoong2, and Yong-Hak Kim1* |
| Address | 1Department of Microbiology, Daegu Catholic University School of Medicine, Daegu 42472, Republic of Korea, 2Kolon R&D Division, Seoul 07793, Republic of Korea |
| Bibliography | Journal of Microbiology, 57(8),644–654, 2019, |
| DOI | 10.1007/s12275-019-9116-5 |
| Key Words | Gram-negative bacteria, lipopolysaccharide (LPS), quality index, effective does (ED50), macrophage activation |
| Abstract | Lipopolysaccharide (LPS) is one of the major components in the outer membrane of Gram-negative bacteria. However, its heterogeneity and variability in different bacteria and differentiation conditions make it difficult to extract all of the structural variants. We designed a solution to improve quality and biological activity of LPS extracted from various bacteria with different types of LPS, as compared to conventional methods. We introduced a quality index as a simple measure of LPS purity in terms of a degree of polysaccharide content detected by absorbance at 204 nm. Further experiments using gel electrophoresis, endotoxin test, and macrophage activation test were performed to evaluate the performance and reliability of a proposed ‘T-sol’ method and the biological effectiveness and character of the LPS products. We presented that the T-sol method had differential effects on extraction of a RAW 264.7 cell-activating LPS, which was effective in the macrophage activation with similar effects in stimulating the production of TNF-alpha. In conclusion, the T-sol method provides a simple way to improve quality and biological activity of LPS with high yield. |