Detection of submicrogram quantities of Escherichia coli lipopolysaccharides by agarose-gel electrophoresis

Anal Biochem. 2003 Nov 15;322(2):185-9. doi: 10.1016/j.ab.2003.08.017.

Abstract

A sensitive agarose-gel electrophoresis method has been developed for the visualization of lipopolysaccharide (LPS) samples from several Escherichia coli serotypes, such as 026:B6, 055:B5, 0128:B12, 0111:B4, 0127:B8, and K235. This method can detect as little as 0.5-6 microg of LPS depending on the serotype by treatment of the plates with toluidine blue, and it is able to measure sub-microgram amounts of samples, approx. 0.05-0.5 microg, when the staining with toluidine blue followed by Stains-All procedure is adopted. Treatment of LPS with alkali under anhydrous conditions removes the ester-linked fatty acids and the phosphate groups of the Lipid A component, producing the detoxified LPS. The carbohydrate neutral moiety obtained from the hydrolysate does not migrate during electrophoresis. This was utilized to monitor quantitatively the removal process of the Lipid A component for the formation of the detoxified LPS.

MeSH terms

  • Coloring Agents / chemistry
  • Electrophoresis, Agar Gel / methods*
  • Escherichia coli / chemistry*
  • Escherichia coli / classification
  • Lipid A / analysis
  • Lipid A / chemistry
  • Lipopolysaccharides / analysis*
  • Lipopolysaccharides / isolation & purification
  • Reproducibility of Results
  • Sensitivity and Specificity
  • Serotyping
  • Tolonium Chloride / chemistry

Substances

  • Coloring Agents
  • Lipid A
  • Lipopolysaccharides
  • Tolonium Chloride