Analysis of lipopolysaccharides ofBacteroides fragilisby sodium dodecyl sulphate-polyacrylamide gel electrophoresis and electroblot transfer

Abstract

Lipopolysaccharides (LPS) from three strains ofBacteroides fragilis were run on SDS-polyacrylamide gels and stained with silver. Each LPS produced a similar pattern, consisting of a series of regularly spaced discrete bands which decreased in intensity as they increased in M r value. Electroblot transfer from duplicate SDS gels onto nitrocellulose membrane were reacted with antisera raised to whole cells of two of the strains and antigens were visualised with horse-radish peroxidase-antirabbit-IgG conjugate and colour reagent. Results revealed that the two lowest M r bands of the LPS preparation (rough LPS) represented common antigens. INTRODUCTIONB. fragilis, the most commonly isolated anaerobic bacterium from clinical specimens, possessed surface factors, often referred to as "capsule", which appear to confer it with a greater pathogenic potential than that of closely related species [1].In 1976 Kasper suggested that the capsular polysaccharide may be a species-specific antigen The.investigations into the roles of surface components as virulence factors and species-specific antigens have therefore been compromised by attempting to use preparations of unknown purity.In this study we show that LPS prepared by classical aqueous phenol technique from B. fragilis can be analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) and stained with silver. It can also be transferred to nitrocellulose membrane and probed with antisera raised against whole cells of B. fragilis. From the results obtained we have been able to comment on the purity of the LPS and suggest the nature of the species-specific antigen. MATERIALS AND METHODS Bacterial strains and growth conditionsB. fragilis NCTC9344, and two clinical strains 0378-1097/83/$03.00