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Elution of lipopolysaccharides from polyacrylamide gels

Abstract:

A simple procedure for elution in water of bacterial lipopolysaccharides (LPS) from sodium dodecyl sulfate-polyacrylamide gels is described. It consists of the combination of three principle steps: first, highly sensitive on-gel LPS detection (1-10 ng/band) with zinc-imidazole (negative or reverse staining); second, washing of the individual LPS band in a solution of a zinc-complexing agent (e.g., 100 mM EDTA); and finally, elution of the LPS (100-200 μl water for a 0.5-μg LPS band) from gel microparticles for 3 h at room temperature. Using the procedure, we have successfully eluted a variety of LOPS forms from Bordetella pertussis, Escherichia coli 0111:B4, E. coli K- 235, Salmonella enteritidis, and Pseudomonas aeruginosa. Elution recovery of rough or semismooth LOPS was about 70-80%, while that of smooth LPS was only about 10%. Eluted LPS was biologically active as tested by limulus amebocyte lysate and TNF-α assays.