ELUTION OF LIPOPOLYSACCHARIDES FROM POLYACRYLAMIDE GELS

A simple procedure for elution in water of bacterial lipopolysaccharid es (LPS) from sodium dodecyl sulfate-polyacrylamide gels is described. It consists of the combination of three principal steps: first, highl y sensitive on-gel LPS detection (1-10 ng/band) with zinc-imidazole (n egative or reverse staining); second, washing of the individual LPS ba nd in a solution of a zinc-complexing agent (e.g., 100 mM EDTA); and f inally, elution of the LPS (100 - 200 mu l water for a 0.5-mu g LPS ba nd) from gel microparticles for 3 h at room temperature. Using this pr ocedure, we have successfully eluted a variety of LPS forms from Borde tella pertussis, Escherichia coli 0111:B4, E. coli K-235, Salmonella e nteritidis, and Pseudomonas aeruginosa. Elution recovery of rough or s emismooth LPS was about 70-80%, while that of smooth LPS was only abou t 10%. Eluted LPS was biologically active as tested by limulus amebocy te lysate and TNF-alpha assays. (C) 1998 Academic Press.