Surfactant-assisted lipopolysaccharide conjugation employing a cyanopyridinium agent and its application to a competitive assay

Abstract

The activation of a lipopolysaccharide (LPS) with 1-cyano-4- dimethylaminopyridinium tetrafluoroborate (CDAP) in the presence of a surfactant allows an efficient conjugation with dansyl hydrazine or horseradish peroxidase (HRP) in an aqueous medium maintaining its biological activity. In order to promote the reaction a series of amphiphilic compounds were tested, sodium deoxycholate being the most suitable. The method presents several advantages: it is carried out in a mild environment, good conjugation ratios are obtained, it is suitable for any label bearing amino, hydrazine, or hydrazide groups, and the LPS endotoxic and HRP enzymatic activities are preserved. The HRP conjugate is applied in an amperometric competitive assay for the detection of lipopolysaccharides in an electrode array combined with a multipotentiostat able to carry out simultaneous determinations. The system is able to detect samples in concentrations as low as 100 pg mL-1 of LPS.