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Methods in molecular biology (Clifton, N.J.)

Two efficient methods for the conjugation of smooth-form lipopolysaccharides with probes bearing hydrazine or amino groups. I. LPS activation with cyanogen bromide.


PMID 21567325

Abstract

This chapter presents a conjugation method for coupling probes bearing hydrazine or primary amino groups to a smooth(S)-form lipopolysaccharide (LPS). LPS is modified by the activation of the hydroxyl groups present in its O-antigen moiety with cyanogen bromide in aqueous acetone. The method yields conjugates with good labeling ratios, preserving the endotoxic activity of the lipid A moiety. Conjugation of smooth-form LPS from Salmonella enterica sv. Minnesota with dansyl hydrazine and horseradish -peroxidase yields labeling ratios above 300 nmol dansyl per mg LPS, with nearly no loss of the original endotoxin activity. In the case of horseradish peroxidase, introducing a spacer, a ratio of 28 nmol HRP per mg LPS is obtained, preserving 65% of the original endotoxic activity.