Journal of Endotoxin Research

 

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Journal of Endotoxin Research, Vol. 1, No. 4, 207-215 (1994)
DOI: 10.1177/096805199400100402

Design of a semi-synthetic (polymyxin-B•NHS-LC-biotin)4lavidin tetramer and evaluation of its ability to inhibit lipopolysaccharide induced TNF{alpha} synthesis

C.P. Coyne

Veterinary Pharmacology Research Laboratory, Veterinary Research Program, College of Veterinary Medicine, Wise Center, Mississippi State University, MS, USA

J.T. Moritz

Veterinary Pharmacology Research Laboratory, Veterinary Research Program, College of Veterinary Medicine, Wise Center, Mississippi State University, MS, USA

A method is described for the semi-synthetic production of (polymyxin-B•NHS-LC-biotin) 4lavidin tetramer. In phase I of semi-synthesis procedures, polymyxin-B was reacted with NHS-LC-biotin reagent resulting in the generation of the reactive intermediate, polymyxin-B•NHS-LC-biotin.

In phase II of the conjugation procedure, the polymyxin-B•NHS-LC-biotin complex was combined with purified avidin producing the tetramer complex, (polymyxin-B•NHS-LC-biotin) 4lavidin. A 'long-chain' biotinylation reagent, NHS-LC-biotin minimized steric hinderance phenomenon thereby maximizing the binding of multiple lipopolysaccharide molecules.

Binding avidity of (polymyxin-B•NHS-LC-biotin) 4lavidin for lipopolysaccharide was established with the application of fluorescein isothiocyanate Escherichia coli (055:B5) lipopolysaccharide conjugate (FITC-LPS). Applying a tissue culture based biological assay, (polymyxin-B•NHS-LC-biotin) 4lavidin was capable of inhibiting Salmonella minnesota (RS) lipopolysaccharide induced synthesis of tumor necrosis factor{alpha} (TNF{alpha}).


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