Polymyxin B-horseradish peroxidase conjugates as tools in endotoxin research.
The peptide antibiotic Polymyxin B (PMB) binds to bacterial endotoxin (lipopolysaccharide, LPS). We prepared covalent conjugates of PMB and horseradish peroxidase (HRP) by periodation of HRP-linked oligosaccharides followed by direct condensation with PMB. In addition we prepared monoclonal antibodies (Mabs) to PMB. The PMB-HRP conjugates and anti-PMB Mabs were used to study in ELISA the binding of PMB to LPS from Escherichia coli, Klebsiella pneumoniae, and Pseudomonas aeruginosa. In addition, PMB-HRP was used to quantify lipid A in ELISA, and to stain gram-negative bacteria histochemically. For the study of PMB-LPS interaction, PMB-HRP proved to be superior to the anti-PMB Mabs. PMB-HRP conjugates are useful general probes to detect or measure lipid A and LPS of various species using very simple methods and to stain bacteria, and they may obviate the need for many specific antisera. Thus, PMB-HRP conjugates are useful probes for endotoxin research.
Appelmelk
B J
BJ
Department of Medical Microbiology, Vrije Universiteit, Amsterdam, The Netherlands.
Su
D
D
Verweij-van Vught
A M
AM
Thijs
B G
BG
MacLaren
D M
DM
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial / Q000032:analysis*; D004731:Endotoxins / Q000032:analysis*; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods; D004926:Escherichia coli; D006090:Gram-Negative Bacteria / Q000166:cytology / Q000302:isolation & purification; D006735:Horseradish Peroxidase; D007150:Immunohistochemistry / Q000379:methods; D007711:Klebsiella pneumoniae; D008050:Lipid A / Q000032:analysis*; D008070:Lipopolysaccharides / Q000032:analysis*; D011112:Polymyxin B / Q000276:immunology; D011550:Pseudomonas aeruginosa
D000911:Antibodies, Monoclonal; D000942:Antigens, Bacterial; D004731:Endotoxins; D008050:Lipid A; D008070:Lipopolysaccharides; D006735:Horseradish Peroxidase; D011112:Polymyxin B