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Anal Biochem. 1995 Jan 1;224(1):390-4.

Detection of glycoprotein receptors on blotting membranes by binding of live bacteria and amplification by growth.

Author information

1
Phagocyte Research Laboratory, Department of Medical Microbiology and Immunology, Göteborg, Sweden.

Abstract

Conditions have been adapted for detecting bacteria bound to glycoprotein receptors on blotting membranes using a self-enhancing detection method based on bacterial growth. Neutrophil plasma membrane proteins, mediating adherence of mannose-binding type-1-fimbriated Escherichia coli and concanavalin A (Con A) to intact human neutrophils, were separated by SDS-PAGE and transferred onto a polyvinylidene difluoride (PVDF) membrane. The PVDF membrane was immersed in a suspension of mannose-binding type-1-fimbriated E. coli, and after repeated washings, bound bacteria were allowed to multiply into bacterial colonies by placing the membrane on a solid nutrient substratum. About one major and eight minor glycoproteins, some of which also were detected by Con A, selectively induced colony formation in a mannose-inhibitable fashion. Binding of [35S]methionine metabolically labeled E. coli to PVDF membranes produced a virtually identical binding pattern, demonstrating further the accuracy of this self-enhancing detection method which is rapid, simple, and sensitive and avoids radioisotopes.

PMID:
7710097
DOI:
10.1006/abio.1995.1055
[Indexed for MEDLINE]

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