Optimization of immobilized bacterial disaccharides for surface plasmon resonance imaging measurements of antibody binding.
The interactions between proteins and immobilized carbohydrates are crucial to biological events such as cell signaling and immune response. The modification of surfaces with carbohydrates to create sensing platforms provides a pathway to study these interactions in a laboratory setting. In this work, a family of structurally related Salmonella disaccharide epitopes is immobilized on thin gold films in an array format to probe antibody binding with surface plasmon resonance (SPR) imaging. The disaccharides are modified with an alkyl thiol linker for facile immobilization to gold. Small differences in the stereochemistry of the immobilized, modified disaccharides are shown to greatly influence the binding of a monoclonal antibody. Specifically, binding is only observed to an immobilized abequose dideoxyhexose relative to a tyvelose or a paratose analogue. However, both the amount and relative strength of bound antibody depends on the distribution of disaccharide moieties in a mixed monolayer of the epitope and a nonbinding diluent molecule. We thoroughly characterize the mixed monolayers with a variety of techniques to understand the optimal density and distribution of the disaccharide for antibody capture. This work reinforces the importance of controlling the density of ligands at the interface for optimized surface based bioassays.
Grant
Chris F
CF
Department of Chemistry and National Institute for Nanotechnology, University of Alberta, Edmonton, AB, Canada.
Kanda
Vishal
V
Yu
Henry
H
Bundle
David R
DR
McDermott
Mark T
MT
D000906:Antibodies / Q000276:immunology*; D000915:Antibody Affinity; D004187:Disaccharides / Q000737:chemistry*; D000939:Epitopes / Q000276:immunology*; D005233:Fatty Alcohols / Q000737:chemistry; D015394:Molecular Structure; D012475:Salmonella / Q000737:chemistry*; D013237:Stereoisomerism; D013438:Sulfhydryl Compounds / Q000737:chemistry; D020349:Surface Plasmon Resonance / Q000379:methods*
C521238:11-mercaptoundecanol; D000906:Antibodies; D004187:Disaccharides; D000939:Epitopes; D005233:Fatty Alcohols; D013438:Sulfhydryl Compounds