Conjugation of biotin-coated luminescent quantum dots with single domain antibody-rhizavidin fusions

• Single domain antibody—rhizavidin fusion bioconjugated biotin coated quantum dots.
• Provided facile and effective method to orient antibodies on QD surface.
• Accomplished improved production of His-tagged rhizavidin (RZh) in E. coli.
• Demonstrated utility of RZh as a replacement for tetrameric biotin binders.

Straightforward and effective methods are required for the bioconjugation of proteins to surfaces and particles. Previously we demonstrated that the fusion of a single domain antibody with the biotin binding molecule rhizavidin provided a facile method to coat biotin-modified surfaces with a highly active and oriented antibody. Here, we constructed similar single domain antibody—rhizavidin fusions as well as unfused rhizavidin with a His-tag. The unfused rhizavidin produced efficiently and its utility for assay development was demonstrated in surface plasmon resonance experiments. The single domain antibody-rhizavidin fusions were utilized to coat quantum dots that had been prepared with surface biotins. Preparation of antibody coated quantum dots by this means was found to be both easy and effective. The prepared single domain antibody-quantum dot reagent was characterized by surface plasmon resonance and applied to toxin detection in a fluoroimmunoassay sensing format.

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