A sensor for the lethal bacterial enzyme， botulinum neurotoxin type A (BoNT/A)， was developed using self-assembled monolayers (SAMs). SAMs consisting of an immobilized synthetic peptide that mimicked the toxin's in vivo SNAP-25 protein substrate were formed on Au and interfaced with arrayed microfluidic channels. Efforts to optimize SAM composition and assay conditions for greatest reaction efficiency and sensitivity are described in detail. Channel design provided facile fluid manipulation， sample incubation， analyte concentration， and fluorescence detection all within a single microfluidic channel， thus avoiding sample transfer and loss. Peptide SAMs were exposed to varying concentrations of BoNT/... More
A sensor for the lethal bacterial enzyme， botulinum neurotoxin type A (BoNT/A)， was developed using self-assembled monolayers (SAMs). SAMs consisting of an immobilized synthetic peptide that mimicked the toxin's in vivo SNAP-25 protein substrate were formed on Au and interfaced with arrayed microfluidic channels. Efforts to optimize SAM composition and assay conditions for greatest reaction efficiency and sensitivity are described in detail. Channel design provided facile fluid manipulation， sample incubation， analyte concentration， and fluorescence detection all within a single microfluidic channel， thus avoiding sample transfer and loss. Peptide SAMs were exposed to varying concentrations of BoNT/A or its catalytic light chain (ALC)， resulting in enzymatic cleavage of the peptide substrate from the surface. Fluorescence detection was achieved down to 20 pg/mL ALC and 3 pg/mL BoNT/A in 3 h. Toxin sensing was also accomplished in vegetable soup， demonstrating practicality of the method. The modular design of this microfluidic SAM platform allows for extension to sensing other toxins that operate via enzymatic cleavage， such as the remaining BoNT serotypes B-G， anthrax， and tetanus toxin.
