Fast, Ratiometric FRET from Quantum Dot Conjugated Stabilized Single Chain Variable Fragments for Quantitative Botulinum Neurotoxin Sensing
- Fast, Ratiometric FRET from Quantum Dot Conjugated Stabilized Single Chain Variable Fragments for Quantitative Botulinum Neurotoxin Sensing
- 이준석; Melissa B Brennan; Rosemarie Wilton; Clare E Rowland; Elena A Rozhkova; Sara Forrester; Daniel C Hannah; Julia Carlson; Elena V Shevchenko; Daniel S Schabacker; Richard D Schaller
- Issue Date
- Nano letters
- VOL 15-7167
- Botulinum neurotoxin (BoNT) presents a significant hazard under numerous realistic scenarios. The standard detection scheme for this fast-acting toxin is a labbased mouse lethality assay that is sensitive and specific, but slow (∼2 days) and requires expert administration. As such, numerous efforts have aimed to decrease analysis time and reduce complexity. Here, we describe a sensitive ratiometric fluorescence resonance energy transfer scheme that utilizes
highly photostable semiconductor quantum dot (QD) energy donors and chromophore conjugation to compact, single chain variable antibody fragments (scFvs) to yield a fast, fieldable sensor for BoNT with a 20− 40 pM detection limit, toxin quantification, adjustable dynamic range, sensitivity in the presence of interferents, and sensing times as fast as 5 min. Through a
combination of mutations, we achieve stabilized scFv denaturation temperatures of more than 60 °C, which bolsters fieldability. We also describe adaptation of the assay into a microarray format that offers persistent monitoring, reuse, and multiplexing.
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