期刊
NANO LETTERS
卷 15, 期 10, 页码 7161-7167出版社
AMER CHEMICAL SOC
DOI: 10.1021/acs.nanolett.5b03442
关键词
botulinum neurotoxin; FRET; protein sensor; scFv; quantum dot; microarray
类别
资金
- U.S. Department of Energy, Office of Basic Energy Sciences [DE-AC02-06CH11357]
- U.S. Department of Energy, Office of Science, Office of Basic Energy Sciences [DE-AC02-06CH11357]
- Argonne National Laboratory Director's Postdoctoral Fellowship
- National Science Foundation Graduate Research Fellowships [DGE-0824162]
Botulinum neurotoxin (BoNT) presents a significant hazard under numerous realistic scenarios. The standard detection scheme for this fast-acting toxin is a lab-based mouse lethality assay that is sensitive and specific, but slow (similar to 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 degrees 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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