Real-Time Monitoring of a Botulinum Neurotoxin Using All-Carbon Nanotube-Based Field-Effect Transistor Devices

The possibility of exposure to botulinum neurotoxin (BoNT), a powerful and potential bioterrorism agent, is considered to be ever increasing. The current gold-standard assay, live-mouse lethality, exhibits high sensitivity but has limitations including long assay times, whereas other assays evince rapidity but lack factors such as real-time monitoring or portability. In this study, we aimed to devise a novel detection system that could detect BoNT at below-nanomolar concentrations in the form of a stretchable biosensor. We used a field-effect transistor with a p-type channel and electrodes, along with a channel comprising aligned carbon nanotube layers to detect the type E light chain of BoNT (BoNT/E-Lc). The detection of BoNT/E-Lc entailed observing the cleavage of a unique peptide and the specific bonding between BoNT/E-Lc and antibody BoNT/E-Lc (Anti-BoNT/E-Lc). The unique peptide was cleaved by 60 pM BoNT/E-Lc; notably, 52 fM BoNT/E-Lc was detected within 1 min in the device with the antibody in the bent state. These results demonstrated that an all-carbon nanotube-based device (all-CNT-based device) could be produced without a complicated fabrication process and could be used as a biosensor with high sensitivity, suggesting its potential development as a wearable BoNT biosensor.

[1]  Seungwoo Ham,et al.  Effect of oxygen plasma treatment on carbon nanotube-based sensors. , 2014, Journal of nanoscience and nanotechnology.

[2]  D. Gill,et al.  Bacterial toxins: a table of lethal amounts , 1982, Microbiological reviews.

[3]  Lin Kang,et al.  An Ultrasensitive Gold Nanoparticle-based Lateral Flow Test for the Detection of Active Botulinum Neurotoxin Type A , 2017, Nanoscale Research Letters.

[4]  J. Rogers,et al.  Guided growth of large-scale, horizontally aligned arrays of single-walled carbon nanotubes and their use in thin-film transistors. , 2005, Small.

[5]  S. Nauenburg,et al.  Proteolysis of SNAP‐25 Isoforms by Botulinum Neurotoxin Types A, C, and E , 1999, Journal of neurochemistry.

[6]  Chao Li,et al.  Complementary detection of prostate-specific antigen using In2O3 nanowires and carbon nanotubes. , 2005, Journal of the American Chemical Society.

[7]  M. Prato,et al.  Chemistry of carbon nanotubes. , 2006, Chemical reviews.

[8]  Yanyan Tang,et al.  Label-free study of the function of ion channel protein on a microfluidic optical sensor integrated with artificial cell membrane. , 2014, Lab on a chip.

[9]  Frank Gessler,et al.  Evaluation of lateral flow assays for the detection of botulinum neurotoxin type A and their application in laboratory diagnosis of botulism. , 2007, Diagnostic microbiology and infectious disease.

[10]  Rong-Hwa Shyu,et al.  Monoclonal antibody-based lateral flow assay for detection of botulinum neurotoxin type A. , 2008, Hybridoma.

[11]  Kenzo Maehashi,et al.  Label-free protein biosensor based on aptamer-modified carbon nanotube field-effect transistors. , 2007, Analytical chemistry.

[12]  G. Schiavo,et al.  Tetanus and botulism neurotoxins: a new group of zinc proteases. , 1993, Trends in biochemical sciences.

[13]  Suzanne R. Kalb,et al.  Mass Spectrometric Detection of Bacterial Protein Toxins and Their Enzymatic Activity , 2015, Toxins.

[14]  Ji-Joon Song,et al.  Facile electrochemical detection of botulinum neurotoxin type E using a two-step proteolytic cleavage. , 2015, Biosensors & bioelectronics.

[15]  Bo Liedberg,et al.  Highly manufacturable graphene oxide biosensor for sensitive Interleukin-6 detection , 2015 .

[16]  Eun Sung Kim,et al.  CRITERIA FOR PRODUCING YARNS FROM VERTICALLY ALIGNED CARBON NANOTUBES , 2010 .

[17]  Edwin R Chapman,et al.  Using fluorescent sensors to detect botulinum neurotoxin activity in vitro and in living cells. , 2004, Proceedings of the National Academy of Sciences of the United States of America.

[18]  E J Schantz,et al.  Properties and use of botulinum toxin and other microbial neurotoxins in medicine. , 1992, Microbiological reviews.

[19]  R. C. Whiting,et al.  Detection of Type A, B, E, and F Clostridium botulinum Neurotoxins in Foods by Using an Amplified Enzyme-Linked Immunosorbent Assay with Digoxigenin-Labeled Antibodies , 2006, Applied and Environmental Microbiology.

[20]  Lianxi Zheng,et al.  Strong carbon-nanotube fibers spun from long carbon-nanotube arrays. , 2007, Small.

[21]  Yi Wang,et al.  A review on electronic bio-sensing approaches based on non-antibody recognition elements. , 2016, The Analyst.

[22]  Philip K. Russell,et al.  Botulinum toxin as a biological weapon: medical and public health management. , 2001, JAMA.

[23]  Suzanne R. Kalb,et al.  Optimization of peptide substrates for botulinum neurotoxin E improves detection sensitivity in the Endopep-MS assay. , 2015, Analytical biochemistry.

[24]  R. Mizanur,et al.  Cleavage of SNAP25 and Its Shorter Versions by the Protease Domain of Serotype A Botulinum Neurotoxin , 2014, PloS one.

[25]  Christian Lévêque,et al.  A substrate sensor chip to assay the enzymatic activity of Botulinum neurotoxin A. , 2013, Biosensors & bioelectronics.

[26]  H. Adams,et al.  Neurological aspects of biological and chemical terrorism: a review for neurologists. , 2003, Archives of neurology.

[27]  M. H. van der Veen,et al.  Bandgap opening in oxygen plasma-treated graphene , 2010, Nanotechnology.

[28]  Larry H. Stanker,et al.  Rapid Microfluidic Assay for the Detection of Botulinum Neurotoxin in Animal Sera , 2016, Toxins.

[29]  G. Sakaguchi Clostridium botulinum toxins. , 1982, Pharmacology & therapeutics.

[30]  Gengfeng Zheng,et al.  Multiplexed electrical detection of cancer markers with nanowire sensor arrays , 2005, Nature Biotechnology.

[31]  C. Shone,et al.  Monoclonal antibody-based immunoassay for type A Clostridium botulinum toxin is comparable to the mouse bioassay , 1985, Applied and environmental microbiology.

[32]  SinghAjay,et al.  Evaluation of an enzyme-linked immunosorbent assay (ELISA) kit for the detection of botulinum neurotoxins A, B, E, and F in selected food matrices. , 2015 .

[33]  P. Avouris,et al.  Engineering Carbon Nanotubes and Nanotube Circuits Using Electrical Breakdown , 2001, Science.

[34]  Randy F. Stout,et al.  Nanopore Sensing of Botulinum Toxin Type B by Discriminating an Enzymatically Cleaved Peptide from a Synaptic Protein Synaptobrevin 2 Derivative , 2014, ACS applied materials & interfaces.