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Multivalent nano-robot sensors for counting of Covid-19 coronavirus particles in aerosols

Molecular and Cell Biology

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Dr Andrey Revyakin No more applications being accepted Competition Funded PhD Project (European/UK Students Only)

About the Project

Airborne transmission of CoV-19 is believed to play a significant role in the ongoing pandemic. Recent studies indicate that a significant fraction (or the majority) of shed viral RNA is found in fine aerosol particles. In addition, 20-50% of outbreaks are shown to originate from asymptomatic (i.e. cough-free) persons, and that infected individuals typically shed ~10E4-10E6 RNA genomes per hour. At normal breathing rates, the majority of particles emitted by a non-coughing person falls into the fine aerosol range (<1 micron diameter), produced at a rate of 10E4-10E5 per hour. Such small particles remain airborne for hours and cause infection despite social distancing. A real-time bio-specific CoV-19 virion counter, if deployed at a specific location (e.g. in crowded indoor workspaces) would evaluate the current risk of infection before the infection has taken place in a non-invasive manner (similar to a Geiger counter). However, state-of-the-art aerosol sampling methods do not permit real-time (results in <1 hr) counting of virions in aerosols

In this project, we will develop a new type of fluorescent nanoprobes (“nanorobots”) that rapidly and specifically bind to multi-valent targets, such as the ‘corona’ of CoV-19. The nanoprobes will be an integral part of a prototype device which we call the Coronavirus Aerosol Monitor (CAM). In CAM, indoor air would be pumped through a thin, transparent filter membrane comprised of ultra-fine (~100 nm) synthetic nano-fibers. The membrane would then be ‘developed’ with the nanorobots which bind to single virions captured in the membrane and make the virions fluorescent. This, in turn, would enable counting of single virions using a low-cost digital microscope (integrated, for instance, with a smartphone). The work will be carried out as a collaboration between the Leicester Institute for Chemical Biology (LISCB), the Leicester School of Engineering, and the Health & Safety Executive Laboratory (HSE).

In Year 1, we will be make nanorobot probes featuring: (i) the ability to specifically interact with CoV-19 particles via multi-valent interactions; (ii) super-bright fluorescence to enable counting of virions using a low-cost microscope; and (iii) modular design for rapid adaptation towards future pandemics. To this end, we will use the “DNA origami” technology which enables making 3D objects 20-50 nm in size rapidly and cost-effectively (in comparison, a typical CoV virion is ~80-120 nm in diameter). In DNA origami, a long ssDNA molecule (‘scaffold’) is folded into the desired 3D shape by annealing together with ~250 short ssDNA fragments (‘staples’). We will explore several nanorobot designs, beginning with a simple 17 nm nanocube functionalized with the ectodomain of the human receptor protein ACE2 (the key binding partner for the “Spike” protein of the CoV-19 “corona”). The nanorobots will be characterized via Transmission-Electron and Cryo-Electron Microscopy.

In Year 2, we will use biochemical and biophysical (single-molecule fluorescence imaging) methods to measure the affinity and specificity of the ACE2-nanorobots. As a model target for the nanorobots, we will employ decoy (non-infectious) particles equipped with recombinant “Spike” proteins and, at later stages, genome-free Virus-Like Particles (VLPs). These measurements will allow us to further optimize the nanorobots’ affinity and specificity.

In Year 3, we will create a prototypical CAM device in which the microscope slide (from Year 2) will be replaced with a membrane prepared from ultrafine nanofibers by eletrospinning. In a typical experiment, we will pulverize CoV-19 decoy particles and/or VLPs, and pump them through the nano-membrane, resulting in capture of the particles. Then, we will incubate the nano-membrane with the nanorobots, and will use a single-molecule fluorescence microscope to visualize the particles as bright defined ‘spots’ stained with the nanorobots. These measurements will be used optimize the binding conditions for nanorobots, and the data will be used in future research to scale the CAM prototype up for in real-life measurements of live CoV-19 virions in aerosols

BBSRC Strategic Research Priority: Understanding the Rules of Life: Microbiology & Structural Biology

Techniques that will be undertaken during the project:

DNA origami nanotechnology, DNA purification, protein expression and purification, ultracentrifugation, chemical and photo-chemical conjugation (click-chemistry, UV-crosslinking), electrophoresis, transmission electron microscopy, cryo-electron microscopy, single-molecule fluorescence microscopy, optics, surface chemistry, super-resolution microscopy, electrospinning, rapid prototyping (3D printing), engineering, basic Python and Matlab coding, basic electronics.

The first supervisor for this project is Dr Andrey Revyakin
The second supervisor for this project is Prof Nicholas Brindle

Funding Notes

4 year fully funded BBSRC MIBTP studentship
UK/EU fees and stipend at UKRI rates. For 2020 this will be £15,285 pa

Application is to be submitted via the University of Leicester MIBTP program (click "APPLY MIBTP" at the bottom of the page). The deadline is January 10th, 2021

Please email [Email Address Removed] if you have questions about the application process, or if you would like to inquire about the status of your application

Please email science-specific questions to Dr Andrey Revyakin: [Email Address Removed]
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