Nanocapillary electrokinetic system for particle tracking / counting and microscopy
Tracking the motion of single nanoparticles in liquid solution is a gateway to high accuracy particle counting as well as to understanding and monitoring physical, chemical, and biological processes at the nanoscale. This technology has been recently demonstrated to carry out high-speed tracking of nanoparticles and macromolecules using elastic light scattering.
The weak scattering of single small viruses (26 nm) was successfully detected. For the first time, their fast thermal diffusion was tracked at a frame rate of more than 2 kHz (see Figure 1). As a step forward towards clinical applications, single urinary vesicles as small as 35 nm were also tracked by elastic light scattering (the first successful attempt of detecting biological vesicles that are smaller 70 nm in freely diffusing suspension). These vesicles possess low-refractive index (n<1.4), as confirmed by comparing their thermal diffusion and light scattering cross section.
Figure 1: a. Schematic representation of our single nanoparticle/virus tracking setup. b. CCMV virus that we have tracked. c. The nanofluidic access of single-mode optical fiber. d. Quasi-1D tracking of a single CCMV virus.
This particle-tracking system embeds a silica-based single-mode optical fibre with a hollow-core (nanometer scale) to suppress the free-diffusion of single nanoparticles and direct them into the detection volume. When light is coupled to the nanoparticle-filled optical fiber and detection is performed with a microscope lens at a right angle to the guided illuminating light, the untethered motion of nanoparticles can be imaged and tracked in a quasi-1D geometry for a virtually unlimited duration with negligible disturbance. For details see ACS Nano, 9 (12), 12349–12357, Open Access.
- Platform capable of detecting, tracking, counting, and measuring single nanoparticles, vesicles, and biomolecules.
- It uses a unique (patented) step-index hollow core fiber to propagate light without distorting the particle image (unlike other structured hollow core fibers).
- This method does not involve any nanofabrication processes, which makes it affordable to any research labs or diagnostic labs.
- Consisting of a plug and play cartridge along with a nanofluidic optical platform, the system does not require any modification of existing microscopy setup. Hence, it is a true add-on to standard optical microscopes.
- The platform will enable measurements at single nanoparticle level, electrophoretic separation, and studying diffusion and aggregation dynamics.
- This method does not require any fluorescent labels. Hence, it will be a label-free method.
Figure 2: The portable stage setup. The green outlined portion is the fibre cartridge is magnetically attached to the stage setup. The cartridge is plug-and--play, which means it detachable or mountable without any mechanical tools. The red masked regions are part of a conventional optical microscope.
Ensemble scattering-based methods such as diffuse wave spectroscopy and dynamic light scattering spectroscopy have proven to meet the demand of a large market.
The current particle sizing methods are based on ensemble measurements. They are fundamentally prone to the uncertainties caused by inhomogeneous size distributions and stochastic phase-differences. Since dynamic processes are triggered stochastically, ensemble measurements cannot resolve fast processes that are washed out by averaging. The outcomes of such ensemble measurements are dominated by the slowest processes due to averaging of asynchronous processes.
One interesting and fast-growing field of application whereby this novel method can make a difference is the one of neurodegenerative diseases. The World Alzheimer Report of 2015 states that the current expenditure in the USA alone for dementia is USD 818 billion. By 2018 this cost will rise to USD 1 trillion. Currently, biochemical assays being used in any diagnostics are based on ensemble averaging experiments. However, neurodegenerative diseases are primarily caused by protein misfolding. The complexity of protein folding process is difficult or impossible to study using the available ensemble methods. Ensemble averaging experiments conceal molecular interactions, and thereby, early biomolecular interactions responsible for diseases, such as Alzheimer’s, Parkinson’s, prion diseases, and amyotrophic lateral sclerosis remain undetected. This limitation is overcome by this method, whereby the capability to track sub-micron particle individually is crucial to track the evolution of protein folding and misfolding.
Another area of biomedical research where particle tracking is in high demand is related to extracellular vesicles which are found in human body fluids. The scientific research on that is exponentially increasing every year (see Figure 3).
Figure 3: An yearly exponential behaviour of number of scientific outcome (papers) on small biological extracellular vesicles (Edwin van der Pol, PhD Thesis, Amsterdam Medical Center).
Nanoparticle tracking has proven to be a viable technological market, too. This system can characterize nanoparticles from 10 nm to 2000 nm in solution, and allows for single particle tracking. However, trajectories being short, it gathers statistics by collecting many tracks, through which particle size distribution and concentration can be derived. This has an immediate impact on studying protein aggregation related neurodegenerative diseases. The primary advantage over existing alternative methods is that this system can resolve pure single nanoparticle level dynamics.
Prospective users of this technology can be from the following sectors:
- Biomolecular research
- Biomedical diagnostics
- Colloid chemistry
- Environment pollution
- Colloid chemistry
- Chemical identification
This invention is a user friendly, plug-and-play add-on device for conventional optical microscope or method to tackle biologically relevant questions based on detecting single nanoparticles. The name of this device is ‘nanoCET’ (nano Capillary Electrophoretic Tracking) and encompasses a cartridge and a stage. Keeping the needs of the biomedical research into account, the nanoCET cartridge will be disposable and cost-effective, and can be detachable from the nanoCET stage. The nanoCET stage is an add-on the conventional microscope, which can be rented, leased, and purchased as one-time investment. Interested parties can perform test experiments in the inventor’s lab with the resident existing setups.
Luris reference numberINV-028.048
Patent rights are already granted in the UK and being acquired in the EU and in the USA.