Nanoparticle Tracking Analysis (NTA)
Nanoparticle Tracking Analysis (NTA) is a unique method of visualising and analysing particles in liquids, from 10-2000nm, that relates the rate of Brownian motion to particle size. The rate of movement is related only to the viscosity of the liquid, the temperature and size of the particle and is not influenced by particle density or refractive index.
The particles contained in the sample are visualised by virtue of the light they scatter when illuminated by laser light. Typical analysis time is approximately 30s however longer analysis times improve accuracy and statistics.
The light scattered by the particles is captured using a scientific digital camera and the motion of each particle is tracked from frame to frame by the specially developed software. This rate of particle movement is related to a sphere equivalent hydrodynamic radius as calculated through the Stokes-Einstein equation. The technique calculates particle size on a particle-by particle basis overcoming inherent weaknesses in ensemble techniques. Also, since video clips form the basis of the analysis, accurate characterisation of real time events such as aggregation and dissolution is possible.
Measuring Concentration with Nanoparticle Tracking Analysis
NanoSight’s unique single particle detection system allows particle concentration to be measured in the 10-2000nm range in liquid suspension. The ability to measure concentration and apply this to product performance, goes well beyond simply measuring particle size and provides the user with a much fuller understanding of their sample. Measuring particle concentration is a requirement across a range of applications, please see the Applications section above for a more detailed description.
The NanoSight technique provides high resolution number distributions which play a pivotal role in understanding the kinetics of aggregation. Absolute numbers of particles can be measured and the relative number of monomer versus aggregated particles is calculated in real time. Understanding aggregation and the kinetics of the reaction is a ubiquitous requirement across a huge range of applications from Drug Delivery and Vaccine Preparation to CMP slurries and Protein Aggregation.
The NanoSight NTA Software can be used to set up a time course experiment and the kinetics of aggregation, studied in real time. Aggregation can be monitored in three ways; what does the sample look like from a qualitative point of view, what is the change in the particle size with time, and also what is the change in the particle concentration as the sample aggregates with respect to time.
Whole Virus Vaccine
The NanoSight technology can measure ‘Total Viral Titer’ in minutes. The total viral titer is termed as the number of infectious and non-infectious viruses within a preparation. It is often critical to be able to measure the total number of viruses and compare this to the infectious titer as measured by infectivity assays for example. It is often the case that only 1 in 100 viruses may be infectious and by monitoring this ratio it gives the manufacturer the ability to optimize the purification process.
Through fluorescent labeling it is also possible to discriminate target particles in a complex unpurified sample.
Virus Like Particles
Virus Like Particles (VLPs) are neither infectious nor contain nucleic acid and hence cannot be measured by qPCR or infectivity assays. The NanoSight technology can measure the particle size, degree of sample aggregation and particle concentration of a VLP preparation, which is crucial information when understanding the clinical performance of the product and in optimizing the production processes.
Exosomes and microvesicles are secreted by a wide range of cell types. They contain numerous membrane bound protein indicative of their cellular origin and may also contain microRNA, they have been implicated in the progression of a wide range of disease types (cancer, neurological disease, heart disease, preeclampsia, diabetes etc) and are thought to be produced at early stages in these diseases and at elevated concentrations. Accordingly, their role as potential biodiagnostic markers of such diseases is the subject of intense research. Given their size, detection by conventional methodologies such as Flow Cytometry is not possible, making NanoSight’s unique methodology the current instrument of choice for detection and measurement of both particle concentration and size. Fluorescent labeling of specific biomarkers allows phenotyping of exosomes and measurement of exosomes relating to a specific condition.
Measuring the concentration of liposomes, polymeric nanoparticles and other delivery vehicles is a key parameter in understanding the performance of a drug delivery system. The particle size will determine the distribution of the particles within the body and the concentration and size of the particles will govern the amount of drug delivered. NanoSight’s NTA delivers number weighted data through particle-by-particle analysis of the whole distribution and is especially applicable to polydisperse and complex samples.