Fluorescence labeling can be utilized in any situation where it is necessary to distinguish a particular subset of particles within a complex background. Fluorescent nanoparticles can be measured in terms of particle size and concentration to understand the nature of those particles in complex media such as blood serum which may induce particle aggregation. This has applications in understanding the fate of drug delivery vectors as well as understanding the toxiciology of nanoparticles in biological environments for example. Alternatively, fluorescently labeled antibodies may be used to specifically bind to target particles within a sample thus providing a mechanism to identify specific particles within an unknown sample. This has implications in diagnostic applications such as research into exosomes and microvesicles in that specific markers can be identified and monitored in the detection of disease.
The NanoSight range of instruments can operate in light scatter mode where all particles within the instrument are visualized and measured (and thus does not require the sample to be fluorescently labeled to take a measurement) or alternatively the instrument can also operate in fluorescence mode where only fluorescent nanoparticles are measured.
The NanoSight system uses either a 405 nm (violet), 488 nm (blue), 642 nm (red) or 532 nm (green) laser source to excite suitable fluorophores whose fluorescence can then be determined using matched long or short-pass filters.
In the following example an approx. 50:50 mixture of fluorescently labeled (Fluoresbrite) 100 nm and unlabeled 100 nm polystyrene beads were analyzed under light scatter mode (red line and top image) and when fluorescently filtered (white line and bottom image).