Fluorescent microparticles and nanobeads, widely used as molecular tags, are also effective calibrants in fluorescence microscopy. They can be fabricated from a range of materials and come in different sizes, shapes (hollow vs filled) and colours. Their stable properties help to assess key instrument performance characteristics in machinery and microscopes, such as spatial resolution, alignment, and stability. However, as a wide range of fluorescence microscopes and techniques exist, it can be challenging to produce standard materials which are widely suitable.
Stream Bio develop and manufacture bioimaging molecular probes, Conjugated Polymer Nanoparticles (CPNs), for applications ranging from R&D to therapeutic research. They wanted to better understand the performance and stability of their product in order to ensure proof of concept and competitivity in the market.
Stream Bio approached NPL to help test their CPNs using a range of fluorescence microscopy techniques such as confocal laser scanning microscopy (CLSM), super-resolution structured illumination microscopy (SIM) and light sheet microscopy (LSM). A range of Stream Bio’s CPNs, with different spectral properties, were imaged and characterised to assess their key properties including brightness, photostability and uniformity. The data was analysed to assess the suitability of different CPNs for use as calibration tools, with results compared to alternative products currently available.
Super resolution SIM image of CPNs with different spectral properties.
NPL’s quantitative imaging of Stream Bio’s CPNs validated particle uniformity and other physical attributes, such as performance and stability. This work provided Stream Bio with key data and detailed sample preparation protocols for use as calibrants. The work supports the wider use of CPNs as microscopy calibration standards as well as helping further advancements of their product in applications such as cell structure labelling