Development of biofunctional ruby nanoparticles for optical imaging of opioid receptors

There is an increasing demand for high performance photoluminescent nanoparticles in the life sciences, where the targeted labelling and optical imaging of biomolecules are needed. Photostability, low environmental sensitivity and narrow spectra make nanoparticles superior over existing molecular fluorophores. Ruby nanocrystals, termed nanorubies, represent an excellent example, especially their long photoluminescence lifetime allowing dramatic improvement of the optical contrast. However, premature surface functionalisation procedures have hindered the widespread use of nanorubies. To address this challenge, the first aim was to develop facile and reproducible protocols to functionalise nanoruby with biomolecules, specifically, NeutrAvidin. The second aim was to demonstrate the use of developed biofunctional nanorubies to label opioid receptors in live and fixed cells, followed by high-sensitivity optical imaging. Silica nanoparticles formed a test-bed to develop conjugation methods based on amide and click chemistries. Biotin-binding assays and receptor labelling studies showed that click chemistry was most specific, efficient, reproducible and controllable. Amide-based chemistries, in comparison, resulted in non-specific binding and low-contrast imaging. The developed click chemistry protocol, when extended to silica-coated nanorubies, allowed specific labelling and high-sensitivity imaging of opioid receptors. This study enables the wide use of nanoruby for ultrasensitive and real-time imaging of a broad class of receptor-ligand systems.