Development of Non-Metallic Quantum Dots for Bioimaging and Biosensing

Abstract

Biocompatible and photostable fluorescent probes are crucial yet challenging to develop for visualizing and tracking biological functions and interactions that occur in living organisms. We synthesized biocompatible carbon quantum dots (CQDs) with 15% quantum yield (QY) and tested them for cellular and scaffold imaging at variable depths. The CQDs, synthesized from collagen under controlled and benign conditions in a hydrothermal reactor, were characterized for their fundamental physicochemical properties. The fluorescence characteristics were determined using two-photon microscopy and based on our results we propose a mechanism for CQD luminescence by combining the carbogenic core and edge-effect contributions to the photoluminescent (PL) behavior. The bioimaging of cells embedded in a luminescent 3D printed scaffold showed that CQDs enable imaging at depths of about 1500 μm under biomimetic conditions. Real-time videography and imaging tests showed a differential visualization of individual cells and scaffold. The excellent photostability and non-photobleaching characteristics of CQDs make them suitable for long-term whole cell and tissue imaging via multi-photon microscopy. Nanoparticles are key vehicles for targeted therapies because they can pass through biological barriers, enter into cells, and distribute within cell structures. We investigated the synthesis of blue and green emissive hexagonal boron nitride quantum dots (hBNQDs) using a liquid-exfoliation technique followed by hydrothermal treatment. A distinct shift from blue to bright-green emission was observed upon surface passivating the dots using poly (ethylene glycol) or PEG200 under the same UV irradiation. The quantum yield of the hBNQDs increased with the surface passivation. Multiplexed imaging was accomplished using the hBNQDs in conjunction with organic dyes. The hBNQDs provided images with distinctive emission wavelengths and fluorescence lifetimes. Although the fluorescence signals of blue- and green-emissive hBNQDs overlap spectrally with those of the emission wavelengths of the organic dyes, the fluorescence lifetime data were resolved temporally using software-based time gates. The blue-emissive hBNQD-b quantum dots were validated as sensitive platforms for detecting intracellular ferric ions with a low limit of detection (20.6 nM). The green-emissive hBNQD-g quantum dots successfully identified intracellular variations in pH, and the localization in human breast cancer cells was determined during their life cycles via fluorescence lifetime imaging. Sensitive and selective detection of Fe3+ merits attention since its deficiency can cause significant physiological dysfunction. Herein, we explored the interplay of synthesis parameters and size of the Spirulina derived carbon dots to optimize and develop a bright (51% quantum yield), selective and ultra-sensitive sensor to detect variations in intracellular Fe3+ ion concentrations. The final product showed a lower detection limit of 380pM and a response time of only 30 seconds. Several spectroscopic methods were used to elucidate the fluorescence and quenching mechanism of the carbon dots (CDs). Fluorescence lifetime measurements and Stern-Volmer analysis revealed that both static and dynamic quenching processes are dominant at low and high concentrations of Fe3+ ions respectively. The developed CDs were successfully applied to track the dynamic generation of endogenous Fe3+ in living cells under stress induced conditions.