Multicolor Imaging from Bacteria to 3D Cellular Models Using Carbon Quantum Dots

Bacterial and cellular imaging at the outset allows us to visualize microbial and cellular behavior and how they interact and respond to their surroundings. However, developing biocompatible and photostable fluorescent probes for visualizing and tracking biological processes in living systems remains a significant challenge. Conventional staining agents, particularly organic dyes, are limited by photobleaching, cytotoxicity, and reduced performance in complex biological environments. In this work, we present citric-acid-based carbon quantum dots (CQDs) as a versatile fluorescent probe capable of enabling multicolor bioimaging across diverse biological systems. The CQDs exhibit strong and tunable photoluminescence, excitation-dependent emission, and high photostability, allowing sustained imaging without signal deterioration. Their performance was validated in both Gram-positive and Gram-negative bacteria, mammalian cells such as stem cells, muscle cells, and cancerous cells, demonstrating efficient uptake and bright fluorescence. Importantly, these CQDs enable multicolor imaging under different excitation wavelengths, eliminating the need for multiple fluorophores. Beyond conventional imaging, they facilitate simultaneous visualization of bacteria-infected mammalian cells in both live and fixed conditions and show effective penetration and staining in three-dimensional spheroid models. The ability to reliably stain and image diverse biological systems, from bacteria to complex 3D cellular structures, underscores the distinct advantage of CQDs over conventional probes. This work thus highlights their potential as a cost-effective, stable, and biocompatible tool for advanced bioimaging, intracellular tracking, and diagnostic applications.