Wide-field imaging and flow cytometric analysis of cancer cells in blood by fluorescent nanodiamond labeling and time gating

Nanodiamonds containing high density ensembles of negatively charged nitrogen-vacancy (NV-) centers are promising fluorescent biomarkers due to their excellent photostability and biocompatibility. The NV-?' centers in the particles have a fluorescence lifetime of up to 20ns ...ns, which distinctly differs from those (<10 ...ns) of cell and tissue autofluorescence, making it possible to achieve background-free detection in vivo by time gating. Here, we demonstrate the feasibility of using fluorescent nanodiamonds (FNDs) as optical labels for wide-field time-gated fluorescence imaging and flow cytometric analysis of cancer cells with a nanosecond intensified charge-coupled device (ICCD) as the detector. The combined technique has allowed us to acquire fluorescence images of FND-labeled HeLa cells in whole blood covered with a chicken breast of ?0.1-mm thickness at the single cell level, and to detect individual FND-labeled HeLa cells in blood flowing through a microfluidic device at a frame rate of 23 ...Hz, as well as to locate and trace FND-labeled lung cancer cells in the blood vessels of a mouse ear. It opens a new window for real-time imaging and tracking of transplanted cells (such as stem cells) in vivo.