Dye-loaded polymeric nanorods as bright fluorescent nano-platforms for efficient cellular uptake and bioimaging

Fluorescent dye-loaded polymeric nanoparticles have been extensively developed for cellular imaging as they are bright and versatile tools with large choice of polymers, dyes and surface chemistry. Here, we report preparation of fluorescent polymeric nanomaterials with high aspect ratio – nanorods, and study their cellular uptake for bioimaging applications. Fluorescent polymeric nanorods are prepared by scission of electrospun nanofibers using ultrasonication. To achieve high brightness, they are loaded with cationic fluorescent dyes (rhodamine or cyanines) with bulky hydrophobic counterions that prevent aggregation-caused quenching. The obtained nanorods of 400 nm diameter and 2-4 µm length show good fluorescence quantum yields (40% at 5% dye loading) and 800-fold higher brightness compared to 60-nm polymeric nanoparticles. Cellular studies reveal several remarkable features of nanorods vs NPs. First, nanorods internalization depends on the cell type, which was not the case of NPs. Second, we found that internalization of nanorods is nearly 100-fold more efficient than that of polymeric NPs, reaching ~3 wt% of the cell mass. The internalization of nanorods is an active process (endocytosis), which is inhibited by low temperature, absence of growth medium or nanorods treatment by a non-ionic surfactant. Despite this efficient internalization, nanorods exhibit negligible cytotoxicity according to three different types of assays. Due to high accumulation inside the cells, the labelled cells present ~100-fold higher fluorescence signal vs NPs, which remains for at least 6 days. This exceptional cell brightness enables single-cell tracking in two near-infrared spectral regions under the chicken skin. This work reveals exceptional capacity of polymeric nanorods to accumulate inside the cells, which allows efficient fluorescence labelling of cells for robust imaging and long-term tracking.