Single-Molecule Detection of H₂O₂ Mediating Angiogenic Redox Signaling on Fluorescent Single-Walled Carbon Nanotube Array

Reactive oxygen species, specifically hydrogen peroxide (H(2)O(2)), activate signal transduction pathways during angiogenesis and therefore play an important role in physiological development as well as various pathophysiologies. Herein, we utilize a near-infrared fluorescent single-walled carbon nanotube (SWNT) sensor array to measure the single-molecule efflux of H(2)O(2) from human umbilical vein endothelial cells (HUVEC) in response to angiogenic stimulation. Two angiogenic agents were investigated: the pro-angiogenic cytokine, vascular endothelial growth factor A (VEGF-A) and the recently identified inorganic pro-angiogenic factor, europium(III) hydroxide in nanorod form. The nanosensor array consists ofa SWNT embedded within a collagen matrix that exhibits high selectivity and sensitivity to single molecules of H(2)O(2). A calibration from 12.5 to 400 nM quantifies the production of H(2)O(2) at nanomolar concentration in HUVEC with 1 s temporal and 300 nm spatial resolutions. We find that the production of H(2)O(2) following VEGF stimulation is elevated outside of HUVEC, but not for stimulation via nanorods, while increased generation is observed in the cytoplasm for both cases, suggesting two distinct signaling pathways.