(558a) Single Walled Carbon Nanotubes as Single Molecule Chemical Sensors within Living Cells

Chemical agents which damage or bind to nucleic acids are under continuous study for prediction, diagnosis, and treatment of cancer, Alzheimer's disease, and the effects of aging.1,2 Their activity is difficult to measure in real time and in vivo, however, preventing many types of studies on chemotherapeutic agent activity or oxidative stress.3,4 We demonstrate that single-walled carbon nanotubes (SWNT), hollow graphene cylinders with diameters between 0.5 and 2 nm, optically transduce subtle conformational changes in synthetic DNA via spectral changes in near-infrared fluorescence.5,6 We show that a pair of carbon nanotubes can uniquely detect and identify agents of DNA damage in real time within live mammalian cells. Nucleic acid encapsulated nanotube near-infrared photoluminescence is modified upon chemotherapeutic alkylation and reactive oxygen species (ROS) activity via spectral shifts of up to 30 meV. Singlet oxygen, the hydroxyl radical, and alkylating drugs such as melphalan and cisplatin induce differentiating spectral changes between two carbon nanotube species which elucidates the agent involved. These differences are detected from within live mammalian cells. This finding demonstrates the first multiplexed optical detection from a nanoscale sensor. The work provides the first tool to measure alkylating and ROS activity in real-time in living tissue due to the unique photostability and non-invasive near-infrared emission of carbon nanotubes.

This platform has several advantages for fundamental studies of the cellular uptake of nanomaterials. Single-walled carbon nanotubes (SWNT) fluoresce in the near-infrared (n-IR) and are one of a very few classes of fluorophores with no observable photobleaching when excited at moderate fluence. We use this property to track over 10,288 individual trajectories as particles are incorporated into and expelled from NIH-3T3 cells over a period of 127 min on a perfusion microscope stage. A total of 5223 particle trajectories (49.2%) exhibit purely convective diffusion without cellular interaction in the applied flow field. The remaining 5065 trajectories (50.8%) show distinct processes of cellular membrane adsorption (6.2%), surface diffusion (18.4%), endocytosis (12.7%), exocytosis (5.9%) or membrane desorption (7.4%). An analysis of each mean square displacement allows for the first time the complete construction of the network of pathways experienced by nanoparticles as they are trafficked into and within the cell. We observe the first conclusive evidence of nanoparticle exocytosis in this system, and show that the rate closely matches the endocytosis rate with negligible temporal offset. We identify and study a unique pathway that leads to the previously observed aggregation and accumulation of SWNT within the cells. The results have significant implications for the use of nanoparticles in biological systems.

1. Shi, H. L., Hudson, L. G. & Liu, K. J. Oxidative stress and apoptosis in metal ion-induced carcinogenesis. Free Radical Biology and Medicine 37, 582-593 (2004).

2. Povirk, L. F. & Shuker, D. E. DNA-Damage and Mutagenesis Induced by Nitrogen Mustards. Mutation Research-Reviews in Genetic Toxicology 318, 205-226 (1994).

3. Berliner, L. J., Khramtsov, V., Fujii, H. & Clanton, T. L. Unique in vivo applications of spin traps. Free Radical Biology and Medicine 30, 489-499 (2001).

4. Brandes, R. P. & Janiszewski, M. Direct detection of reactive oxygen species ex vivo. Kidney International 67, 1662-1664 (2005).

5. Saito, R., Dresselhaus, G. & Dresselhaus, M. S. Physical Properties of Carbon Nanotubes (Imperial College Press, London, 1998).

6. Heller, D. A. et al. Optical detection of DNA conformational polymorphism on single-walled carbon nanotubes. Science 311, 508-511 (2006).