Photoacoustic imaging of living subjects offers high spatial resolution at increased tissue depths
compared to purely optical imaging techniques. We have recently shown that intravenously injected
single walled carbon nanotubes (SWNTs) can be used as targeted photoacoustic imaging agents in
living mice using RGD peptides to target αvβ3 integrins. We have now developed a new targeted
photoacoustic imaging agent based on SWNTs and Indocyanine Green (SWNT-ICG) with
absorption peak at 780nm. The photoacoustic signal of the new imaging agent is enhanced by ~20
times as compared to plain SWNTs. The particles are synthesized from SWNT-RGD that noncovalently
attach to multiple ICG molecules through pi-pi stacking interactions. Negative control
particles had RAD peptide instead of RGD. We measured the serum stability of the particles and
verified that the RGD/RAD conjugation did not alter the particle's absorbance spectrum. Finally,
through cell uptake studies with U87MG cells we verified that the particles bind selectively to αvβ3
integrin. In conclusion, the extremely high absorption of the
SWNT-ICG particles shows great
promise for high sensitivity photoacoustic imaging of molecular targets in-vivo. This work lays the
foundations for future in-vivo studies that will use the SWNT-ICG particles as imaging agents
administered systemically.
Photoacoustic molecular imaging is an emerging technology offering non-invasive high resolution imaging of the molecular expressions of a disease using a photoacoustic imaging agent. Here we demonstrate for the first time the utility of single walled carbon nanotubes (SWNTs) as targeted imaging agents in living mice bearing tumor xenografts. SWNTs were conjugated with polyethylene-glycol-5000 connected to Arg-Gly-Asp (RGD) peptide to target the αvβ3 integrin that is associated with tumor angiogenesis. In-vitro, we characterized the photoacoustic spectra of the particles, their signal linearity and tested their uptake by αvβ3-expressing cells (U87MG). The photoacoustic signal of SWNTs was found not to be affected by the RGD conjugation to the SWNTs and was also found to be highly linear with concentration (R2 = 0.9997 for 25-400nM). The cell uptake studies showed that RGD-targeted SWNTs gave 75% higher photoacoustic signal than non-targeted SWNTs when incubated with U87MG cells. In-vivo, we measured the minimal detectable concentration of SWNTs in living mice by subcutaneously injecting SWNTs at increasing concentrations. The lowest detectable concentration of SWNTs in living mice was found to be 50nM. Finally, we administered RGDtargeted and non-targeted SWNTs via the tail-vein to U87MG tumor-bearing mice (n=4 for each group) and measured the signal from the tumor before and up to 4 hours post-injection. At 4 hours post-injection, tumors of mice injected with RGD-targeted SWNTs showed 8 times higher photoacoustic signal compared with mice injected with non-targeted SWNTs. These results were verified ex-vivo using a Raman microscope that is sensitive to the SWNTs Raman signal.
In photoacoustic (optoacoustic) medical imaging, short laser pulses irradiate absorbing structures found in tissue, such as blood vessels, causing brief thermal expansions that in turn generate ultrasound waves. These ultrasound waves which correspond to the optical absorption distribution were imaged using a two dimensional array of capacitive micromachined ultrasonic transducers (CMUTs). Advantages of CMUT technology for photoacoustic imaging include the ease of integration with electronics, ability to fabricate large two dimensional arrays, arrays with arbitrary geometries, wide-bandwidth arrays and high-frequency arrays. In this study, a phantom consisting of three 0.86-mm inner diameter polyethylene tubes inside a tissue mimicking material was imaged using a 16 x 16 element CMUT array. The center tube was filled with India-ink to provide optical contrast. Traditional pulse-echo data as well as photoacoustic image data were taken. 2D cross-sectional slices and 3D volume rendered images are shown. Simple array tiling was attempted, whereby a 48 x 48 element array was simulated, to illustrate the advantages of larger arrays. Finally, the sensitivity of the photoacoustics setup to the concentration of ink in the tube was also explored. For the sensitivity experiment a different phantom consisting of only one 1.14-mm inner diameter polyethylene tube inside a tissue mimicking material was used. The concentration of the ink inside the tube was varied and images were taken.
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