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Molecular imaging is a powerful tool for investigating disease processes and potential therapies in both in vivo and in vitro systems. However, high resolution molecular imaging has been limited to relatively shallow penetration depths that can be accessed with microscopy. Optical coherence tomography (OCT) is an optical analogue to ultrasound with relatively good penetration depth (1-2 mm) and resolution (approximately 1-10 microm). We have developed and characterized photothermal OCT as a molecular contrast mechanism that allows for high resolution molecular imaging at deeper penetration depths than microscopy. Our photothermal system consists of an amplitude-modulated heating beam that spatially overlaps with the focused spot of the sample arm of a spectral-domain OCT microscope. Validation experiments in tissuelike phantoms containing gold nanospheres that absorb at 532 nm revealed a sensitivity of 14 ppm nanospheres (weight/weight) in a tissuelike environment. The nanospheres were then conjugated to anti-EGFR, and molecular targeting was confirmed in cells that overexpress EGFR (MDA-MB-468) and cells that express low levels of EGFR (MDA-MB-435). Molecular imaging in three-dimensional tissue constructs was confirmed with a significantly lower photothermal signal (p&lt;0.0001) from the constructs composed of cells that express low levels of EGFR compared to the overexpressing cell constructs (300% signal increase). This technique could potentially augment confocal and multiphoton microscopy as a method for deep-tissue, depth-resolved molecular imaging with relatively high resolution and target sensitivity, without photobleaching or cytotoxicity.

Photothermal Optical Coherence Tomography of Epidermal Growth Factor Receptor in Live Cells Using Immunotargeted Gold Nanospheres