Molecularly targeted fluorescent probes are currently being developed to improve the endoscopic detection of intestinal pathologic conditions.

We report on the development and testing of a novel multichannel microendoscope capable of quantitatively reporting such probes simultaneously at different wavelengths in real time. We assessed the feasibility of detecting and quantifying beacons that can be activated by protease and correlating imaging with disease state.

The microendoscope consisted of a 20-gauge fiberoptic catheter and dichroic beam splitters that simultaneously display visible light, 700 nm and 800 nm near infrared (NIR) fluorescent light. NIR interchannel separation was tested on in vitro phantoms. Two mouse models were used (Apcmin^+/− mice for colonic adenomas and CT26 murine colon cancer). A perfusion probe and one activated by protease at a separate wavelength were injected before endoscopic evaluation.

The microendoscope fluorochrome detection limit was approximately 10 fmol; ratio imaging in the NIR was accurate (±8% of true probe concentration between 0.3 to 100 μg/ml of a protease sensor). Both colonic adenomas and adenocarcinomas were clearly visible in the NIR channel on protease probe administration in live mice. Ratio imaging of protease activity/perfusion increased from healthy colon to adenomas to adenocarcinomas.

Evaluation across additional spontaneous tumor models may provide more data on the translation of these findings.

Our data show the feasibility of multichannel microendoscopic imaging of molecular targets in vivo and that ratio imaging may provide a novel means for characterizing colonic lesions. When scaled up clinically, this could aid in increasing lesion detection and quantitative assessment of distinct molecular markers.