Tissue penetration of laser light and measuring depth (depth sensitivity) for laser Doppler flowmetry in the gastrointestinal application were studied in the feline and human bowel and stomach with a wall thickness of from 2.1 to 7.0 mm. Mucosal and serosal recordings from corresponding sites in the gastrointestinal tract were comparable and in the intestine correlated to total blood flow. Laser light was reflected by a mirror placed on the opposite side of the bowel wall, and hyperemic reactions were similarly recorded from both sides of the bowel with wall thickness up to 6 mm. Placing layers of unperfused feline small intestine (average thickness, 2.1 mm) between the probe and the perfused feline gastric wall attenuated the laser Doppler signal in an exponential manner. We conclude that with this instrument, the tissue penetration and measuring depth of laser Doppler flowmetry are at least 6 mm in the gastrointestinal application.