Dpo7254

The photoacoustic waves were generated with a pulsed 6 ns FWHM Nd:YAG laser (EKSPLA NL301G NdYAG) that when necessary was coupled with an OPO (EKSPLA OPO PG-122) or with a 30 ps FWHM Nd:YAG laser (EKSPLA 2143A), with a diameter of 2 mm. With the ns pulse laser, a fluence of 10 mJ cm⁻² was used, while with the ps laser, a fluence of 2 mJ cm⁻² was used. The presence of high frequencies was analyzed using a front-face irradiation setup (Fig. 1) developed by our group for photoacoustic calorimetry experiments,^{29 (link)} with a 225 MHz contact transducer (Panametrics/Olympus, model V2113). We used a quartz mirror capable of reflecting >99% of the incident light to ensure the safety of the detector. It must be noted that the results were obtained using this quartz mirror of 1 cm between the material to be analyzed and the transducer, so high frequencies may be significantly attenuated. The piezophotonic material was placed on top of the mirror, which is inside of a support, and we applied silicon or water gel to improve the acoustic coupling. Finally, a heavy material (1.5 kg) with an optical window was placed over the film to ensure confinement. The signal was recorded using an oscilloscope (DPO7254 Tektronix, 2.5 GHz bandwidth) with an average of at least 200 waveforms. Films were replaced whenever they appeared to be damaged.

Tissue autofluorescence was excited with 355 nm pulsed laser (<0.6 ns per pulse, >2 $\mathsf{\mu }$ J per pulse, repetition rate 4 kHz; TEEM photonics STV-02E, Meylan, France). A 400 $\mathsf{\mu }$ m core diameter multimode fiber placed perpendicular to the sample was used to deliver and collect the light. A second multimode fiber (600 $\mathsf{\mu }$ m in core diameter) was used to guide the tissue autofluorescence to a monochomator (MicroHR, Horiba, Kyoto, Japan). Light was detected with a gated microchannel plate photomultiplier tube (R5916U-50, Hamamatsu, Hamamatsu, Japan) connected to a high-speed amplifier (C5594, Hamamatsu), and digitized by an oscilloscope (DPO7254, Tektronix, Beaverton, OR, USA). The laser power at the sample plane was kept under 5 mW. Fluorescence decays were acquired from 370 to 600 nm with 2 nm increments and a 0.08 ns resolution. Each data point was an average of 64 decays.
Data were acquired from three points along the length of the colon corresponding to the proximal, the middle, and the distal ends. Three measurements were taken on each position. The results presented in Figure 2 represent the mean (solid thick line) over 4 independent samples (2 female and 2 male) and their corresponding standard deviation (shaded area). Samples were kept in phosphate buffer saline (PBS) for the duration of imaging.