Photoacoustic Microscopy (PAM)

Photoacoustic microscopy (PAM) is a hybrid in vivo imaging technique that detects optical contrast via the photoacoustic effect. Unlike the pure optical microscopy techniques discussed later, PAM takes advantage of the weak acoustic scattering in tissue and thus breaks through the optical depth penetration limit (~ 1 mm) in soft tissues. Since ultrasonic scattering in tissue is three orders of magnitude weaker than the optical scattering, PAM can effectively image at depths up to a few millimeters.

In PAM (see Figure 1), a laser beam is focused using an objective lens and the light passes through a water tank that is in contact with the sample to be imaged. This light is absorbed at the focus, heats the sample, and induces an initial pressure rise which propagates through the tissues as a wide-band acoustic wave. An ultrasonic transducer then detects the acoustic wave. The lateral resolution is determined by the focus of the laser beam, and is typically a few microns, while the axial resolution is about 15 µm. Both endogenous and exogenous contrast agents have been used successfully in PAM imaging. The laser sources typically have ns pulse durations with wavelengths dictated by the absorption of the contrast agents. A representative laser source for PAM is a low repetition rate (10-30 Hz) ns-pulsed Nd:YAG laser. Such a system typically operates at 1064 nm, but this wavelength can be converted to 355 nm via THG and produce pulse energies of several hundreds of millijoules. This source is then used to pump an OPO, which generates ns pulses throughout the VIS portion of the spectrum (see Laser Spectral Tunability for details regarding wavelength conversion and generation).