Imaging 3D cell culture systems using an optical resolution photoacoustic microscope

Optical-resolution photoacoustic microscopy (OR-PAM) provides high sensitivity and resolution to optical absorption and opens a new window to study biological systems at the cellular level. Cancer cells propagated in vitro partially retain the characteristics of cancer, such as immortality and resistance to apoptosis. For 3D cell imaging, the most widely used imaging modalities, the laser scanning optical microscopies (e.g., confocal and multi-photon microscopy) have limited tissue penetration, and they usually require the use of a fluorescent chromophore as the probe as well. PAM system has deeper penetration, and it does not need depth scanning. In other words, 2D scanning can generate 3D photoacoustic (PA) images. In this paper, a combined laser-scanning OR-PAM with optical microscope (OM) system has been setup in the transmission mode. The proposed OR-PAM system has achieved a lateral resolution of 0.59 μm at cellular level with a penetration depth of 0.60 mm, so it has the ability of do cell imaging and other sub-micron cellular experiments. In our study, B16-F10 murine melanoma cells were measured at different layers in 3D cell culture system mode. Two-dimensional optical or mechanical scanning of the object platform, in combination with optical/ultrasonic detection, can provide volumetric optical and 2D/3D PA images. From proof-of-principle experiments, 2D scanning generated 3D PA images and the distribution of cells at different layers agreed closely to the optical image of their localizations. Thus, the OR-PAM system demonstrated great potential in cell imaging.