The optical clearing technique is able improve light penetration depth by applying optical clearing agent (OCA) with high refractive indices and hyperosmolarity. However, the mechanism of tissue optical clearing is not much clear. In this work, both physical and physiological mechanisms of optical clearing are discussed. For physical mechanisms, a tissue phantom, Intralipid, was mixed with OCAs, and then the reduced scattering coefficient was theoretically predicted and experimentally measured. It was found that the physical mechanisms for optical clearing are due to the structure change of scatters or the enhancement of the background refractive index. For physiological mechanisms, the dehydration of porcine skin in vitro caused by topical application of OCA was investigated with NIR reflectance spectrum analysis. The results showed that dehydration is major factor during the initial few minutes, and some other changes of tissue structure maybe occurs with increase of time. And the microstructure and ultrastructure of rat skin was examined after in vivo application of glycerol, we found that both the thickness of skin and the size of fiber reduce. This work is helpful for clarifying the mechanisms of tissue optical clearing, which will contribute to the improvement and clinical application of optical clearing technique.
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