In this thesis we utilize multiphoton microscope as a tool to investigate an enhancement method of transdermal delivery system, that is, iontophoresis. We choose the charge carried by fluorescent model drugs as a varient factor. We study the dependence of molecular charge in transdermal iontophoretic molecular delivery. We chose 6,8-dihydroxy-1,3-pyrenedisulfonic acid disodium salt, 8- hydroxypyrene-1,3,6-trisulfonic acid, trisodium salt, and calcein with the respective charges of -2, -3, and -4. These molecules are used as model drugs to probe the difference in iontophoretic delivery pathways when using molecules with different charges. Because of the visualization ablilty of fluorescent microscopy, it becomes able for us to observe the distribution of model drug inside the skin tissue, which is different from the traditional ways used for this experiment. The result shows that the charge of the molecules can affect the delivery efficiency and the delivery path, and is of physical meaning.