The Applications of Two-photon Microscopy in Biomedicine and Its Development

The history of the development of the light microscopy was a history of constant improvement of the resolution and the contrast of the microscopy.Two photon microscopy represented the development of nonlinear microscopy in the recent 30 years.The resolution limit of two photon microscopy was close to that of confocal microscopy,whereas with using two photon microscopy the imaging contrast and depth of intact sectioning had been improved obviously.Furthermore phototoxicity and photobleaching had been reduced in large extent.Because there were many kinds of autofluorescent fluorophores in tissue and cell,two photon microscopy had been widely applied in imaging of skin tissue,even cancerous tissue and cell.Based on laser scanning confocal microscopy,two photon microscopy can be combined easily with second harmonic generation microscopy,so the important component of skin tissue including collagen fiber can be imaged simultaneously.Besides,two photon microscopy can also couple with other nonlinear optical phenomenon for imaging tissues and living cells,which implies its prospective development in future.Following by the appearance of portable and low-cost two photon devices,we expect the effective use of the two photon microscopy in the clinical medicine.