Three photon excitation

 

A recent emphasis of the lab has been the use of three photon excitation (3PE) microscopy to access normally <300nm excited native indoleamine fluorescence, for example serotonin to the right. Red light 3PE alleviates the need for deep UV optics and their associated aberrations and low transmissivities. Nonlinear excitation is also intrinsically localized to the focal plane and thus is potentially less invasive than single photon techniques. Using this method we have imaged serotonin-containing secretory granules in RBL-2H3 cells (Maiti, 1997), which are viable and release upon stimulation. The feasibility of these experiments was first suggested by our measured 3PE monoamine cross sections shown below:
For more MPE cross section measurements and measurement techniques see:

C. Xu, J. Guild, W.W. Webb and W. Denk. Opt. Lett., 20: 2372, 1995.

C. Xu and W.W. Webb. J. Opt. Soc. Am. B, 13:481, 1996.

C. Xu, W.R. Zipfel, J.B. Shear, R.M. Williams and W.W. Webb. Proc. Nat. Acad. Sci, 93:10763, 1996.

C. Xu, R.M. Williams, W.R. Zipfel and W.W. Webb. Bioimaging, 4:198, 1996.

S. Maiti, J.B. Shear, R.M. Williams, W. R. Zipfel and W.W. Webb. Science, 275:530, 1997.