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. Author manuscript; available in PMC: 2012 Jan 17.
Published in final edited form as: Chem Sci. 2011;2(6):1080–1085. doi: 10.1039/C0SC00637H

Figure 5.

Figure 5

Modulated Alexa 594 fluorescence in solution as a function of primary laser intensity. As the primary excitation increases, the STED fluorescence overcomes the constant depletion-laser-induced two photon excited fluorescence. A. Amplitude of the 10-Hz frequency component initially decreases, then increases as the signal-degrading two-photon excited fluorescence from the depletion laser is overcome by the STED signal. B. The DC (non-modulated) total fluorescence signal increases linearly with increasing primary laser intensity. Fluorescence from both primary and two-photon excited secondary laser excitation give rise to the signal in B. The STED component in A is seen to be out of phase, and offset by the degrading two-photon signal, when demodulated.