© 2002 J.Paul Robinson, Purdue University
Excitation Saturation
•The rate of
emission is dependent upon the time the
molecule remains within the excitation state (the excited state lifetime f)
•Optical saturation occurs when the
rate of excitation exceeds the reciprocal of f
•In a scanned image of 512 x 768 pixels (400,000 pixels)
if scanned in 1 second
requires a dwell time per pixel of 2 x 10-6 sec.
•Molecules that remain in the excitation beam for
extended periods have higher
probability of interstate
crossings and thus phosphorescence
•Usually, increasing
dye concentration can be the most effective
means of increasing
signal when energy is not the limiting
factor (i.e. laser based confocal
systems)