Practice question

Fluorescence involves absorption of high-energy photons exciting electrons from ground state to excited singlet, followed by rapid non-radiative vibrational relaxation losing some energy as heat. When electrons return to ground state they emit photons with less energy than absorbed, as described by Stokes. Less energy corresponds to longer wavelength, so emission spectrum is red-shifted relative to excitation. This wavelength difference allows dichroic mirrors and emission filters to separate weak fluorescence from intense excitation light, essential for visualizing labeled proteins in cells without interference from illumination source itself.