Past exam of the short options paper shared by Prelims and Parts A and B of the physics course of the University of Oxford / 2015 / S12 / 2 / 2

The excitation rate in the weak, unsaturated fluorescence regime is proportional to the time-averaged squared electric field amplitude. Thus the illumination profile relevant to excitation isThe amplitude decay length $1/\kappa$ is twice the intensity decay length. This profile is often called the evanescent excitation intensity; it does not imply a propagating normal energy flux through an ideal lossless totally reflecting interface.

Total internal reflection fluorescence microscopy restricts excitation to a shallow layer adjacent to the glass, typically on the scale of hundreds of nanometres for visible wavelengths and suitable angles. Fluorophores near the basal cell membrane are excited while much of the cytoplasm and bulk solution remain dark. The resulting reduction in out-of-focus fluorescence and autofluorescence improves the signal-to-background ratio for individual molecules, allows camera-based parallel observation, and reduces unnecessary excitation of deeper regions. Its limitation is the same selectivity: it does not image arbitrary molecules deep inside a cell. This near-interface selectivity was directly demonstrated in cell–substrate contact imaging.^[ref]