Standing-wave excitation of fluorescence in a laser-scanning microscope allows precise contour mapping of the red blood cell membrane

We demonstrate fluorescence excitation at multiple planes in a laser-scanning microscope by using the standing wave from a mirror placed close to the specimen. We have observed precise modulation of the standing waves close to a mirror, with a frequency proportional to the Stokes shift, corresponding to a moire pattern between the excitation and emission standing-wave fields. We use standing-wave excitation to plot the exact contour maps of the red blood cell membrane, with an axial resolution of ≈90 nm. The method may prove useful in the study of diseases which involve the surface membrane of red blood cells.