Measurement of the 4Pi-confocal point spread function proves 75 nm axial resolution

TLDR

A 4Pi‑confocal microscope illuminates and observes specimens coherently from above and below, increasing the effective numerical aperture. The authors measured the point spread functions of both 4Pi‑confocal and conventional confocal microscopes. Measurements show a three‑ to seven‑fold improvement in axial resolution, achieving as fine as 75 nm, enabling powerful 3‑D imaging.

Abstract

In a 4Pi-confocal microscope the specimen is illuminated and observed coherently from above and below such that the numerical aperture is increased [S. W. Hell, European Patent Application 91121368.4 (filed 1990, published 1992), S. W. Hell and E. H. K. Stelzer, J. Opt. Soc. Am. A 9, 2159 (1992)]. The point spread functions of 4Pi-confocal and confocal microscopes were measured. Our measurements prove a three- to seven-fold increase of axial resolution, thus opening the prospect for a powerful three-dimensional imaging technique with an axial resolution down to 75 nm.