International Business Machines Corporation

Cornell Research Foundation Inc.

Winfried Denk

Watt W Webb

Johannes Georg Bednorz

Wolfgang D Pohl

Martin A Lanz

James P Strickler

A laser scanning microscope produces molecular excitation in a target material by simultaneous absorption of two photons to thereby provide intrinsic three-dimensional resolution. Fluorophores having single photon absorption in the short (ultraviolet or visible) wavelength range are excited by a stream of strongly focused subpicosecond pulses of laser light of relatively long (red or infrared) wavelength range. The fluorophores absorb at about one half the laser wavelength to produce fluorescent images of living cells and other microscopic objects. The fluorescent emission from the fluorophores increases quadratically with the excitation intensity so that by strongly focusing the laser light, fluorescence as well as photobleaching are confined to the vicinity of the focal plane. This feature provides depth of field resolution comparable to that produced by confocal laser scanning microscopes, and in addition reduces photobleaching. Scanning of the laser beam, by a laser scanning microscope, allows construction of images by collecting two- photon excited fluorescence from each point in the scanned object while still satisfying the requirement for very high excitation intensity obtained by focusing the laser beam and by pulse time compressing the beam. The focused pulses also provide three-dimensional spatially resolved photochemistry which is particularly useful in photolytic release of caged effector molecules.

Two-photon laser microscopy

This light waveguide consists of an optically transparent body cut at one end to a sharp tip and polished optically flat at the other end. A metallization layer on its surface is thick enough to be opaque. By pressing the waveguide against a rigid plate the metallization is plastically deformed so as to expose a tiny aperture at the tip of the body through which light can pass. By carefully controlling the deformation of the metallization the diameter of the aperture can be kept between 10 and 500 nm. The waveguide can be incorporated in a semiconductor laser of a read/write head used in an optical storage device.

Light waveguide with a submicron aperture, method for manufacturing the

Growing community of inventors

Zurich, Switzerland

Winfried Denk