Apparatus and methods relating to spatially light modulated microscopy

Abstract

Apparatus and methods relating to microscopes having specific control of the light that contacts a sample and / or a light detector, such as the eye of the user, a charge couple device or a video camera. The improved control includes enhanced, selective control of the angle of illumination, quantity of light and location of light reaching the sample and / or detector. The microscopes comprise one or more spatial light modulators in the illumination and / or detection light path of the microscope at one or both of the conjugate image plane of the aperture diaphragm of the objective lens and the conjugate image plane of the sample.

Description

RELATED APPLICATIONS [0001] This application claims priority to U.S. Provisional Application No. 60 / 063,893 filed on Oct. 29, 1997, the contents of which are incorporated herein by reference.FIELD OF THE INVENTION [0002] The field of the present invention is the magnification of images of objects, and more particularly microscopes and the field of microscopy. BACKGROUND OF THE INVENTION [0003] Microscopy is used to produce magnified representations of both dynamic and stationary objects or samples. There are many different modes of microscopy such as brightfield microscopy, darkfield microscopy, phase contrast microscopy, fluorescence microscopy, reflectance or reflected light microscopy and confocal microscopy. All of these forms of microscopy deliver illumination light in a controlled, fashion to the sample and collect as much of the light containing the desired information about the sample as possible. Typically, this is accomplished using Kohler illumination in any of reflectanc...

AI Technical Summary

Benefits of technology

[0008] The present invention provides microscopes that have significant advantages in controlling the light that contacts a sample and / or that is detected emanating from a sample. The improved control includes enhanced, selective control of the angle of illumination, the quantity of light and the location of light reaching the sample and / or detector. The present invention provides these advantages by placing one or more spatial light modulators in the illumination and / or detection light path of the microscope at one or both of the conjugate image plane of the aperture diaphragm of the objective lens and the conjugate image plane of the sample.

[0029] In still further embodiments, the computer-implemented programming causes the light detector to detect only light impinging on a central detection pixel aligned with and corresponding to an individual light transmission pixel of the second spatial light modulator that is on, and can compile the data provided by the at least one adjacent detection pixel and combine it with the data provided by the central detection pixel. Thus the computer-implemented programming can compile the data provided by the adjacent detection pixels and combine it with the data provided by the central detection pixel such that the focus of the microscope is enhanced compared to the focus without the data from the adjacent detection pixels.

[0030] In yet further embodiments, the microscopes provide time-delayed fluorescence detection, wherein the computer-implemented programming causes at least one of the spatial light modulators to illuminate the sample for an amount of time suitable to induce fluorescence in the sample and then ceasing illuminating the sample, and then causing the detector to begin detecting fluorescence emanating from the sample.

[0043] In still other aspects, the present invention provides methods for providing uniform intensity of illumination light to a sample in a microscope, wherein the microscope comprises a spatial light modulator comprising an illumination array of individual light transmission pixels and disposed in an illumination light path at a conjugate image plane of a sample and a light detector comprising a detection array of individual detection pixels and disposed downstream from the upstream spatial light modulator at a conjugate image plane of the sample and that receives light directly from the upstream spatial light modulator, wherein the spatial light modulator and the light detector are operably connected to at least one controller containing computer-implemented programming that controls transmissive characteristics of the upstream spatial light modulator and that compiles the light intensity detection data provided by the light detector, the methods comprising: varying the on / off status of individual light transmission pixels of the spatial light modulator to compensate for non-uniform light intensities detected by the light detector, thereby providing a uniform intensity of illumination light that is transmitted to the sample.

Problems solved by technology

Still other known confocal scanning systems have used a laser beam rastered with rotating mirrors to scan a specimen or a laser beam that scans a slit rather than a spot; such slit scanning increases imaging speed but slightly degrades resolution.

Conventional confocal microscopes can be slow to acquire images for certain applications and become even slower as the scan line density increases and the aperture separation decreases.

In addition, it is difficult to practically adjust the perimeters of the confocal microscope in commercial systems, and the signal to noise ratio (SNR) is sacrificed to increase the imaging rate.

In addition, proper alignment of conventional confocal systems can be critical and difficult to maintain.

In addition, laser-based systems are more expensive than white light systems, but such laser systems do not offer a selection of illumination wavelengths and can also lead to photo toxicity and rapid photo bleaching.

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