Radiant energy – Luminophor irradiation
Patent
1996-11-21
1998-03-24
Porta, David P.
Radiant energy
Luminophor irradiation
2504591, G02B 2100, G01N 2164
Patent
active
057315884
DESCRIPTION:
BRIEF SUMMARY
The invention concerns a device for optically measuring a point on a sample with high local resolution, with a light source to send out a exciting light beam suitable for exciting an energy state on the sample, a lens for focusing the exciting light beam on the points, the sample which can be arranged in the focal area of the lens, a separating device for separating out the emission light spontaneously emitted from the sample based on the excitation of the energy state and a detector for detecting the emission light.
The invention also concerns a process for optically measuring a point on a sample with high local resolution, in which an exciting light beam is focused on the point to be measured by means of a lens and excites the energy state there, and in which the emission light spontaneously emitted by the point, based on the excitation of the energy state, is separated out and detected.
FIELD OF THE INVENTION
The invention relates generally to an optical measurement device and more specifically to a device which performs optical measurements of a sample with high local resolution.
BACKGROUND
Such a device and such a process are known from practice. They have applications, for example, in microscopes, particularly raster microscopes. With a raster microscope, individual points on a sample are scanned and measured. In this way, the sample can be measured three-dimensionally. Luminescent, particularly fluorescent or phosphorescent samples, or samples with corresponding dyes, are used.
In such a device and such a process, it is desirable to achieve good local resolution. The local resolution is given by the spatial expansion of the so-called effective point-imaging function. This is a place-dependent function, which quantifies the probability with which a photon will be spontaneously emitted from a certain point in the focal range. It is identical to the spatial distribution of the probability that the energy state is excited at a certain point in the focal range. In conventional fluorescent microscopes,the effective point-imaging function of the lens at the wavelength of the excitation light, which gives the distribution of the intensity of the excitation light in the focal range of the lens and quantifies the probability with which an exposure photon will be met at a certain point in the focal range from a quantum-mechanics standpoint. In a raster microscope, the raster division is limited by the local resolution. With better local resolution, a finer division can therefore be selected, with which a better resolution of the reconstructed image can be achieved.
It is known that in a raster microscope, the resolution can be improved by having the light detected by the sample imaged on the point detector, which is arranged in one of the planes conjugated to the focal plane of the lens. Such an arrangement is called confocal. The better resolution is caused by the fact that two point-imaging functions determine the image in the confocal raster microscope: The effective point-imaging function and the detection-imaging function, which describes the image of the light to be detected that is emitted by the sample in the point detector and quantifies the probability with which a photo emitted from the focal range goes into the point detector from a quantum-mechanics standpoint. Since both exposure and detection must take place, the point-imaging function of a confocal raster microscope is the product of both probability distributions, i.e., from the effective point-imaging function and the detection-point-imaging function. This leads to a clearly narrower main maximum of the confocal point-imaging function compared to a microscope not arranged confocally. This corresponds to a higher resolution of the confocal microscope and brings about discrimination of all points that are not in the direct vicinity of the focus. The latter is the prerequisite for making three-dimensional images in the raster process.
SUMMARY
The task of the invention is to improve the generic device and the generic process in such a wa
REFERENCES:
"Breaking the Diffraction Resolution Limit by Stimulated Emission: Stimulated-Emission-Depletion Fluorescence Microscopy", Stefan W. Hell, et al, Jun. 1, 1994, Optical Society of America, vol. 19, No. 11, pp. 780-782.
"Ground-State-Depletion Fluorescence Microscopy: A Concept for Breaking the Diffraction Resolution Limit", S.W. Hell, et al, 1995, Applied Physics B, pp. 495-497.
Hell Stefan
Wichmann Jan
Hanig Richard
Porta David P.
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