Scanning microscope and method for determining the point spread function (PSF) of a scanning microscope
First Claim
1. A method for operating a scanning microscope and for determining point spread functions, with which sample images are recorded with the scanning microscope, comprisingscanning a sample with at least one illuminating light beam;
- recording at least one sample image with a detector device of the scanning microscope during a scan by said at least one illuminating light beam;
computing the point spread function, with which a sample image is recorded by the scanning microscope, from the at least one sample image,said detector device having receiving elements, where the distance between said receiving elements is smaller than a diffraction disk that generates a sample point on the detector device,said receiving elements generating detector signals which are read out for each of the different positions of the illuminating light beam on the sample, as a result of which the scanning of the sample allows the read out detector signals to generate a plurality of sample images, wherein the point spread functions with respect to the different detector signals are defined in each instance by means of an illumination point spread function and a detection point spread function,with respect to all of the detector signals, a matching illumination point spread function is shifted in accordance with the scanning motion for different detector signals, andwith respect to all of the detector signals, a matching detection point spread function takes account of a spatial offset between the detector elements, andusing the plurality of sample images to compute the illumination point spread function and the detection point spread function, said point spread functions being used to compute the point spread functions with respect to the different detector signals, said sample images are used, together with the computation of the point spread functions, to compute a complete image.
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Abstract
A method for operating a scanning microscope and for determining point spread functions, with which sample images are recorded with the scanning microscope. The method includes scanning a sample with at least one illuminating light beam; recording at least one sample image with a detector device during a scan by the illuminating light beam; and comprising the point spread function, with which a sample image is recorded, from the at least one sample image. A detector device having receiving elements is used, where the distance between the receiving elements is smaller than a diffraction disk that generates a sample point on the detector device. Detector signals, generated by means of the receiving elements, are read out for each of the different positions of the illuminating light beam on the sample, as a result of which the scanning of the sample allows the detector signals, which are read out, to generate a plurality of sample images. In this case the point spread functions with respect to the different detector signals are defined in each instance by means of an illumination point spread function and a detection point spread function. With respect to all of the detector signals, a matching illumination point spread function is assumed that is shifted in accordance with the scanning motion for different detector signals. In addition, with respect to all of the detector signals, a matching detection point spread function is assumed that takes account of a spatial offset between the detector elements. The plurality of sample images are used to compute the illumination point spread function and the detection point spread function, and these functions are used to compute the point spread functions with respect to the different detector signals. In addition, the invention also relates to a corresponding scanning microscope.
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Citations
15 Claims
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1. A method for operating a scanning microscope and for determining point spread functions, with which sample images are recorded with the scanning microscope, comprising
scanning a sample with at least one illuminating light beam; -
recording at least one sample image with a detector device of the scanning microscope during a scan by said at least one illuminating light beam; computing the point spread function, with which a sample image is recorded by the scanning microscope, from the at least one sample image, said detector device having receiving elements, where the distance between said receiving elements is smaller than a diffraction disk that generates a sample point on the detector device, said receiving elements generating detector signals which are read out for each of the different positions of the illuminating light beam on the sample, as a result of which the scanning of the sample allows the read out detector signals to generate a plurality of sample images, wherein the point spread functions with respect to the different detector signals are defined in each instance by means of an illumination point spread function and a detection point spread function, with respect to all of the detector signals, a matching illumination point spread function is shifted in accordance with the scanning motion for different detector signals, and with respect to all of the detector signals, a matching detection point spread function takes account of a spatial offset between the detector elements, and using the plurality of sample images to compute the illumination point spread function and the detection point spread function, said point spread functions being used to compute the point spread functions with respect to the different detector signals, said sample images are used, together with the computation of the point spread functions, to compute a complete image. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14)
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15. A scanning microscope, comprising
a light source device for emitting at least one illuminating light beam, comprising a scanning device for generating a scanning motion of the at least one illuminating light beam across a sample, a detector device for recording at least one sample image during a scan by the illuminating light beam, electronic control and evaluation means, for computing a point spread function, with which a sample image is recorded, from the at least one sample image, said detector device having receiving elements, where the distance between said receiving elements is smaller than a diffraction disk that generates a sample point on the detector device, said electronic control and evaluation means further being designed to read out the detector signals, generated by means of the receiving elements, for each of the different positions of the illuminating light beam on the sample, as a result of which the scanning of the sample allows the detector signals, which are read out, to generate a plurality of sample images, wherein the point spread functions with respect to the different detector signals are defined in each instance by means of an illumination point spread function and a detection point spread function, and wherein said electronic control and evaluation means are designed to compute an illumination point spread function and a detection point spread function with the plurality of sample images; - and these two functions are used to compute the point spread functions with respect to the different detector signals,
and wherein with respect to all of the detector signals, there is a matching illumination point spread function that is shifted in accordance with the scanning motion for different detector signals; and
that with respect to all of the detector signals there is a matching detection point spread function that takes account of a spatial offset between the detector elements, said sample images are used, together with the computation of the point spread functions, to compute a complete image.
- and these two functions are used to compute the point spread functions with respect to the different detector signals,
Specification