Patent ID: 8704196

Claim:
A method for generating an image of a sample by microscopy methods which provide different spatial resolutions, wherein at least two of the following microscopy methods are combined: a first microscopy method including exciting the sample to luminescence by structured line-type or wide-field illumination; if structured line-type illumination is used, rotating a structuring and shifting the structuring several times for each rotation position, wherein at least three rotation positions and at least three shift positions per rotation position are realized; imaging the luminescent sample onto a 2D detector with a predetermined optical resolution for each combination of rotation position and shift position; and generating a first microscopy image with spatial resolution increased beyond the predetermined optical resolution from thus-obtained images by a computational processing comprising Fourier analysis, a second microscopy method including: labelling the sample with label molecules which can be excited to emit particular luminescence radiation only after being activated by a switching signal; applying the switching signal to the sample such that only a sub-set of the label molecules present in the sample are activated, wherein there are partial areas in the sample in which activated label molecules have a distance to their closest neighbouring activated label molecules which is greater than or equal to the predetermined optical resolution; exciting the activated molecules to emit luminescence radiation; imaging the sample emitting luminescence radiation onto the 2D detector with the predetermined optical resolution; analyzing the image; and generating image data from the image which give geometric locations of the label molecules emitting luminescence radiation with a local resolution increased above the predetermined optical resolution; and generating a second microscopy image from the image data, a third microscopy method including generating a third microscopy image by application of laser scanning microscopy, and combining at least two obtained microscopy images from the first, second and third microscopy images into a composite image.