Method for three-dimensional high resolution localization microscopy

US10024793B2 · kind B2 · utility

Assignee

• Carl Zeiss Microscopy GmbH · DE

Inventors

• Jörg Ritter · Baden-Baden, DE
• Jörg Siebenmorgen · Jena, DE
• Thomas Kalkbrenner · Jena, DE

Key dates

• Filing date: May 13, 2014
• Grant date: Jul 17, 2018
• Expiry date: Apr 7, 2036

Classification

• Technology area (CPC H): Electricity
• CPC primary: H04N13/20
• WIPO field: Measurement
• WIPO sector: Instruments

Abstract

A three-dimensional high-resolution localization microscopy method including illuminating a sample by excitation radiation to excite fluorescence markers in the sample to luminesce, and imaging the sample in an image frame via imaging optics along an imaging direction, wherein the image frame contains images of the luminescing fluorescence markers, and the imaging optics have a plane of focus and an optical resolution. The excitation step and imaging steps are repeated multiple times to generate a plurality of image frames, wherein the excitation steps are performed to isolate the images of the luminescing fluorescence markers in each image frame for at least some of the luminescing fluorescence markers. The location of the corresponding fluorescence marker is determined in each instance in the generated plurality of image frames from the isolated images of the luminescing fluorescence markers, and a highly resolved total image is generated from the locations determined in this way.