LIGHT-PAD MICROSCOPE FOR HIGH-RESOLUTION 3D FLUORESCENCE IMAGING AND 2D FLUCTUATION SPECTROSCOPY

• US 20120206798A1
• Filed: 02/13/2012
• Published: 08/16/2012
• Est. Priority Date: 02/14/2011
• Status: Active Grant

• Current Assignee: European Molecular Biology Laboratory
• Original Assignee: European Molecular Biology Laboratory
• Inventors: Wachsmuth, Malte, Knop, Michael, Capoulade, Jérémie
• Granted Patent: US 10,908,403 B2
• Time in Patent Office: Days
• US Class Current: 359/385
• CPC Class Codes: G01N 21/6408 with measurement of decay time, time resolved fluorescence
• CPC Class Codes: G02B 21/002 Scanning microscopes scanning near field optical microscopes G01Q60/18
• CPC Class Codes: G02B 21/0032 Optical details of illumination, e.g. light-sources, pinholes, beam splitters, slits, fibers G02B21/0036 - G02B21/008; means for illumination of specimens in general G02B21/06
• CPC Class Codes: G02B 21/06 Means for illuminating specimens
• CPC Class Codes: G02B 21/16 adapted for ultraviolet illumination ; Fluorescence microscopes G02B21/0076 takes precedence
• CPC Class Codes: G02B 21/367 providing an output produced by processing a plurality of individual source images, e.g. image tiling, montage, composite images, depth sectioning, image comparison

Abstract

A microscope is described, having an illumination light path for illuminating a sample or object and a viewing light path for viewing the sample. The microscope includes an illumination light path focussing arrangement in the illumination light path, defining a substantially two-dimensional sample or object illumination region extending along an illumination direction of the illumination light path and transversely thereto. The microscope further includes an illumination region-confining device in the illumination light path for selectively illuminating a portion of the substantially two-dimensional object illumination region, wherein the portion of the substantially two-dimensional object illumination region is confined at least in the illumination direction and/or in the direction transversely thereto.

Claims

1. A microscope having an illumination light path for illuminating a sample and at least one viewing light path for viewing the sample, the microscope comprising

an illumination light path focussing arrangement in the illumination light path, the illumination light path focussing arrangement defining a substantially two-dimensional object illumination region extending along an illumination direction of the illumination light path and transversely thereto, and

an illumination region-confining device in the illumination light path for selectively illuminating a portion of the substantially two-dimensional object illumination region, wherein the portion of the substantially two-dimensional object illumination region is confined at least in one of the illumination direction and the direction transverse thereto.

2. The microscope of claim 1, wherein the illumination light path focussing arrangement comprises at least one of a cylindrical lens, an anamorphically shaped lens, a one-dimensional array of spherical or of aspherical lenses.

3. The microscope of claim 1, wherein the illumination light path focussing arrangement comprises at least one anamorphically shaped mirror.

4. The microscope of claim 1, wherein the illumination region-confining device comprises at least a first aperture for confining the portion of the substantially two-dimensional object illumination region in the in the illumination direction.

5. The microscope of claim 1, wherein the illumination region confining device comprises at least a second aperture for confining the portion of the substantially two-dimensional object illumination region in a direction transversely to the illumination direction.

6. The microscope of claim 1, wherein the illumination region-confining devices comprises a light beam shaper.

7. The microscope of claim 1, wherein a viewing direction of the at least one viewing light path is substantially perpendicular to the illumination direction and to the substantially two-dimensional object illumination region.

8. The microscope of claim 1, further comprising an adjustable detection aperture in the detection path that allows to reduce an effectively viewed region in one or two dimensions.

9. The microscope of claim 8, wherein the portion of substantially two-dimensional object illumination region and the effectively observed/viewed region are congruent and/or coincident.

10. The microscope of claim 9, wherein the portion of substantially two-dimensional object illumination region and the effectively observed/viewed region are congruently and/or coincidentally moveable through the sample (8).

11. The microscope of claim 1, further comprising an additional objective lens for illumination and viewing of a sample arranged in the sample illumination region.

12. The microscope of claim 11, wherein the additional objective lens is part of an epifluorescence microscope.

13. The microscope of claim 11, wherein the additional objective lens is part of an inverted microscope.

14. A method for detecting a sample, the method comprising:

illuminating a two-dimensional portion of a sample by focussing an illumination light beam into a substantially two-dimensional object illumination region extending in an illumination direction of the illumination light beam and transversely thereto;

wherein illuminating the two-dimensional portion further comprises confining the substantially two-dimensional object illumination region for selectively illuminating a portion of the substantially two-dimensional object illumination region, wherein the portion of the substantially two-dimensional object illumination region is confined at least in the illumination direction.

15. The method of claim 14, further comprising viewing the substantially two-dimensional object illumination region in a viewing direction, the viewing direction being substantially perpendicular to the illumination direction.

16. The method of claim 14, further comprising moving at least one of the substantially two-dimensional object illumination region or the portion of the substantially two-dimensional object illumination region through the sample.

17. The method of claim 14, further comprising measuring a signal fluctuation in the portion of the substantially two-dimensional object illumination region.

18. The method of claim 14, further comprising measuring a fluorescence intensity contrast for alignment.