Systems and methods for three dimensional imaging
View Patent ↗Methods, devices and systems for up to three-dimensional scanning of target regions at high magnification are disclosed.
1. A microscope, comprising:
an objective;
a further lens assembly, wherein the objective and the further lens assembly have an identical configuration with respect to at least axial focusing aberrations, and wherein the objective and further lens assembly are arranged so that image light is received by the objective lens and subsequently passes through the further lens assembly;
a light source that generates illumination light;
a scanning/de-scanning optical assembly having a movable light-redirecting element positioned to route the illumination light into the objective such that an illumination sheet will emanate obliquely from the objective and sweep a volume region, the swept illumination sheet scanning across the volume region; and
a light sensor,
wherein the movable light-redirecting element is further positioned to redirect image light received from the volume region via the objective into the further lens assembly so as to hold a focus of light returning from a planar region illuminated by the illumination sheet onto the light sensor as said sheet of light is swept.
2. The microscope of claim 1 , further comprising a beam splitter arranged to form a junction positioned such that the image light and the light from the light source follow adjacent paths into and from the objective and follow divided paths remote from the objective.
3. The microscope of claim 1 , further comprising a stationary reflector positioned to reflect the image light back through the further lens assembly.
4. The microscope of claim 3 , wherein the stationary reflector is oriented at an angle with respect to the optical axis of the further lens assembly, the angle being selected responsively to an angle of an intermediate image that would be formed at the position of the mirror by the objective and further lens assembly.
5. The microscope of claim 1 , wherein the movable light-redirecting element is a motor-controlled reflecting element.
6. The microscope of claim 4 , wherein the reflecting element has a reflecting characteristic that makes identical angular changes in the direction of the light from the light source and the light received from the volume region as would a single planar reflecting surface.
7. The microscope of claim 4 , wherein the reflecting element has a single planar reflecting surface on which both the light from the light source and the light received from the volume region are incident.
8. The microscope of claim 1 , wherein the light source generates light that is selected to generate fluorescence in a sample containing fluorescent dye.
9. The microscope of claim 1 , wherein the light sensor includes a camera.
10. The microscope of claim 1 , wherein the light sensor includes a linear array of photomultiplier elements.
11. The microscope of claim 1 , wherein the light sensor includes a single photomultiplier element.
12. The microscope of claim 1 , wherein the light source includes a beam flattening optical element that flattens a round beam into said sheet of light.
13. The microscope of claim 1 , wherein scanning/de-scanning optical assembly includes a movable optical element that can be selectively positioned in the path of light from the light source to change the illumination beam between a sheet of light directed toward an edge of a back aperture of said objective (identifiable as a scan mode) and a broad illumination beam extending across said back aperture (identifiable as a confocal mode).
14. The microscope of claim 13 , wherein the movable optical element is refractive and expands a width of the light from the light source.
15. The microscope of claim 13 , wherein the movable optical element is refractive and shifts a centerline of the light from the light source.
16. The microscope of claim 13 , wherein the scanning-de-scanning assembly is configured to fix a position of said light-redirecting element at a time when said movable optical element is selectively positioned to provide a broad illumination beam extending across said back aperture (confocal mode).
17. The microscope of claim 13 , wherein the scanning-de-scanning assembly is configured to fix a position of said light-redirecting element in response to a confocal mode control command that selectively positions said movable optical element.
18. The microscope of claim 13 , wherein the scanning-de-scanning assembly includes:
image forming optics, with an optical axis, that, in scan mode, generate an intermediate real image of image light returned from said illumination sheet and a detection module that images said intermediate real image, the light sensor including a linear or two-dimensional array of pixels onto which an image of said intermediate real image is focused;
the detection module having an optical axis that is oblique to said image forming optics optical axis, whereby one or selected ones of said pixels may function as a confocal pinhole or slit in confocal mode and whereby, in scan mode, the angle between said detection module and image forming optics optical axes is effective for correcting for variations in axial positions of points along said illumination sheet that are imaged by said detection module.