Light sheet-based imaging device with extended depth of field
Light sheet microscope for recording 3D images of a specimen, comprising a light sheet generator, an objective lens, an image detector and a wavefront encoder element or system positioned between the objective lens and the image detector for extending the depth of field of the objective lens. By doing so, the restriction on the distance between the collecting objective and the light sheet is relaxed, allowing the use of a light sheet scanning unit. The resulting light sheet microscope allows for a robust, aberration insensitive, fast 3D imaging without the need to move or perturb the specimen.
1. A light sheet microscope for recording 3D images of a specimen, comprising:
a light sheet generator and sheet scanning unit that produces a light sheet that is scanned through the specimen,
an objective lens positioned to receive the signal generated by the scanned light sheets and an image detector positioned to receive light from the objective lens,
a wavefront encoder (WFE) element or system positioned between the objective lens and the image detector at an exit pupil plane or a conjugated plane of the objective lens, wherein the WFE extends the depth of field of the objective lens, and wherein the WFE is selected to generate an optical path difference resulting in a phase modulation that permits creating a 3D image from 2D images generated by scanning the light sheet with the scanning unit while the objective lens and the specimen are static.
2. The light sheet microscope according to claim 1 , wherein the wavefront encoder element is a single optical element that is adapted to introduce an optical path difference that results in a phase modulation described by a linear combination of Legendre polynomials, a cubic profile or a polynomial with rotation symmetry.
3. The light sheet microscope according to claim 2 , where the WFE element works in a transmission configuration or a reflection configuration.
4. The light sheet microscope according to claim 2 , where the WFE element is a phase mask.
5. The light sheet microscope according to claim 2 , where the WFE element is an axicon.
6. The light sheet microscope according to claim 2 , where the WFE element or system is actively changed to adapt the depth of field to a specific requirement.
7. The light sheet microscope according to claim 1 , where the wavefront encoder system is a combination of optical elements, the combination adapted to introduce an optical path difference that results in a phase modulation described by a linear combination of Legendre polynomials, a cubic profile or a polynomial with rotation symmetry.
8. The light sheet microscope according to claim 7 , where the WFE system works in a transmission configuration or a reflection configuration.
9. The light sheet microscope according to claim 7 , wherein the wavefront encoding system comprises a deformable mirror and a relay system formed by a set of relay lenses that conjugate the exit pupil of the objective lens to the surface of the deformable mirror whose surface profile is modulated to result in an extended DoF.
10. The light sheet microscope according to claim 9 , wherein a wavefront sensor is placed at a conjugated plane of the surface of the deformable mirror for measuring and controlling the phase modulation introduced by the deformable mirror.
11. The light sheet microscope according to claim 7 where the WFE element or system is actively changed to adapt the depth of field to a specific requirement.
12. The light sheet microscope according to claim 1 further comprising a digital processing unit for decoding by an adequate deconvolution process the 2D images obtained by the image detector and transforming them into 3D images.