Optical tissue sectioning using full field optical coherence tomography

1. A multimodal optical sectioning microscope for full-field imaging of a volumic and scattering sample comprising:

a full-field Optical Coherence Tomography (OCT) system for providing an image of a first section in depth of the sample comprising a first illumination sub-system and a full-field imaging interferometer, wherein the full-field imaging interferometer comprises:

a first beamsplitter for defining a reference arm and an object arm for receiving the sample, wherein the reference arm comprises a reference mirror and the object arm comprises a microscope objective,

a detection sub system, and

an optical conjugation device for optically conjugating the sample and said detection sub system, wherein said optical conjugation device comprises the microscope objective;

a supplementary full-field optical sectioning imaging system for providing a fluorescent image of a second section in depth of said sample comprising a structured illumination microscope, wherein the structured illumination microscope comprises:

a second illumination sub system separate from the first illumination sub system,

an imaging channel comprises the microscope objective of the full-field imaging interferometer and means for generating at the focal plane of said microscope objective a variable spatial pattern illumination, wherein the imaging channel is optically coupled with the object arm of the full-field imaging interferometer using the first beamplitter, and

a detection sub system, optically conjugated with said focal plane of the microscope objective; and

a second dichroic beamsplitter, wherein light emitted by the first and second illumination sub systems is combined in the object arm using the first beamsplitter and the second dichroic beamsplitter.

2. The multimodal optical sectioning microscope according to claim 1 , wherein the supplementary full-field optical sectioning imaging system and the full-field OCT system have the same detection sub-system.

3. The multimodal optical sectioning microscope according to claim 1 , wherein the supplementary full-field optical sectioning imaging system and the full-field OCT system have two separate detection sub-systems, and wherein the multimodal optical sectioning microscope further comprises a dichroic beamsplitter able to separate light retro diffused by said first section and fluorescent light emitted by said second section.

4. The multimodal optical sectioning microscope according to claim 1 , wherein said means for generating the spatial pattern comprises a mask with variable spatial transmittance or reflectance to be illuminated by the illumination sub-system of said supplementary full-field optical sectioning imaging system and conjugated with the focal plane of said microscope objective.

5. The multimodal optical sectioning microscope according to claim 1 , wherein the second illumination sub-system of said supplementary full-field optical sectioning imaging system is a laser, and said means for generating the spatial pattern comprises a diffuser to be illuminated by said laser and conjugated with the back aperture of said microscope objective.

6. A method for acquiring in depth section images of a first section of a volumic and scattering sample using a Full-Field Optical Coherence Tomography (FF-OCT) system comprising a full-field imaging interferometer and in depth section fluorescent images of a second section of said sample using a supplementary full-field optical sectioning imaging system comprising a structured illumination microscope, the method comprising:

acquiring the images of said first section of the sample by:

emitting light using a first illumination sub system,

defining, in the full-field imaging interferometer, using a first beamsplitter, a reference arm and an object arm for receiving the sample, wherein said reference arm comprises a reference mirror and said object arm comprises a microscope objective,

optically conjugating said sample with a detection sub-system of the full-field imaging interferometer using the microscope objective; and

acquiring fluorescence images of said second section of the sample by:

emitting light using a second illumination sub-system, separate from the first illumination sub-system, wherein the light emitted by said first and second illumination sub-systems is combined in the object arm of the full-field imaging interferometer using a second dichroic beamsplitter and said first beamsplitter,

optically coupling using said first beamsplitter an imaging channel of said structured illumination microscope with the object arm of the full-field imaging interferometer, wherein said imaging channel comprises said microscope objective and means for generating at the focal plane of said microscope objective a variable spatial pattern illumination,

generating at the focal plane of said microscope objective of said full-field imaging interferometer a variable spatial pattern illumination,

forming a plurality of images for different spatial pattern illumination on a detection sub-system optically conjugated with the focal plane of said microscope objective, and

processing said plurality of images to obtain said fluorescence images.

7. A method according to claim 6 , further comprising simultaneous acquisition of images from said supplementary full-field optical sectioning imaging system and said FF-OCT system.

8. A method according to claim 7 , wherein simultaneous acquisition is obtained using spectral separation of light retro diffused by said first section and fluorescent light emitted by said second section.

9. A method according to claim 7 , wherein simultaneous acquisition is obtained using a multispectral array detector common for said supplementary full-field optical sectioning imaging system and said FF-OCT system.

10. A method according to claim 7 , wherein simultaneous acquisition is obtained using a single detection sub-system and the discrimination of data coming from said supplementary full-field optical sectioning imaging system, and wherein said FF-OCT system is performed using post processing of data acquired by said detection sub-system.

11. A method according to claim 6 , further comprising displaying said first and second fluorescent images, and displaying merged images from said first and second fluorescent images.

12. A method according to claim 6 , wherein acquiring images of said first section of the sample using the FF-OCT system further comprises acquiring using said imaging interferometer at least a first and a second interferometric images and processing said interferometric images to obtain said in depth section image of said first section.