This invention is an optical imaging method for biological specimens based on physically expanding the specimen.
Microscopy has facilitated the discovery of many biological insights by optically magnifying images of structures in fixed cells and tissues. However, the resolution to a light microscope is limited. The existing super-resolution techniques are limited in three aspects: imaging depth, throughput, and complexity/cost. High resolution in conventional imaging techniques is only achieved in thin layers. Additionally, in super-resolution imaging techniques based on single-molecule imaging, the throughput is not high enough to image a large specimen, such as an entire organ. Lastly, super-resolution imaging techniques generally require relatively expansive and complex optical systems. The Inventors addressed these issues by developing Expansion Microscopy, which physically expands the specimen, resulting in physical magnification.
The Inventors have developed a method for optically imaging biological specimens with a resolution better than predicted by the classical diffraction limit, based on physically expanding the specimen. Expansion microscopy involves embedding a tissue into a swellable material that is formed by infusing the tissue with reactive monomers which then undergo polymerization. Expansion microscopy can be performed repeatedly on the same sample, essentially enabling arbitrary magnification. Additionally, proteins can be visualized within the specimen by anchoring fluorophores directly to the expandable gel matrix. Nucleic acid can also be anchored to the gel for both in situ genomic and transcriptomic assessment, as well as to enable nucleic acid barcodes to be used to identify essentially arbitrary numbers of molecules. To adapt the expansion microscopy technology in the clinical setting, the Inventors have designed variants of expansion microscopy that incorporate de-paraffinization, antigen retrieval and aggressive protease digestion into a comprehensive workflow to expand various common clinical specimens. De-paraffinization and antigen retrieval address the recovery of archived clinical samples for immunohistochemistry, while aggressive protease digestion is critical for the success of sample expansion, as most of the human tissues contain abundant hard-to-digest structural proteins.
- Swollen material with embedded biological specimen can be imaged on any optical microscope
- Allows for effective imaging of features below the classical diffraction limit
- Provides a feasible and a robust way to homogenously expand clinical tissue samples