Open-Top Bessel Beam Two-Photon Light Sheet Microscopy for 3D Pathology

Core Concepts

The author presents the development of Open-Top Two-Photon Light Sheet Microscopy (OT-TP-LSM) as a non-destructive and high-throughput method for 3D pathology, enabling rapid and accurate visualization of cellular structures in various human cancer specimens.

Abstract

The content introduces the innovative OT-TP-LSM technique for 3D pathology, offering enhanced imaging depths and detailed visualization of cells and extracellular matrix. The study showcases applications in skin, pancreas, and prostate cancer specimens, demonstrating the potential for rapid and precise non-destructive 3D pathology. By utilizing deep learning networks for style transfer to virtual H&E images, OT-TP-LSM provides comparable histopathological information to traditional methods without thin sectioning.
The study highlights the technical setup of OT-TP-LSM, emphasizing its ability to generate an extended depth of field light sheet using a Bessel beam. Detailed imaging results from human skin, pancreatic, and prostatic cancers illustrate the capability of OT-TP-LSM to distinguish between normal tissues and cancerous structures with high contrast. Additionally, the use of CycleGAN for virtual H&E staining showcases the potential for enhancing pathologists' interpretation of OT-TP-LSM images.
Overall, this research demonstrates the promising application of OT-TP-LSM in advancing non-destructive 3D pathology by providing detailed cellular information in various human cancer specimens through innovative imaging techniques and deep learning-based image processing.

Stats

High imaging depths were achieved owing to long excitation wavelengths.
Imaging depth was approximately three times higher than conventional one-photon LSM.
Image resolution was measured to be 0.9 μm in all dimensions.
Imaging throughput was 0.24 mm2/s at an acquisition rate of 400 fps.
Imaging depth in human skin was approximately 90 μm in a tilted image plane.

Quotes

"OT-TP-LSM may have the potential for histopathological examination in surgical and biopsy applications by rapidly providing 3D information." - Author

Key Insights Distilled From

Open-top Bessel beam two-photon light sheet microscopy for three-dimensional pathology

by Park,W. Y., ... at www.biorxiv.org 10-20-2023

https://www.biorxiv.org/content/10.1101/2023.10.18.562941v2

Deeper Inquiries

How can OT-TS-LMS address challenges related to spherical aberration?

OT-TP-LSM can address challenges related to spherical aberration by implementing an immersion objective lens with a high numerical aperture (NA). By using an immersion objective lens, the issue of spherical aberration arising from the refractive index mismatch between the liquid prism and air can be minimized. The higher NA of the immersion objective lens allows for improved image resolution and enhanced emission light collection efficiency. This adjustment helps in reducing distortion in imaging caused by spherical aberration, particularly in the imaging arm of OT-TP-LSM.

What are the implications of using CycleGAN for virtual H&E staining beyond pancreatic specimens?

The use of CycleGAN for virtual H&E staining beyond pancreatic specimens has significant implications for histopathological practices. One key implication is that it enables rapid conversion of images obtained through OT-TP-LSM into virtual H&E images, mimicking traditional histological staining without actual tissue sectioning or staining procedures. This technology could revolutionize how pathologists visualize and interpret tissue samples, allowing for quick generation of pseudo-colored images that resemble conventional H&E-stained slides.
Beyond pancreatic specimens, CycleGAN's application opens up possibilities for creating virtual H&E images across various tissue types such as skin, prostate, and other organs. This versatility allows researchers and clinicians to explore 3D cellular structures non-destructively while obtaining detailed histopathological information comparable to traditional methods. Additionally, this approach could streamline pathology workflows by providing a faster alternative to generating stained slides manually.

How might advancements in microscopy technology impact traditional histopathological practices?

Advancements in microscopy technology have the potential to significantly impact traditional histopathological practices by offering new capabilities and improving diagnostic accuracy:

Enhanced Visualization: Advanced microscopy techniques like OT-TP-LSM provide high-resolution 3D imaging capabilities that allow pathologists to visualize cellular structures with greater detail than conventional 2D methods. This enhanced visualization can lead to more accurate diagnoses and better understanding of disease processes.

Non-Destructive Examination: Non-destructive imaging technologies reduce the need for tissue sectioning and staining, preserving valuable samples for further analysis or archival purposes. Techniques like two-photon microscopy enable deep-tissue imaging without damaging tissues, offering a comprehensive view of pathological features.

Virtual Histology: Tools like CycleGAN facilitate virtual staining of tissue samples captured through advanced microscopy systems, enabling rapid generation of pseudo-colored images resembling traditional H&E-stained slides. Virtual histology streamlines image processing workflows and provides pathologists with familiar visual cues for interpretation.

Automation and Efficiency: Advancements in microscopy automation enhance workflow efficiency by enabling high-throughput imaging and analysis of large sample volumes within shorter timeframes compared to manual methods. Automated image processing tools help extract relevant information from complex datasets quickly.

5Interdisciplinary Collaboration: As microscopy technologies evolve, they encourage interdisciplinary collaboration between pathologists, biologists, engineers,and computer scientists working together on innovative solutions at the intersectionof biologyandtechnology.Thiscollaborativeapproachdrivesfurtheradvancesinhistopathologyandbiomedicalresearchbyleveragingcutting-edgeimagingtechniquesandcomputationaltools.
Overall,theintegrationofadvancedmicroscopytechnologiesintotraditionalhistopathologicapracticessignificantlyenhancesthequality,speed,andaccuracyofdiagnosticprocedureswhileopeningupnewavenuesforresearchandclinicalapplications.