Chip-based multimodal super-resolution microscopy for histological investigations of cryopreserved tissue sections

Luis E Villegas-Hernández,Firehun T Dullo,Gery Barmettler,Balpreet Singh Ahluwalia,Aud-Malin Karlsson Hovd,Jean-Claude Tinguely,Vishesh Dubey,Kristin Andreassen Fenton,Ganesh Acharya,Åsa Birna Birgisdottir,Anish Priyadarshi,Azeem Ahmad,David A Coucheron,Mona Nystad,Krishna Agarwal,José M Mateos,Urs Ziegler,Sebastian Acuña

doi:10.1038/s41377-022-00731-w

Abstract

Histology involves the observation of structural features in tissues using a microscope. While diffraction-limited optical microscopes are commonly used in histological investigations, their resolving capabilities are insufficient to visualize details at subcellular level. Although a novel set of super-resolution optical microscopy techniques can fulfill the resolution demands in such cases, the system complexity, high operating cost, lack of multi-modality, and low-throughput imaging of these methods limit their wide adoption for histological analysis. In this study, we introduce the photonic chip as a feasible high-throughput microscopy platform for super-resolution imaging of histological samples. Using cryopreserved ultrathin tissue sections of human placenta, mouse kidney, pig heart, and zebrafish eye retina prepared by the Tokuyasu method, we demonstrate diverse imaging capabilities of the photonic chip including total internal reflection fluorescence microscopy, intensity fluctuation-based optical nanoscopy, single-molecule localization microscopy, and correlative light-electron microscopy. Our results validate the photonic chip as a feasible imaging platform for tissue sections and pave the way for the adoption of super-resolution high-throughput multimodal analysis of cryopreserved tissue samples both in research and clinical settings.

Highlights

Histology is the field of biology that studies the microanatomy and structure of tissues regarding their function in organisms
While whole slide imaging scanners provide fast imaging of several histological slides in a day, these automated optical microscopes are limited to a resolution power of ~250–500 nm, which is insufficient for the observation of individual subcellular structures such as nanovesicles, filaments, tubules, brush border, gap junctions and several other intracellular features with an average size below ~250 nm
Chip-based multicolor TIRFM imaging In this part of the study, we used chorionic villi tissue from the human placenta to assess the suitability of the photonic chip for histological observations

Summary

Introduction

Histology is the field of biology that studies the microanatomy and structure of tissues regarding their function in organisms. A typical histological analysis involves the extraction of a tissue sample from the body, fixation, and preservation followed by sectioning and labeling before observation through a microscope. Contrast, and resolution are critical parameters for histological assessment. While whole slide imaging scanners provide fast imaging of several histological slides in a day, these automated optical microscopes are limited to a resolution power of ~250–500 nm The visualization of such features was only possible through other imaging techniques such as electron microscopy[2,3,4], which supports a resolving power down to ~10 nm for fixed and embedded histological samples. While available published methods for fast[5] and automated correlation microscopy exist since years[6], the combination of traditionally lengthy sample preparation

Methods

Results

Conclusion