Development of novel optical clearing agent for 3D deep-tissue spatial transcriptomics

Abstract

To understand cell function in relation to histological data, there have been approaches integrating transcriptomics with spatial information However, conventional spatial transcriptomics have limitations of quality compared to single cell RNA sequencing and they are unable to analyze 3D tissue. Optical clearing techniques are required to get 3D tissue images for tissue analysis, but conventional optical clearing methods are accompanied with removal and loss of components in the cell, compromising cellular analysis. In this paper, we propose a novel optical clearing technique based on deep eutectic solvents (DESs) which circumvents the exclusion of cell elements. Our approach utilized not only a reversible crosslinker but also a liquid urea-based DES solution containing solid urea, choline chloride, and other components. We conducted ex vivo and in vitro experiments to assess the preservation of cell morphology and fluorescence expression with DES. From the results, this novel DES agent showed that this agent is able to preserve not only cell morphology but also internal cell contents, thereby providing a potential for further 3D spatial transcriptomic analysis termed Histo-Omics.