Optical Projection Tomography with a Tissue Clearing Agent for Developmental and Reproductive Toxicology Studies

Abstract

3Background: Developmental and reproductive toxicology (DART) testing represents an expensive and time-consuming stage in determining the toxicological profile of new chemical entities. Within DART studies, morphological evaluation of fetal skeletons for developmental abnormalities typically requires 7 to 14 days. Current processing techniques involve digestion of soft tissue using a strong base (KOH), followed by qualitative assessment of the remaining skeletal tissue by a fetal morphologist. Microcomputed tomography (micro-CT) has been proposed as a nondestructive image-based alternative for quantitative assessment of skeletal morphology. Such methods eliminate the need for extensive tissue processing and can be paired with quantitative analysis algorithms. However, due to the significant capital and operational expenses required for micro-CT imaging, this approach has yet to gain widespread traction and regulatory acceptance. Methods: A novel tissue clearing agent was used in 1-week-old rats to render soft tissue optically transparent. Alizarin red was used to stain the skeleton.High dynamic range optical trans-illumination images were then acquired with an optical-CT imaging system and rendered as three-dimensional images of skeletal structure. Results: High dynamic range-based optical-CT imaging of chemically cleared tissues can rapidly generate high resolution (50-250 lm) reconstructions of whole skeletons. Conclusion: In summary, this study demonstrates that the combination of tissue clearing, optical imaging, and novel reconstruction algorithms may present a new paradigm for highthroughput evaluation of tissues in DART testing.

Current Status of Tissue Clearing and the Path Forward in Neuroscience

“…When coupled with tissue clearing, OPT enables rapid, bright imaging of centimeter-scale samples with micron resolution . For example, Magsam et al used OPT to produce a low-resolution 3D whole mouse brain model in about 5 min after clearing with the BABB protocol. Recently, a label-free OPT technique visualized the whole cleared mouse embryo at different developmental stages and successfully reconstructed the volumetric bone structures in a cleared mouse embryo, a powerful tool for high-throughput phenotype screening and animal development studies …”

Three‐dimensional tissue volume generation in conventional brightfield microscopy

“…Another alternative for 3D tissue volume reconstruction has been presented for whole slide imaging stacks using registration algorithms (Paknezhad et al, 2020). However, the technologies that have been presented in literature (Belay et al, 2021; Davis et al, 2019; Du et al, 2020; Liu et al, 2019; Magsam et al, 2018; Nguyen et al, 2017; Prunskaite‐Hyyrylainen, 2019; Sharpe, 2003; Vallejo Ramirez et al, 2019; Zhang et al, 2020) a/ require the sample to be fully rotated around the light source, b/ necessitate the design of a special instrument devoted for this purpose and c/ have limited application in clinical practice.…”