Optimizing a three-dimensional spheroid clearing method for the imaging-based evaluation of cardiotoxicity

Toxicity evaluation based on two-dimensional cell culture shows differences from clinical results and has the disadvantage of not accurately reflecting cell-to-cell cross-signaling. Since almost all cells in the human body are arranged in a three-dimensional structure and constitute a tissue, the in vitro reproduction of three-dimensional tissues composed of human cells can be used as effective models for drug development and toxicity evaluation. The clearing technique improves image resolution and can be used to generate three-dimensional bio-images throughout the organized structure, improving the efficiency of toxicity evaluation for disease models using spheroids. Herein, we report the first optical spheroid clearing protocol for an image-based toxicity prediction model. In our results, spheroid clearing significantly increased fluorescence intensity and enabled image-based toxicity prediction. We propose that this spheroid clearing method can be utilized for image-based cardiotoxicity evaluation. Furthermore, we also present the possibility that our protocol can be utilized for patient-tailored toxicity prediction.