Ahmadkhani Nima, Benson James D, Eroglu Ali, Higgins Adam Z
School of Chemical, Biological, and Environmental Engineering, Oregon State University, Corvallis, USA.
Department of Biology, University of Saskatchewan, Saskatoon, Canada.
Sci Rep. 2025 Jan 13;15(1):1862. doi: 10.1038/s41598-025-85509-x.
Cryoprotective agent (CPA) toxicity is the most limiting factor impeding cryopreservation of critically needed tissues and organs for transplantation and medical research. This limitation is in part due to the challenge of rapidly screening compounds to identify candidate molecules that are highly membrane permeable and non-toxic at high concentrations. Such a combination would facilitate rapid CPA permeation throughout the sample, enabling ice-free cryopreservation with minimal toxicity. This study presents a new method for rapidly assessing the cell membrane permeability and toxicity of candidate CPAs. The new method enables ~ 100 times faster permeability measurement than previous methods, while also allowing assessment of CPA toxicity using the same 96-well plate. We tested five commonly used CPAs and 22 less common ones at both 4 °C and room temperature, with 23 of them passing the screening process based on their favorable toxicity and permeability properties. Considering its advantages such as high throughput measurement of membrane permeability along with simultaneous toxicity assessment, the presented method holds promise as an effective initial screening tool to identify new CPAs for cryopreservation.
冷冻保护剂（CPA）毒性是阻碍用于移植和医学研究的急需组织和器官冷冻保存的最主要限制因素。这种限制部分归因于快速筛选化合物以鉴定在高浓度下具有高膜渗透性且无毒的候选分子所面临的挑战。这样的组合将有助于CPA在整个样品中快速渗透，实现毒性最小的无冰冷冻保存。本研究提出了一种快速评估候选CPA细胞膜渗透性和毒性的新方法。该新方法的渗透性测量速度比以前的方法快约100倍，同时还能使用同一96孔板评估CPA毒性。我们在4°C和室温下测试了5种常用的CPA和22种不太常用的CPA，其中23种基于其良好的毒性和渗透性特性通过了筛选过程。考虑到其具有膜渗透性高通量测量以及同时进行毒性评估等优点，所提出的方法有望成为一种有效的初始筛选工具，用于鉴定用于冷冻保存的新型CPA。
