BioMEMS Resource Center, Center for Engineering in Medicine, & Department of Surgery, Massachusetts General Hospital, Harvard Medical School, Boston, MA, USA.
Centers for Disease Control and Prevention, Atlanta, GA, USA.
Cryobiology. 2021 Apr;99:106-113. doi: 10.1016/j.cryobiol.2020.12.007. Epub 2020 Dec 28.
Laboratory rearing of mosquitoes is commonly practiced by researchers studying mosquito-borne infectious diseases and vector control methods. In the absence of cryopreservation methods to stabilize unique or genetically modified strains, mosquito lines must be continuously maintained, a laborious process that risks selection effects, contamination, and genetic drift. Towards the development of a cryopreservation protocol, several commonly used cryoprotectants were systematically characterized here both individually and as cocktails. Among first instar, feeding-stage An. gambiae and An. stephensi larvae, cryoprotectant toxicity followed the order of dimethyl sulfoxide > ethylene glycol > methanol. The resulting LD values were used as the basis for the development of cryoprotectant cocktail solutions, where formulation optimization was streamlined using Taguchi methods of experimental design. Sensitivity to hypothermia was further evaluated to determine the feasibility of cryoprotectant loading at reduced temperatures and slow cooling approaches to cryopreservation. The information described here contributes to the knowledge base necessary to inform the development of a cryopreservation protocol for Anopheles larvae.
实验室饲养蚊子通常是研究蚊媒传染病和病媒控制方法的研究人员的做法。在缺乏稳定独特或基因改良品系的冷冻保存方法的情况下，必须不断维持蚊子品系，这是一个费力的过程，存在选择效应、污染和遗传漂移的风险。为了开发冷冻保存方案，本文系统地研究了几种常用的冷冻保护剂，包括单独使用和作为混合物使用的情况。在第一龄、摄食阶段的冈比亚按蚊和斯蒂芬斯按蚊幼虫中，冷冻保护剂毒性的顺序为二甲基亚砜>乙二醇>甲醇。根据获得的 LD 值，开发了冷冻保护剂混合溶液，利用实验设计的田口方法简化了配方优化。进一步评估了对低温的敏感性，以确定在降低温度和缓慢冷却方法下进行冷冻保护剂加载的可行性。这里描述的信息有助于为冈比亚按蚊幼虫的冷冻保存方案的开发提供必要的知识库。
