冷冻扫描电子显微镜冷冻断裂法在生物样品观察中的应用  

Application of Freeze Fracture Method of Cryo-Scanning Electron Microscopy in Observation of Biological Samples

作　　者：黄克让 王珍[1] 陈蕾[1] 崔凤展 彭潔莹 张敏[1] HUANG Ke-rang;WANG Zhen;CHEN Lei;CUI Feng-zhan;PENG Jie-ying;ZHANG Min(Division of Laboratory Safety and Services College of Food Science and Engineering of Northwest A&F University,Yangling 712100,Shaanxi China)

机构地区：[1]西北农林科技大学实验室安全与条件保障处食品科学与工程学院,陕西杨凌712100

出　　处：《分析测试技术与仪器》2022年第4期433-438,共6页Analysis and Testing Technology and Instruments

基　　金：西北农林科技大学试验技术研究与实验室管理创新项目(22800/A1070221004)。

摘　　要：冷冻扫描电子显微镜(Cryo-SEM)具有能在高真空状态下观察含水样品、分辨率高、制样简单快速、可对样品进行断裂刻蚀等优点,是生命科学研究中的有力工具.利用Cryo-SEM冷冻断裂法进行生物类样品表面形貌观察,既能观察到表面结构信息,又可观察样品内部结构信息.以沙漠梭梭成熟分支中上部同化枝的植物样品和乳清蛋白纤维(WPIF)的液体样品为试验材料,对样品进行冷冻断裂处理,观察并对比冷冻断裂处理前后样品的结构.结果表明:植物样品冷冻断裂后在升华温度为-100℃、升华时间10 min、冷台温度为-175℃、观察电压5 kV的试验条件下可以观察到植物细胞的叶绿体和细胞之间的细毛组织. WPIF冷冻断裂后在升华温度为-100℃、升华时间5 min、冷台温度为-175℃、观察电压5 kV的试验条件下可以观察到中链甘油三酯和蛋白纤维堆积而成的片层结构.未冷冻断裂处理的样品无法观察到以上结果.Cryo-scanning electron microscopy(Cryo-SEM) is a powerful tool in the life science research because of its advantages such as observing aqueous samples under high vacuum condition, high resolution, simple and rapid sample preparation, fracture etching of samples, etc. Through the surface morphology observation of biological samples using Cryo-SEM’s freeze fracture method, both the surface structural information and the internal structural information of the samples can be observed. The plant samples from the upper and middle assimilated branches of Haloxylon Ammodendron and the liquid samples of whey protein fiber(WPIF) were taken as the experimental materials, and the samples were subjected to a freeze fracture treatment to observe and compared the structures of the samples before and after the freeze fracture treatment. The results showed that the chloroplasts of plant cells and the fine hairs between the cells could be observed under the test conditions of a sublimation temperature of-100 ℃, a sublimation time of 10 min, a cold platform temperature of-175 ℃, and an observation voltage of 5 kV. And the lamellar structure of medium chain triglyceride and protein fiber can be observed under the test conditions of a sublimation temperature of-100 ℃, a sublimation time of5 min, a cold stand temperature of-175 ℃, and observation voltage of 5 kV after freeze fracture of WPIF. However, the above results could not be observed for the samples without a frozen fracture treatment.

关 键 词：冷冻扫描电子显微镜 冷冻断裂 形貌观察 生物样品 

分 类 号：O657[理学—分析化学] TN16[理学—化学]