The application of highpressure freezing-freeze substitution technique in ultrastructure of nervous tissue
ZHANG Bin;WU Liang-liang;WU Ying-ying;PAN Li-jun;LOU Sen-sen;KONG Yu;Institute of Neuroscience,State Key Laboratory of Neuroscience,Shanghai Institutes for Biological Sciences,Chinese Academy of Science;
To promote the application of high pressure freezing-freeze substitution( HPF-FS) technique in neurobiology,the ultrastructure of nervous tissue of two model organisms( mouse and nematode) were investigated via the sample preparation,ultrathinsectioning and TEM observation. Besides,the influences of three different fixation methods including chemical fixation( CF),high pressure freezing( HPF) and the combinationfixation( CF + FS) to the ultrastructure of nervous tissue were further explored. The results of TEM demonstratethat HPF provides more advantages for nervous tissue and could greatly minimize the artificial illusion to the ultrastructure which given rise to traditional chemical fixatives. In addition,it was also found that the suitable fixation way varies from different nervous tissues. For example,it is suitable for mouse brain tissue that directly to befixed by HPF after biopsy sampling. The resulting ultrastructure is better than CF or CF + HPF in both completeness and exquisite degree of membrane,indicating that there is no need to use chemical fixatives before HPF fixation. Due to its thick integument,the nematode is more suitable to be fixed by direct HPF treatment so as to shorten the residence time before fixation and effectively maintain the ultrastructure of its abdominal neurons.