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Glutaraldehyde and uranyl acetate dual fixation combined sputtering/unroofing enables intracellular fatty acids
Chi Zhang1, Makoto Horikawa1,2,3, Tomoaki Kahyo1,2
1Department of Cellular and Molecular Anatomy, Hamamatsu University School of Medicine, 1-20-1 Handayama, Higashi-ku, Hamamatsu, Shizuoka 431-3192, Japan.
Abstract:
Fatty acids (FAs) have diverse functions in cellular activities. The intracellular distribution of FAs is critical for their functions. Imaging of FAs by time-of-flight secondary ion mass spectrometry (TOF-SIMS) has been achieved. However, TOF-SIMS images of FAs so far do not have subcellular distribution due to inadequate sample preparation methods. In this study, we developed a chemical fixation method using glutaraldehyde (GA) with uranyl acetate (UA), which preserved cellular structure and intracellular FA distribution well. Combining GA+UA fixation with sputtering-based methods and unroofing-based methods, respectively, we successfully imaged intracellular lipids with the subcellular distribution.
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01:20Preparation of Samples for Electron Microscopy
01:03Fixation and Sectioning
The simplest type of preparation is the wet mount, in which the specimen is placed in a drop of liquid on the slide. A liquid specimen can be directly deposited on the slide using a dropper. Solid specimens, such as skin scraping, can be placed on the slide before adding a drop of liquid to prepare the wet mount. Sometimes the liquid is simply water, but stains are often added...