The non-treated material is comparatively soft and cannot be cut to make a thin section (about 30 μm thick). And I want to prevent several seconfary metabolites (water and alchol soluble) dissolving from it by various embedding treatments.
Thank you for the reply, Dr. Costa. Yes, I tried frozen sectioning (mounted by distilled water). Unfortunately, however, the sample might be soaked with water in sectioning and water-soluble compound can be dissolved or move in the surface.
I want to try in situ detection of such compounds on the section, therefore I am afraid I cannot detect the compounds in correct localization by frozen sectioning method. but I am glad to have the answer from you. Thank you very much.
Depending on the tissue, you should be able to make a few sections of an appropriate thickness by hand. It takes practice, but I would start by trimming down a chunk and placing it on a slide, with water to keep it from drying out. Then you can place another slide across it as a guide (like a carpenter's fence). Using a fresh razor-blade make your first cut with the blade edge vertical; and then, not moving the fence, make a second cut with the blade edge angled in slightly. With practice you can produce a nice section in which the thickness varies across the width of the section from very thin at what was the top of the specimen to somewhat thicker at the bottom - wedge-shaped. Once you get the technique down it is quite easy to control the thickness nicely. It's true that the thickness is not uniform across the section, but for most microscopical observation at 20X or higher, the field of view is small enough so that you don't really notice the thickness difference. Works well for herbaceous stems, some leaves, and even stems with some secondary wood formation, as long as the blade is sharp. Use a fresh blade every time! You need to trim the specimen carefully so that it lies flat on the cutting surface (the first slide) and make sure your axial orientation is precise so that you get true cross-sections.
Could you use another media?
Maybe this could be of some usefulness: Peters, Stephen R. A practical guide to frozen section technique. Springer, 2010.
Dr. Lintilhac, thank you for your detail explanation of the sectioning method by hand. The material I've treated is Citrus peel. The presence of oil glands in flavedo (pericarp) and fragile albedo (mesocarp) are sometimes troubles in sectioning. Your method seems to become skillful, but also interesting for me. Thank you very much.
Dr. Costa
Thank you for the reference. I'll check it. If I can find appropriate medial, I try it.
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