I would like to do high resolution microscopy of fresh tissues, which have NOT been stained or embedded.
You can put the tissue between half cut potato and then by using a new shaving blade no. of thin slices can be made. dip these slices in water and remove the tissue section to put it on the slide and view under microscope by putting cover slide.
Microtomy can be the next option
Some years ago, we needed to make thin slices of potato and carrot (approx. 0.1 mm thin), for studying the diffusion of pollutants through these tissues. We tried different techniques and used different types of blades and also dedicated tools for microscopy work (microtomy). Surprisingly, we obtained the best slices with the ham cutter of our canteen: ”A large number of carrot and potato slices were cut using an electric hamslicer (model Euro 2560S from Graef, Arnsberg, Germany). The slice thickness was measured using a digital micrometer with a resolution of 10 micrometer. Slices with a thickness between 90 and 110 micrometer were selected. Circles with a diameter of 6mm were cut from these slices and stored at 100% humidity” (Trapp et al, 2007, Environmental Science & Technology, 41, 3103-3108). This will only work for some kind of plant tissues, and I am not sure whether these slice are good specimens for your microscopy work. However, for our diffusion work this gave the best results and was very practical. The storage at (near) 100% humidity was very crucial to keep the tissue slices intact (up to some hours), and this was easily done by placing some wet cotton pads in a closed petry dish and then placing the thin slices on the wet cotton or next to them.
Hello, we use razor blad (a new one is better). For stems and roots we don't need a support we just take the sample between fingers but for leaves we usea polyestyrene support to facilitate cuting
Hello, I also used microtome. Size of the section is vary depend on each microtome. I my case, I sliced skin of melon (15 µm thickness) and my rotary microtome can slice sample 1-25 µm thickness. Anyway, we need to prepare sample with dehydration series and embedded in paraffin blocks in the first step.
Hello so if the samles are fixed deshydrated and embeded they are not fresh plant sections any more
Hello, I am sorry. For the fresh samples, I also used microtome (Hooker, plant microtome) and no need to stained or embedded, used fresh sample directly.
I have used a vibratome to get slightly thinner sections than I could get by hand. One advantage of the vibratome is that you get a known thickness and it is constant across the cut and from one section to another. Another is that the sample is in water, so it does not dry out.
I held the tissue in place with agarose as Dr. Bogdanovic suggests, though sometimes it is helpful to stick the tissue down with superglue (cyanoacrylate) before covering with agarose.
The thinnest sections I could get with roots was just under 100 µm. I have often heard that 80 µm is the thinnest most fresh tissue can be sectioned without tearing.
That said, I have seen an experienced hand (Root anatomist Margaret McCully) whip out hand sections that were far superior to anything I could do with this tool. So there is the option of practicing on thousands of samples until you have the feel.
Hi Puthmee,
Could you please explain how you used microtome for fresh sectioning?
Hi i need to make the thin leaf section of soybeans through microtome, can any body guide me about thickness of leaf tissues i mean through the chat, i got it can be 1-25 micron meter thickness and paraffin film can be used for the embedding. However, if anyone of you know the best method please tell me
Thanks
Asifa
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