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UkyoKuonji2004 Vice Captain
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Posted: Sun Sep 26, 2010 7:04 pm
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Posted: Sun Sep 26, 2010 7:13 pm
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Posted: Sun Sep 26, 2010 7:20 pm
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Posted: Sun Sep 26, 2010 7:41 pm
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Posted: Sun Sep 26, 2010 7:47 pm
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Posted: Mon Sep 27, 2010 5:24 pm
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Finally got my report done. I hope it's done right...
Myelin Sheath stain: Weil method
The purpose of this stain is to demonstrate myelin in tissue. When axons degenerate their myelin sheathes are broken down to simple lipids. Those simple lipids are then removed. Syphilis is known to cause demyelination, and would therefore be detected with this stain.
This stain is one to demonstrate myelin, therefore I used slides of spinal cord as a control. Medulla tissue is also good for control as well.
The staining solution used in this method is a mixture of 10% alcoholic hematoxylin, ferric ammonium sulfate, and deionized water. The staining solution has to be prepared fresh. The first differentiating solution in this method is 4% ferric ammonium sulfate. When it's made, it should be good for six months. The solution needed to finish differentiating is a mixture of sodium borate and potassium ferricyanide in deionized water. The last solution you need is diluted ammonia water. When I did this method, I used a ratio of six drops of ammonium hydroxide for every 100 mL of water. It is recommended to have all these solutions ready before you begin.
This stain is a regressive stain, which means you have to overstain your slide and then differentiate and/or decolorize.
The procedure is as follows:
Hydrate the slides to water. First, you place your slides in the staining solution which has been warmed to 54 to 56 degrees Celcius. Maintaining that temperature in the water bath, let the slides stain for half an hour. During this time the mordant-hematoxylin solution will attach to the phospholipid component of the myelin sheath. Once the half an hour is over, take the slides out and rince them in two changes of tap water. When you have finished that, the slides have to be differentiated in the 4% ferric ammonium sulfate solution. The goal here is to remove most of the excess dye. From testing the method out, I found that two one second dips in this solution is sufficient for this task. Once that is done, the slides need to be washed in three changes of tap water. This part of the procedure is a little trickier. The differentiation is completed in this step. The sections are dipped in the sodium borate-potassium ferricyanide solution until the grey matter and the white matter are sharply defined. This should be controlled microscopically. When doing this step, have a coplin jar filled with water handy so you can dip the slide in it before putting it under the microscope. The water stops the differentiation. It could take more than six dips in the differentiating solution before getting the desired results. When you are done, wash the slides in two changes of tap water. Then you treat the slides with the diluted ammonia water. The water is slightly alkaline so make sure you don't dip the slides too vigorously, otherwise the sections will float off the slide and you would have to start over again. Wash in deionized water. From here you dehydrate, clear and mount as normal.
The finished product should show the myelin sheath as blue or bluish black. The rest of the tissue should stain pale tan.
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UkyoKuonji2004 Vice Captain
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Posted: Mon Sep 27, 2010 5:35 pm
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Posted: Mon Sep 27, 2010 5:38 pm
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UkyoKuonji2004 Vice Captain
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Posted: Mon Sep 27, 2010 5:42 pm
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Posted: Mon Sep 27, 2010 5:45 pm
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Posted: Mon Sep 27, 2010 5:47 pm
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Posted: Mon Sep 27, 2010 5:49 pm
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UkyoKuonji2004 Vice Captain
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Posted: Mon Sep 27, 2010 5:52 pm
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Posted: Mon Sep 27, 2010 5:52 pm
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UkyoKuonji2004 Vice Captain
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Posted: Mon Sep 27, 2010 5:53 pm
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