19 captures17 Jan 2002 – 31 Dec 2010successfailAbout this captureCOLLECTED BY Organization: Internet Archive The Internet Archive discovers and captures web pages through many different web crawls.At any given time several distinct crawls are running, some for months, and some every day or longer.View the web archive through the Wayback Machine. Collection: Wide Crawl started October 2010 Web wide crawl with initial seedlist and crawler configuration from October 2010 TIMESTAMPS
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Before we start to look at anything it is just as well to review how a slide comes to be. The whole idea is to cut a slice of the tissue to be examined so thin that you can shine light through it. To do this it must be stiff enough not to buckle when the knife hits it. Basically there are three ways of doing this: Touch preparation (smear), frozen section or impregnation section. Touch preparation is what is sounds like – dab a slide on top of the tissue and most often the cells will stick to the slide in a layer thin enough to look through. In other words you avoid the problems of section completely. Frozen section is also conceptually pretty easy. Freeze the tissue and cut the slides in the ice cream section of your supermarket. Impregnation is a mite more complex. Mostly we use plastic or wax. The workhorse of a Diagnostic Lab is wax, so let’s follow that though. The idea is to permeate the tissue with molten wax, let it solidify and then cut it, wax and all. In order to do this one needs to have all the nooks and crannies of the tissue occupied by some wax solvent preferably volatile enough to boil off at, or slightly above, the melting point of wax (about 60oC) so as to drag the wax inside. Most@@ J labs in the USA use Xylene. In order to get the xylene in, one needs to get rid of the water that is naturally in the tissue. This is usually done by using alcohol (superconcentrated vodka will do fine). Tissue is removed from the body and unless you do something quickly it will start to rot, either from its own enzymes or infection. To stop this we fix the tissue in an enzyme poison. Most often we use formalin, but picric acid and mercury salts also work well. Some people use microwaving (just like your handy snack heater). Most fixatives also make the tissue stiffer so handling is easier. So take the tissue, fix, dehydrate, put in Xylene, place in molten wax, allow wax to cool and cut. Place flake of wax containing tissue on a slide then put in an oven, melt off the wax and the tissue is ready to stain.~ The stain we use most of the time is the combination of hematoxylin and eosin. Hematoxylin is an extract of Spanish oak tree bark that is water soluble, dark purple, base that binds to acids in the tissues mostly nuclei acids. Thus it stains all nuclei and the rough endoplasmic reticulum. Eosin was once an extract of beetles (there is now a synthetic equivalent) which is a red, alcohol soluble, acid which binds with bases in the tissue. So to stain the slide – dump it in hematoxylin, then place it in alcohol, then in eosin and now we need to place it in some medium with the same refractive index as glass so that Newton’s rings do not spoil the view when we place it under a microscope. Most of the mounting media are soluble in Xylene so dump some zylene containing some glue with a glass-like refraction on the top of the stained tissue on the slide, put a cover slip on top and you are done! Notice that if there was anything in the tissue that is water, alcohol or xylene soluble then it will not be on the slide for you to see. Also note that you will not see anything that does not bind with either eosin or hematoxylin. For example if there was a lot of edema fluid in the tissue you will not see it because it would be removed by alcohol. If a cell was full of glycogen when we started it will just show as an empty hole by the time we finish. Fat will also be gone. Most organisms do not stain with either H or E so you will have a difficult time seeing them. One other thing and we will actually look at a slide or two. The method of preparation will alter the size of the tissue. Thus a polymorph on a touch preparation will be anywhere between 14 and 20 microns in diameter depending on how hard you pushed and on how long the slide took to dry. In a frozen section a polymorph measures about 10 microns across and in wax about 8 microns.
