I am facing problem in getting good quality paraffin embedded tissue sections. I have prepared paraffin blocks of tissues following a standard protocol. However in some blocks I am unable to get proper ribbons and fine tissue sections (of 5 micron). Although I have used new microtome blade but ribbons and tissue sections are wrinkled, curled and in some instances tissues became powdered instead of a flat-thin section. Please suggest me, what might be the problem and possible solution for the same. I would be thankful if anyone can share a standard protocol for processing of organs (for formalin fixed tissues) and tumor tissue for H&E staining.
Dear Shivani
you can use the protocol of Luna, 1968.. If you want a good sectioning slides you must use a stander materials, like alcohol especially ethanol not methanol, Zylen, liquid paraffin, also the stains eiosin alcoholic or watery, Hematoxyline . Furthermore, the time or period in each steps must be noticed carefully. and good luck
Dr. Suzan
I am with Dr. suzan about her opeion and the most useful protocol is Luna 1968
Dear colleague
you can dissolve the problem by using chemical materials with high quality and especially the ethyl alcohol and stains
Good Luck
hi
if you are sure that all steps before sectioning were good and followed the standards, put your blocks in freezer for 30 minutes and keep using ice cube while cutting.
Good luck.
don't miss inform us about your results..
Hi
Good quality parafin sections rely on several factors including the chemicals (fixative, ethanol, xylol, paraffin, and H&E stains), time required for processing and instruments. If you prepare good blocks, keep your blocks in freeze and set the knife of your microtom by suitable angle. After sections put the ribbons in water path to remove rinkels and deparaffinize it by putting it in the oven 65C for 30 minutes and in the xylol for 15 minutes to complete deparaffinization prior to staining procedures. Plz inform me about your results.
Good luck.
Hi, As for good quality of paraffin embedded tissue sectioning you should first mature the wax that you are using for embedding. For full maturation you can first boil the wax at about 80 degree Celsius in water bath for over night. Then second time when u are using it then u can melt it again.
Thanks & Good Luck
Second thing is that some tissues are more difficult to section than others. Hard tissue (such as bone) and cavernous tissues (such as liver or heart) present special challenges when sectioning. In addition, less than ideal fixation and embedding techniques can make sectioning difficult.
So you should check these steps again that whether u done some mistakes during these steps.
1. Fix Properly
2. Moist the sample
3. Proper chilling of blocks
4. Use new and proper blades
5. Floating of sample in water bath
Some of these steps you check it that either these steps are correct and for proper timing.
Good Luck
Hope this might be useful to you.
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