I am trying to perform HE staining of small intestine. I used OCT solution to make sections.
After HE staining when I look my slides under microscope, I do not get intact villi. In much of the cases, villi are broken. Does anyone can help me?
Dear Sailesh
Did you make any chemical fixation?
How did you make the dissection?
Dear Robson Campos Gutierre,
After washing the intestine with PBS, I fixed the intestine by keeping it in 10% formaldehyde neutral buffer solution for overnight.
why did you use OCT? if you fixed the tissue in formalin, then you had to proceed with FFPE preparation. OCT and formalin are not compatible. if you need to preserve villi, make FFPE .
You can achieve great villi structure using the fixation method, but after fixation you must dehydrate with graded alcohols and clear (Xylene) and then impregnate with paraffin wax. Embed in a mold with more paraffin. Cut on microtome, float section out on water bath, pick up on slide, dry section in oven long enough to remove water and then dewax (xylene) rehydrate (graded alcohols), water and then stain.
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