[Histonet] Frozen section artefact

[Adam Boanas]

Hello,

I have a puzzling artefact that I can't seem to correct in 7 micron frozen sections of rat and mouse Small Intestine. When completing an IHC run, parts of the section look great and the staining has worked fine - on other parts of the same section, the morphology is ruined by the appearance of several spindly striations or smears that run between individual villi and in some cases actually cover the majority of the section. Where this occurs the top half of the villi do not take up either the antibody staining or the Haematoxylin counterstain (which is taken up fine elsewhere). Sometimes the smearing looks so bad that it turns the section into a smeary, gloopy mess. >From what I can see the origin of the smears appears to be the nuclei as I have seen several small spindles from the nuclei leading into a larger "thread".
We have thought about Lysis, Gut Mucus (have stained PAS to highlight), nuclear degredation.
Could it be fixation? We air dry following sectioning and fix in Acetone / alcohol 3:1 for 5 mins.
Any ideas would be great! Thanks in advance.
Adam

Adam Boanas
Senior Research Associate
Epistem Ltd
48 Grafton Street
Manchester, M13 9XX



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