Hello,
I am currently doing an exploratory study which requires me to section and stain lungs of mice, however this is new territory for me. This is a representative image of how my staining currently looks. Should this be considered artifacts from improper preservation or sectioning? Or is this indicative of dysplasia and lung inflammation?
After the necropsy, I place the lungs in 10% formalin and they are embedded between one day to one week from the necropsy date.
Good day, dear Jelonia Rumph.
It is good for first try. But, the section alas to thick, and some areas resemble autolysis of tissue and artificial pressure of lung tissue. Did you use buffered formalin. Why have been you waiting for a week? Such small organs like mice lungs enough for fixation over 24-48 hours. And little less time in eosin during staining, it is hard to find nuclei
Best regards,
Dmitry
Dmitry Zinovkin Thank you for your advice! How thick should the sections be?
Dear Jelonia Rumph sorry for delayed answer.
The thicknes of sections over 5-10 Um, some tissues, for example lymph node, need thinner than 5 sections, but mosr tissues are are cutted in this interval.
Dmitry Zinovkin Okay thanks! The sections were about 7Um. One more question. Is the autolysis a result of prolonged formalin exposure?
Dear, Jelonia Rumph , highly likely it is long fixation. Could you also send foto of tissue block?
Hi Jelonia,
From your image it seems that this lung section indeed suffers from damage caused by improper handling and fixation.
You can try the followings:
a. After you have sacrificed the mouse, promptly do the perfusion with 4％ PFA (pH 7.4)
b. Inflate the lung using 1~1.5 ml 4％ PFA (pH 7.4)
c. Fix overnight in PFA at 4℃
d. After overnight fixation you better start the embedding procedure. Alternatively, you can store it in 70％ ethanol at 4℃
e. Apply the eosin for a much shorter period of time.
I hope that these steps will solve your problem. Let me know if you have any other issues. Good luck.
Regards,
Shafiq
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