Im trying to optimise a fixation protocol for inflated mouse lungs using zinc based fixative. I understand that for ZBF fix the tissue pieces should be `3mm, so for my first experiment I did a whole lung inflated with ZBF and submerged in ZBF for 24hr. The next day, I began resecting the lobes into 3mm pieces for a further 24 hr ZBF fix- only to loose the inflation.
Ive heard that acetic acid may improve the penetration of fixative; my goal is to keep the lungs whole throughout a ZBF fix. Can anyone advise if addition of acetic acid to the ZBF may improve the fixation so I can proceed to paraffin without resection?
Thanks in advance,
Judith
I looked up a Nucleic Acids Research paper that talked about the usefulness of zinc based fixation. Interestingly, while the authors demonstrate the isolation of DNA and RNA and 2D protein analysis from the tissue, they do not show any micrographs of histology nor immunocytochemical images. The fact that your lung sample loses it inflated state, even after 24 h in this "fixative", indicates to me that it is not a very good fix for what you a attempting. I would look for an alternative fix. Adding acetic acid will alter the proteins, thus precluding any immunofluorescence, though it will assist in fixation (if not be the only "fixative" in the solution) but the tissue will only be useful for H and E or other general histological staining approaches. For example, Carnoy's fix which contains acetic acid and ethanol gives excellent nuclear fixation but prevents any other type of staining other than general observation type stains. To me, you are probably better off using a more standard approach to fixation which will allow for the lung tissue to remain inflated and will permit you to do a more diverse array of histological approaches for your study.
- Badges
- Science topic
- Similar topics
- Social Psychology
- Attitude