Heat-Induced Epitope/Antigen Retrieval.
Hi. I'm doing MMP-2 and MMP-9 IHC on mouse lung tissue. I haven't been successful at getting good, consistent staining.... either no staining or too much background. A few people suggested HIER, however, the tissue is not formalin-fixed, nor paraffin-embedded. Once I harvest the tissue, I perfuse the lungs (through the pulmonary arteries) with a 50:50 mixture of OCT (TissueTek) and PBS. Then I freeze them in OCT. Then I section them at 4 or 5 microns in thickness. I store the slides at -20C and when I'm ready to do an experiments, the first thing I do is fix them in cold aceton for 10-15 min, dry the slides for 5-10 min, then rehydrate (on a shaker to remove the OCT) in either dH2O or TBS, depending on the stain I'm doing. So, does anyone think I still need to do the HIER step? From what I read about HIER is that it's used mainly with FFPE tissue, since that's what masks the antigenicity. Thank you for your time.
Nidal E Muvarak
Associate Research Specialist
Department of Biomedical Engineering
University of Wisconsin-Madison
1550 Engineering Dr.; Rm. 2158
Madison, WI 53706-1609
Lab: (608) 265-8921; Office: (608) 265-4205;
Home: (608) 256-7934; Cell: (608) 332-6068
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