Hi. Most of the tissues I'm using are embedded in paraffin for HE staining. However, I also need to analyze those tissues via immunofluorescence staining. The tissue is very limited. So can I use the same tissue fixed in paraffin for immunofluorescence?
Parafin embedded tissues can be used for immunofluorescence staining. However, like fresh frozen tissues, there are caveats, specifically your antigen of interest, optimizations by the antibody manufacturer, and your own optimizations for your particular tissue/antigen. Most manufacturers will list and reference whether or not a particular antibody has been tested in paraffin embedded tissue. Additionally, optimization for immunofluorescence markers is more challenging to produce good quality images, with specific positive staining and reduce background/ non-specific stain. Finally, paraffin tissues are usually cut very thin 5-7uM, as such your depth of field is limited.
More information is needed to fully address this question. One additional point, if your question is that of "can you use the same sample and concurrently stain H&E and Immunofluorescence simultaneously..." that would not be recommended.
FFPE tissue is challenging for IF, because the tissue will often have some degree of autofluorescence, which could be difficult to differentiate from your signal. It is possible to go beyond this using image analysis and gating of the autofluorescence wavelengths, but it requires special software.
Yes.
I am sending You a method. It was very useful in my kidney biopsy material.
(in all cases I used the direct IF on frozen sections, and this method was the same positivity.)
Best:
Peter
Yes. Though it is challenging but still the paraffin-embedded tissue can be used for immunofluorescence staining.
- Badges
- Science topic
- Similar topics
- Biological Science
- Molecular Biology
- Molecular Methods
- Cloning