1. Where 'including paraffin-embedded sections' is not
included in a description, does it mean that the antibody has been
tested and found not to work, or does it mean that it has never
No, it does not mean the antibody has been tested and found not
to work. It is certainly worth trying it. For testing an antibody's
suitability, we recommend performing antigen retrieval on the
sections by heat treatment and using the antibody at a starting
dilution of 1/50 (8ug IgG/ml).
2. If an antibody can be used for paraffin-embedded
sections, what method of antigen retrieval should I
As a starting point, we recommend using sodium citrate buffer
with heat treatment (95°C). However, it is possible that
alternative methods (e.g. enzymatic digestion) may be better for
your sections. We recommend each researcher optimises the antigen
retrieval conditions for their own tissues.
3. What fixation method is best and what dilution of
primary antibody should I use?
For immunofluorescence, we recommend methanol fixation of the
cells and a starting dilution of 1/50 (8ug/ml). For frozen
sections, we recommend acetone fixation of the sections and a
starting dilution of 1/100 (4ug/ml). For formalin-fixed,
paraffin-embedded sections, we recommend antigen retrieval using
heat treatment and a starting dilution of 1/50 (8ug/ml).
To determine the optimal dilution required, we recommend that
each researcher perform a serial titration against relevant
positive and negative controls.