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mouse liver, IHC-P, high background in endothelial cells. Paraffin embedded sections, negative control also showing staining, secondary antibody is fine. H202 block 15 min, protein block 5 min
Asked on Sep 20 2012
Thank you for contacting us.
You had previously been in contact with my colleague heather regarding staining observed in your negative control using these mouse monoclonal Cytokeratin antibodies on mouse tissue. To clarify, is your negative control from a tissue that is not expected to express cytokeratin or is it an isotype control or a no primary antibody control?
When using an anti-mouse secondary antibody on mouse tissue, background staining is often observed due to cross-reactivity with endogenous mouse IgGs in the sample. This can be checked by testing a no-primary antibody control slide.
If you are observing mouse on mouse background staining, this can be avoided by either using a mouse on mouse (MOM) staining kit, such as ab127055. The background can also be blocked by adding an incubation with a F(ab) fragment unconjugated anti-mouse secondary antibody, such as ab6668. The F(ab) fragment will coat any endogenous IgGs in your sample, and can be done just before the serum block at a 1:500 dilution for 1 hour at room temperature.
I hope this helps, if not, please let me know and I will be happy to help you further.
Answered on Sep 20 2012