Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Human Tissue sections (Nasal Polyp and Tonsil)
Specification
Nasal Polyp and Tonsil
Fixative
Formaldehyde
Antigen retrieval step
Enzymatic - Buffer/Enzyme Used: Pronase
Permeabilization
No
Blocking step
20% DMEM and 1% BSA as blocking agent for 20 minute(s) · Concentration: 1% · Temperature: 21°C
Other product details
Dilution
1/2000
Incubation time
18 hour(s) and 0 minute(s) · Temperature: +4°C · Diluent: Tris Buffered Saline
Secondary antibody
Name
Non-Abcam antibody was used: Biotinylated Rabbit anti-mouse (Fab2)
Host species: Rabbit
Clonality: Polyclonal
Conjugation: Biotin
Host species: Rabbit
Clonality: Polyclonal
Conjugation: Biotin
Dilution
1/400
Additional data
Additional Notes
The tissue was fixed in 10% formalin for 24 hours.
An Avidin Biotin blocking step was used in addition to those already mentioned.
The primary antibody is incubated overnight at room temperature. We find it doesn't make any difference whether this is from midday one day to 9am the next or 4pm to 9am - both will give the same results.
We tried the Abcam mast cell tryptase antibody [AA1] (ab2378) at dilutions of 1:50 up to 1:3000, trying out pronase and comparing it to microwaving with a citrate buffer at pH6.
1:2000 with a pretreatment of pronase for 10 minutes gives the best staining - clear positive staining on the mast cells with a nice clean background in both the nasal polyp and the tonsil.
An Avidin Biotin blocking step was used in addition to those already mentioned.
The primary antibody is incubated overnight at room temperature. We find it doesn't make any difference whether this is from midday one day to 9am the next or 4pm to 9am - both will give the same results.
We tried the Abcam mast cell tryptase antibody [AA1] (ab2378) at dilutions of 1:50 up to 1:3000, trying out pronase and comparing it to microwaving with a citrate buffer at pH6.
1:2000 with a pretreatment of pronase for 10 minutes gives the best staining - clear positive staining on the mast cells with a nice clean background in both the nasal polyp and the tonsil.
Abcam response
Thank you very much for this detailed Abreview.
Best regards,
Guillaume
Best regards,
Guillaume
The reviewer received a reward from Abcam’s Loyalty Program in thanks for submitting this Abreview and for helping the scientific community make better-informed decisions.
Abcam user community
Verified customer
提交于 Sep 28 2010