Immunohistochemistry (Frozen sections) abreview for Anti-HA tag antibody - ChIP Grade

Inconclusive
Abreviews
Application
Immunohistochemistry (Frozen sections)
Sample
Mouse Cell (spinal cord)
Permeabilization
Yes - PBS/gelatin/triton/serum
Specification
spinal cord
Blocking step
Serum as blocking agent for 30 minute(s) · Concentration: 10% · Temperature: 20°C
Fixative
Paraformaldehyde

Other product details

Incubation time
15 hour(s) and 0 minute(s) · Temperature: 20°C · Diluent: PBS/gelatin/triton/serum
Dilution
1/650

Secondary antibody

Dilution
1/1000
Name
Non-Abcam antibody was used: Donkey anti rabbit Alexa 555 (Life Technologies)
Host species: Donkey
Clonality: Polyclonal
Conjugation: Alexa Fluor® 555

Additional data

Abcam response
Thank you for your review of this previously untested application, IHC of PFA-fixed, frozen tissue, and for the comparison of results from two batches. For consistent staining after the lengthy fixation (2 weeks in 4% PFA before cryoprotection and freezing, according to the reviewer), an antigen retrieval step before staining may help. For heat-mediated retrieval, a temperature lower than the standard 95-100C may be necessary to minimize damage to fragile tissue. The following reference discusses optimization of an antigen retrieval protocol. J Neurosci Methods. 2000 Dec 15;104(1):27-34.
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Submitted Jul 14 2015

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