Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

Fluorescence IHC

DR Daniel Ryan
JK Jim Koziol
WE Wael M. ElShamy
request Request a Protocol
ask Ask a question
Favorite

IHC was performed as previously described95. Briefly, 5μm thick paraffin-embedded sections of tumor tissue excised from GemOE orthotopic mammary tumors were deparaffinized, rehydrated, and washed in PBS. Antigen retrieval for all antibodies was done using Sodium citrate buffer (10 mM Sodium citrate, 0.05% Tween 20, pH 6.0), 10 min at 95 °C. Slides were blocked with 10% normal serum (2° antibody species) for 1 h at RT, washed, and probed with 1° antibodies overnight at 4 °C. Slides were exposed to Alexa Fluor 568 (red), and 488 (green) conjugated secondary antibody for 1 h at RT and counterstained and mounted with VECTASHIELD mounting medium with DAPI (Vector) and imaged under the fluorescence microscope.

Copyright and License information: The Author(s) ©2019
Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

Do you have any questions about this protocol?

Post your question to gather feedback from the community. We will also invite the authors of this article to respond.

post Post a Question
0 Q&A