Colon tissues were harvested and fixed in 10% formalin. Deparaffinized colon sections were then obtained and incubated with mouse anti-IRF4 Ab (Invitrogen) followed by Alexa546-conjugated anti-mouse IgG Ab (Invitrogen) by using Can Get Signal immunoreaction enhancer solution (Toyobo) as described previously 43 (link). For visualization of interaction between IRF4 and RICK, MyD88, TRAF6 in colon samples, a Duolink In Situ kit was used (Olink Bioscience). Goat anti-IRF4 Ab (Santa Cruz), rabbit anti-RICK Ab, rabbit anti-MyD88 Ab, and rabbit anti-TRAF6 Ab (Abcam) were used to visualize target protein interactions. For visualization of interaction between IRF4 and RICK, MyD88, TRAF6 in human DCs, cells were fixed in 4% paraformaldehyde and subjected to the protocols suggested by the Duolink In Situ kit.
Protocol detail
Visualizing IRF4 Interactions in Colon Samples
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Corresponding Organization :
Other organizations : Center for Innovation, National Institutes of Health, Kyoto University, National Institute of Allergy and Infectious Diseases, Tohoku University, Kindai University
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Variable analysis
independent variables
- Presence of mouse anti-IRF4 Ab (Invitrogen)
- Presence of Alexa546-conjugated anti-mouse IgG Ab (Invitrogen)
- Use of Can Get Signal immunoreaction enhancer solution (Toyobo)
- Presence of Goat anti-IRF4 Ab (Santa Cruz), rabbit anti-RICK Ab, rabbit anti-MyD88 Ab, and rabbit anti-TRAF6 Ab (Abcam)
- Visualization of interaction between IRF4 and RICK, MyD88, TRAF6 in colon samples
- Visualization of interaction between IRF4 and RICK, MyD88, TRAF6 in human DCs
- Colon tissue samples
- Human DCs
- Fixation of colon sections in 10% formalin
- Deparaffinization of colon sections
- Fixation of human DCs in 4% paraformaldehyde
- No positive or negative controls were explicitly mentioned in the provided information.
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