Cells were treated with RIPA buffer (Beyotime, China) to obtain the total cell proteins, which were quantified using the micro bicinchoninic acid (BCA) method. Proteins (50 μg) were separated by SDS-PAGE, transferred to PVDF membranes, and sealed with 5% skimmed milk at 37°C for 2 h. Then, the membrane was combined with RNF6 (ab204506; Abcam, USA) or GAPDH (ab9484; Abcam) antibodies overnight at 4°C. The secondary antibody was horseradish peroxidase-labeled goat anti-rabbit IgG (ab205718; Abcam). The ECL method was used to detect the signal, the UVI gel imaging system was used to collect the image, and Image J software (Media Cybernetics, USA) was used to analyze the gray value.
Ab204506
Manufactured by Abcam
Sourced in United States
Ab204506 is a recombinant monoclonal antibody that recognizes a specific protein target. It is intended for use in laboratory research applications.
Automatically generated - may contain errors
Lab products found in correlation
Ab9484, by Abcam (1 mentions)
Ab205718, by Abcam (1 mentions)
Ripa buffer, by Beyotime (1 mentions)
Imagej software, by Media Cybernetics (1 mentions)
Ab16895, by Abcam (1 mentions)
Ab78393, by Abcam (1 mentions)
Ab9110, by Santa Cruz Biotechnology (1 mentions)
Ab170901, by Abcam (1 mentions)
Ab94483, by Abcam (1 mentions)
Ab7780, by Abcam (1 mentions)
Anti six3, by Santa Cruz Biotechnology (1 mentions)
2 protocols using ab204506
1
Western Blot Analysis of RNF6 Protein
2
Investigating SIX3 Protein Regulation via Co-Immunoprecipitation
Check if the same lab product or an alternative is used in the 5 most similar protocols
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The details of these procedures were described previously [37 (link)]. Anti-SIX3 (Santa Cruz Biotechnology, Dallas, TX, USA; sc-81985), anti-FLAG (Abcam, Cambridge, MA, USA; ab205606), anti-HA (Abcam; ab9110), and anti-IgG (Santa Cruz Biotechnology; sc-2027) antibodies were used for co-immunoprecipitation (Co-IP). Immunoprecipitates were washed at least five times and subjected to western blot analysis using anti-TRIM27 (Abcam; ab78393), anti-NEDD4 (Abcam; ab236512), anti-SMURF2 (Abcam; ab94483), anti-RNF6 (Abcam; ab204506), anti-SYVN1 (Abcam; ab170901), anti-MDM2 (Abcam; ab16895), and Anti-SIX3 (Abcam; ab172131) antibodies. For ubiquitination assays, the lysates of A549 cells transfected with siTRIM27-1 or siNC were used for IP with an anti-IgG (Santa Cruz Biotechnology; sc-2027) or Anti-SIX3 antibody (Santa Cruz Biotechnology; sc-81985) and Protein A/G PLUS-Agarose (Novex, Oslo, Norway), which was performed at 4° C overnight. The eluted proteins were then detected by western blot analysis using an anti-ubiquitin (Ub) antibody (Abcam, ab7780).
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