colorectal carcinoma cell line hct116  (ATCC)

Journal: STAR Protocols

Article Title: Protocol for identifying components of subcellular compartments by antibody-based in situ biotinylation

doi: 10.1016/j.xpro.2025.103803

Figure Lengend Snippet: Detection of biotinylated proteins by streptavidin blot HCT116 cells were treated with DMSO or 20 μM etoposide for 1 h and subjected to in situ biotinylation using an anti-γH2AX antibody. Biotinylation was performed with 0.0015% H 2 O 2 and the indicated biotinylation time (0–300 s) or biotin phenol concentration (0–500 μM). Biotinylated proteins were detected by blotting with streptavidin-HRP. The biotin signal was clearly stronger than in the negative controls.

Article Snippet: Human: HCT116 , ATCC , CCL-247.

Techniques: In Situ, Concentration Assay

Journal: STAR Protocols

Article Title: Protocol for identifying components of subcellular compartments by antibody-based in situ biotinylation

doi: 10.1016/j.xpro.2025.103803

Figure Lengend Snippet: Optimization of avidin pulldown HCT116 cells were treated with 20 μM etoposide for 1 h and subjected to in situ biotinylation using an anti-γH2AX antibody. Biotinylation was performed with 0.0015% H 2 O 2 and 500 μM biotin phenol for 1 min. After lysate preparation, avidin pulldown was performed using the indicated volume of avidin beads for 1.5 h at room temperature. Biotinylated proteins were detected by blotting with streptavidin-HRP. Biotinylated proteins were enriched in the pulldown fraction (PD). Biotin signal was drastically reduced in the flowthrough fraction (FT). In this experiment, 5 μL of beads were not sufficient, and at least 20 μL of beads were needed for a good result.

Article Snippet: Human: HCT116 , ATCC , CCL-247.

Techniques: Avidin-Biotin Assay, In Situ