Sigma-Aldrich
Monoclonal ANTI-FLAG® M2 antibody produced in mouse
F1804
Browse Similar Products: FLAG Western blot Antibodies
The goal of this experiment was to immunoprecipitate a FLAG-tagged protein that was over-expressed in HEK293T cells. Based on my previous experience working with this antibody from Sigma-Aldrich, I expected that this antibody should be a good choice for this experiment.
Immunoprecipitation
HEK293T cell whole cell lysate
2 µg per 2mg lysate for immunoprecipitation and incubated for 1 hr at 4 deg; 1:2000 dilution for Western Blotting incubated for 16 hr at 4 deg
5% Milk powder in TBS-T (0.1%) for Western Blotting
1:5000 dilution with Donkey-anti Mouse HRP antibody for 1 hr at room temperature
NA
ECL
1. FLAG tagged Evc protein was over-expressed in HEK293T cells for 40 hr. 2. Whole cell lysates were prepared in RIPA lysis buffer. 3. 2 mg of clarified extract was incubated with 2 µg of FLAG antibody for 1 hr at 4 deg on a rotator. 4. 10 µl of Protein G Sepharose beads were addred and incubated for 1 hr at 4 deg on a rotator. 5. Beads were extensively washed in RIPA lysis buffer and bound protein complexes were eluted in SDS sample buffer. 6. Western Blotting was done with the same antibody (1:2000 dil) and a good signal was obtained.
None
Very good for immunoprecipitation
A clean antibody for both immunoprecipitation and Western Blotting