Flow Cytometry analysis of K562 cells using anti-KMT6/EZH2 antibody (A00050-1). Overlay histogram showing K562 cells stained with A00050-1 (Blue line). To facilitate intracellular staining, cells were fixed with 4% paraformaldehyde and permeabilized with permeabilization buffer. The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-KMT6/EZH2 Antibody (A00050-1, 1 µg/1x106 cells) for 30 min at 20C. DyLight488 conjugated goat anti-rabbit IgG (BA1127, 5-10 µg/1x106 cells) was used as secondary antibody for 30 minutes at 20C. Isotype control antibody (Green line) was rabbit IgG (1 µg/1x106) used under the same conditions. Unlabelled sample without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.
Western blot analysis of KMT6/EZH2 using anti-KMT6/EZH2 antibody (A00050-1). Electrophoresis was performed on a 5-20% SDS-PAGE gel at 70V (Stacking gel) / 90V (Resolving gel) for 2-3 hours. The sample well of each lane was loaded with 30 ug of sample under reducing conditions. Lane 1: human Jurkat whole cell lysates,Lane 2: human K562 whole cell lysates,Lane 3: human MCF-7 whole cell lysates,Lane 4: rat spleen tissue lysates,Lane 5: rat testis tissue lysates,Lane 6: mouse spleen tissue lysates,Lane 7: mouse testis tissue lysates.After electrophoresis, proteins were transferred to a nitrocellulose membrane at 150 mA for 50-90 minutes. Blocked the membrane with 5% non-fat milk/TBS for 1.5 hour at RT. The membrane was incubated with rabbit anti-KMT6/EZH2 antigen affinity purified polyclonal antibody (Catalog A00050-1) at 0.5 µg/mL overnight at 4C, then washed with TBS-0.1%Tween 3 times with 5 minutes each and probed with a goat anti-rabbit IgG-HRP secondary antibody at a dilution of 1:5000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog EK1002) with Tanon 5200 system. A specific band was detected for KMT6/EZH2 at approximately 98 kDa. The expected band size for KMT6/EZH2 is at 85 kDa.
Immunoprecipitating KMT6/EZH2 in MCF-7 whole cell lysate.Western blot analysis of KMT6/EZH2 using anti-KMT6/EZH2 antibody (A00050-1).Lane 1: MCF-7 whole cell lysates (30ug),Lane 2: Rabbit control IgG instead of anti-KMT6/EZH2 antibody in MCF-7 whole cell lysate,Lane 3: anti-KMT6/EZH2 antibody (2µg) + MCF-7 whole cell lysate (500µg).After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-KMT6/EZH2 antigen affinity purified polyclonal antibody (A00050-1) at a dilution of 0.5 µg/mL and probed with a goat anti-rabbit IgG-HRP secondary antibody (Catalog BA1054). The signal is developed using ECL Plus Western Blotting Substrate (Catalog AR1196-200). A specific band was detected for KMT6/EZH2 at approximately 98 kDa. The expected band size for KMT6/EZH2 is at 85 kDa.
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