PERM1 Antibody

Flow Cytometry analysis of MCF-7 cells using anti-PERM1 antibody. Overlay histogram showing MCF-7 cells (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-PERM1 Antibody (1 µg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-rabbit IgG (5-10 µg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was rabbit IgG (1 µg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.

IF analysis of PERM1 using anti-PERM1 antibody and anti-Beta Tubulin antibody. PERM1 was detected in immunocytochemical section of U2OS cell. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5 µg/mL rabbit anti-PERM1 Antibody and mouse anti-Beta Tubulin antibody overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG (B) and DyLight®550 Conjugated Goat Anti-Mouse IgG (D) were used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI(C). Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Western blot analysis of PERM1 using anti-PERM1 antibody. 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 MCF-7 whole cell lysates, Lane 2: human Caco-2 whole cell lysates, Lane 3: human 293T whole cell 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-PERM1 antigen affinity purified polyclonal antibody at 0.5 µg/mL overnight at 4°C, 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 with Tanon 5200 system. A specific band was detected for PERM1 at approximately 75 kDa. The expected band size for PERM1 is at 85 kDa.