Anti-Rac1 Antibody

• Catalog Number: orb259608
• Category: Antibodies
• Description: Anti-Rac1 Antibody. Tested in IF, ICC, WB applications. This antibody reacts with Human, Mouse, Rat.
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: ICC, IF, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Rabbit IgG
• Immunogen: A synthetic peptide corresponding to a sequence at the C-terminus of human Rac1, identical to the related mouse and rat sequences.
• Antibody Type: Primary Antibody
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 21 kDa
• UniProt ID: P63000
• Storage: Maintain refrigerated at 2-8°C for up to 2 weeks. For long term storage store at -20°C in small aliquots to prevent freeze-thaw cycles.
• Alternative names: Ras-related C3 botulinum toxin substrate 1; Cell m
• Note: For research use only
• Application notes: Western blot, 0.1-0.5μg/ml, Human, Mouse, Rat Immunohistochemistry, 2-5μg/ml, Human Immunocytochemistry/Immunofluorescence, 5μg/ml, Human Flow Cytometry(Fixed), 1-3 μg/1x106 cells, Human. Add 0.2ml of distilled water will yield a concentration of 500ug/ml
• Expiration Date: 12 months from date of receipt.

Flow Cytometry analysis of U87 cells using anti-Rac1 antibody. Overlay histogram showing U87 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-Rac1 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 without incubation with primary antibody and secondary antibody (Red line) was used as a blank control.

IF analysis of Rac1 using anti-Rac1 antibody and anti-Beta Tubulin antibody. Rac1 was detected in immunocytochemical section of A549 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-Rac1 Antibody and mouse anti-Beta Tubulin antibody overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit IgG and DyLight®594 Conjugated Goat Anti-Mouse IgG were used as secondary antibody at 1:500 dilution and incubated for 30 minutes at 37°C. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

IHC analysis of Rac1 using anti-Rac1 antibody. Rac1 was detected in a paraffin-embedded section of human colon cancer tissue. Heat mediated antigen retrieval was performed in EDTA buffer (pH8.0, epitope retrieval solution). The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 2 µg/ml rabbit anti-Rac1 Antibody overnight at 4°C. Peroxidase Conjugated Goat Anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using HRP Conjugated Rabbit IgG Super Vision Assay Kit with DAB as the chromogen.

Western blot analysis of Rac1 using anti-Rac1 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 A431 whole cell lysates, Lane 2: human Hacat whole cell lysates, Lane 3: human MCF-7 whole cell lysates, Lane 4: human PC-3 whole cell lysates, Lane 5: rat small intestine tissue lysates, Lane 6: mouse small intestine 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-Rac1 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 Rac1 at approximately 21 kDa. The expected band size for Rac1 is at 21 kDa.