CD44 Antibody (monoclonal, 7H7)

• Catalog Number: orb738399
• Category: Antibodies
• Description: CD44 Antibody (monoclonal, 7H7)
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: 7H7
• Tested applications: FC, ICC, IF, WB
• Reactivity: Human, Rat
• Isotype: Mouse IgG2b
• Immunogen: A synthetic peptide corresponding to a sequence at the N-terminus of human CD44 (29-56aa RFAGVFHVEKNGRYSISRTEAADLCKAF), different from the related mouse and rat sequences by two amino acids.
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Dilution range: Western blot, 0.25-0.5μg/ml, Human, Rat Immunocytochemistry/Immunofluorescence, 5μg/ml, Human Flow Cytometry, 1-3μg/1x106 cells, Human
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 82 kDa
• UniProt ID: P16070
• Storage: Store at -20˚C for one year from date of receipt. After reconstitution, at 4˚C for one month. It can also be aliquotted and stored frozen at -20˚C for six months. Avoid repeated freeze-thaw cycles.
• Note: For research use only
• Application notes: Tested Species: In-house tested species with positive results. Other applications have not been tested. Optimal dilutions should be determined by end users. Add 0.2ml of distilled water will yield a concentration of 500ug/ml.
• Expiration Date: 12 months from date of receipt.

Western blot analysis of CD44 using anti-CD44 antibody (orb738399). 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 50ug of sample under reducing conditions. Lane 1: human HeLa whole cell lysates, Lane 2: human A549 whole cell lysates, Lane 3: rat C6 whole cell lysates. After Electrophoresis, proteins were transferred to a Nitrocellulose membrane at 150mA for 50-90 minutes. Blocked the membrane with 5% Non-fat Milk/TBS for 1.5 hour at RT. The membrane was incubated with mouse anti-CD44 antigen affinity purified monoclonal antibody (Catalog # orb738399) 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-mouse IgG-HRP secondary antibody at a dilution of 1:10000 for 1.5 hour at RT. The signal is developed using an Enhanced Chemiluminescent detection (ECL) kit (Catalog # orb90502) with Tanon 5200 system. A specific band was detected for CD44 at approximately 82KD. The expected band size for CD44 is at 82KD.

IF analysis of CD44 using anti-CD44 antibody (orb738399). CD44 was detected in immunocytochemical section of A431 cells. Enzyme antigen retrieval was performed using IHC enzyme antigen retrieval reagent (orb90553) for 15 mins. The cells were blocked with 10% goat serum. And then incubated with 5μg/mL mouse anti-CD44 Antibody (orb738399) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Mouse IgG was used as secondary antibody at 1:100 dilution and incubated for 30 minutes at 37°C. The section was counterstained with DAPI. Visualize using a fluorescence microscope and filter sets appropriate for the label used.

Flow Cytometry analysis of U87 cells using anti-CD44 antibody (orb738399). Overlay histogram showing U87 cells stained with orb738399 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with mouse anti-CD44 Antibody (orb738399, 1μg/1x10^6 cells) for 30 min at 20°C. DyLight®488 conjugated goat anti-mouse IgG (5-10μg/1x10^6 cells) was used as secondary antibody for 30 minutes at 20°C. Isotype control antibody (Green line) was mouse IgG (1μg/1x10^6) used under the same conditions. Unlabelled sample (Red line) was also used as a control.