Beta Tubulin Antibody(monoclonal, 5E4)

• Catalog Number: orb623835
• Category: Antibodies
• Description: Beta Tubulin Antibody(monoclonal, 5E4)
• Species/Host: Mouse
• Clonality: Monoclonal
• Clone Number: 5E4
• Tested applications: FC, ICC, IF, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Mouse IgG1
• Immunogen: A synthetic peptide corresponding to a sequence at the C-terminus of human Beta Tubulin (383-412aa EQFTAMFRRKAFLHWYTGEGMDEMEFTEAE), identical to the related mouse and rat sequences.
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Dilution range: Western blot, 0.1-0.5μg/ml, Human, Mouse, Rat Immunocytochemistry/Immunofluorescence, 2μg/ml, Human Flow Cytometry, 1-3μg/1x106 cells, Human
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 55 kDa
• UniProt ID: P07437
• Sensitivity: > 5000 cells
• 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 500μg/ml.
• Expiration Date: 12 months from date of receipt.

Western blot analysis of Beta Tubulin using anti-Beta Tubulin antibody (orb623835). 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 HEK293 whole cell lysates, Lane 2: human HeLa whole cell lysates, Lane 3: human HEPG2 whole cell lysates, Lane 4: human HL-60 whole cell lysates, Lane 5: human Raji whole cell lysates, Lane 6: rat brain tissue lysates, Lane 7: mouse brain tissue 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-Beta Tubulin antigen affinity purified monoclonal antibody (Catalog # orb623835) 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 Beta Tubulin at approximately 55KD. The expected band size for Beta Tubulin is at 55KD.

IF analysis of Beta Tubulin using anti-Beta Tubulin antibody (orb623835). Beta Tubulin 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 2μg/mL mouse anti-Beta Tubulin Antibody (orb623835) 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 SiHa cells using anti-Beta Tubulin antibody (orb623835). Overlay histogram showing SiHa cells stained with orb623835 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with mouse anti-Beta Tubulin Antibody (orb623835, 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.