DIAPH1 Antibody

• Catalog Number: orb763118
• Category: Antibodies
• Description: DIAPH1 Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: ELISA, FC, ICC, IF, WB
• Reactivity: Human, Monkey
• Isotype: Rabbit IgG
• Immunogen: E.coli-derived human DIAPH1 recombinant protein (Position: D1039-Q1153).
• Concentration: Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Dilution range: Western blot, 0.25-0.5 μg/ml, Human, Monkey Immunocytochemistry/Immunofluorescence, 5 μg/ml, Human Flow Cytometry, 1-3 μg/1x106 cells, Human Direct ELISA, 0.1-0.5 μg/ml, Human
• Form/Appearance: Lyophilized
• Conjugation: Unconjugated
• MW: 150 kDa
• UniProt ID: O60610
• Storage: 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 freezing and thawing.
• 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. Adding 0.2 ml of distilled water will yield a concentration of 500 μg/ml.
• Expiration Date: 12 months from date of receipt.

Western blot analysis of DIAPH1 using anti-DIAPH1 antibody (orb763118). 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 K562 whole cell lysates, Lane 2: human Raji whole cell lysates, Lane 3: human HeLa whole cell lysates, Lane 4: monkey COS-7 whole cell lysates, Lane 5: human Caco-2 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-DIAPH1 antigen affinity purified polyclonal antibody (Catalog # orb763118) 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 (Catalog # orb90503) with Tanon 5200 system. A specific band was detected for DIAPH1 at approximately 150 kDa. The expected band size for DIAPH1 is at 150 kDa.

IF analysis of DIAPH1 using anti-DIAPH1 antibody (orb763118). DIAPH1 was detected in an immunocytochemical section of Caco-2 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 5 μg/mL rabbit anti-DIAPH1 Antibody (orb763118) overnight at 4°C. DyLight®488 Conjugated Goat Anti-Rabbit 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 K562 cells using anti-DIAPH1 antibody (orb763118). Overlay histogram showing K562 cells stained with orb763118 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-DIAPH1 Antibody (orb763118, 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.