SGT1/ECD Antibody

• Catalog Number: orb865692
• Category: Antibodies
• Description: SGT1/ECD Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: ELISA, FC, ICC, IF, WB
• Reactivity: Human
• Isotype: Rabbit IgG
• Immunogen: E.coli-derived human SGT1/ECD recombinant protein (Position: L50-Q567).
• 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 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: 80 kDa
• UniProt ID: O95905
• 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 SGT1/ECD using anti-SGT1/ECD antibody (orb865692). 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 PC-3 whole cell lysates, Lane 2: human Caco-2 whole cell lysates, Lane 3: human HL-60 whole cell lysates, Lane 4: human U20S 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-SGT1/ECD antigen affinity purified polyclonal antibody (Catalog # orb865692) 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 SGT1/ECD at approximately 80 kDa. The expected band size for SGT1/ECD is at 80 kDa.

IF analysis of SGT1/ECD using anti-SGT1/ECD antibody (orb865692). SGT1/ECD was detected in an 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 5 μg/mL rabbit anti-SGT1/ECD Antibody (orb865692) 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 HeLa cells using anti-SGT1/ECD antibody (orb865692). Overlay histogram showing HeLa cells stained with orb865692 (Blue line). The cells were blocked with 10% normal goat serum. And then incubated with rabbit anti-SGT1/ECD Antibody (orb865692, 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.