Anti-HSPB8/Hsp22 Antibody

• Catalog Number: orb18992
• Category: Antibodies
• Description: Anti-HSPB8/Hsp22 Antibody
• Species/Host: Rabbit
• Clonality: Polyclonal
• Tested applications: FC, ICC, IF, IHC, IP, WB
• Reactivity: Human, Mouse, Rat
• Isotype: Rabbit IgG
• Immunogen: E.coli-derived human HSPB8/Hsp22 recombinant protein (Position: M1-T196). Human HSPB8/Hsp22 shares 94.4% and 95.4% amino acid (aa) sequence identity with mouse and rat HSPB8/Hsp22, respectively.
• 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: 22 kDa
• UniProt ID: Q9UJY1
• 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: Heat shock protein beta-8; HspB8; Alpha-crystallin
• Note: For research use only
• Application notes: Western blot, 0.1-0.5μg/ml, Human, Mouse, Rat Immunohistochemistry (Paraffin-embedded Section), 0.5-1μg/ml, Human, Mouse, Rat Immunocytochemistry/Immunofluorescence, 2μg/ml, Human Immunoprecipitation, 0.5-2 μg/ml, Human Flow Cytometry (Fixed), 1-3μg/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 U2OS cells using anti-HSPB8/Hsp22 antibody. Overlay histogram showing U2OS 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-HSPB8/Hsp22 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 HSPB8/Hsp22 using anti-HSPB8/Hsp22 antibody. HSPB8/Hsp22 was detected in immunocytochemical section of MCF7 cells. 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 2 µg/mL rabbit anti-HSPB8/Hsp22 Antibody overnight at 4°C. DyLight®594 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.

IHC analysis of HSPB8/Hsp22 using anti-HSPB8/Hsp22 antibody. HSPB8/Hsp22 was detected in paraffin-embedded section of human lung cancer tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 µg/ml rabbit anti-HSPB8/Hsp22 Antibody overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.

IHC analysis of HSPB8/Hsp22 using anti-HSPB8/Hsp22 antibody. HSPB8/Hsp22 was detected in paraffin-embedded section of human mammary cancer tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 µg/ml rabbit anti-HSPB8/Hsp22 Antibody overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.

IHC analysis of HSPB8/Hsp22 using anti-HSPB8/Hsp22 antibody. HSPB8/Hsp22 was detected in paraffin-embedded section of mouse pancreas tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 µg/ml rabbit anti-HSPB8/Hsp22 Antibody overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.

IHC analysis of HSPB8/Hsp22 using anti-HSPB8/Hsp22 antibody. HSPB8/Hsp22 was detected in paraffin-embedded section of rat pancreas tissues. Heat mediated antigen retrieval was performed in citrate buffer (pH6, epitope retrieval solution) for 20 mins. The tissue section was blocked with 10% goat serum. The tissue section was then incubated with 1 µg/ml rabbit anti-HSPB8/Hsp22 Antibody overnight at 4°C. Biotinylated goat anti-rabbit IgG was used as secondary antibody and incubated for 30 minutes at 37°C. The tissue section was developed using Strepavidin-Biotin-Complex (SABC) with DAB as the chromogen.

Immunoprecipitating HSPB8/Hsp22 in MCF-7 whole cell lysate. Western blot analysis of HSPB8/Hsp22 using anti-HSPB8/Hsp22 antibody; Lane 1: MCF-7 whole cell lysates (30 ug); Lane 2: Rabbit control IgG instead of anti-HSPB8/Hsp22 antibody in MCF-7 whole cell lysate; Lane 3: anti-HSPB8/Hsp22 antibody (2 µg) + MCF-7 whole cell lysate (500 µg). After electrophoresis, proteins were transferred to a membrane. Then the membrane was incubated with rabbit anti-HSPB8/Hsp22 antigen affinity purified polyclonal antibody at a dilution of 0.5 µg/mL and probed with a goat anti-rabbit IgG-HRP secondary antibody. The signal is developed using ECL Plus Western Blotting Substrate. A specific band was detected for HSPB8/Hsp22 at approximately 22 kDa. The expected band size for HSPB8/Hsp22 is at 22 kDa.

Western blot analysis of HSPB8/Hsp22 using anti-HSPB8/Hsp22 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 MCF-7 whole cell lysates, Lane 2: human U87 whole cell lysates, Lane 3: rat H9C2 whole cell lysates, Lane 4: rat heart tissue lysates, Lane 5: mouse C2C12 whole cell lysates, Lane 6: mouse skeletal muscle tissue lysates, Lane 7: mouse heart 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-HSPB8/Hsp22 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 HSPB8/Hsp22 at approximately 22 kDa. The expected band size for HSPB8/Hsp22 is at 22 kDa.