Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [Y208] to Bad - BSA and Azide free
  • Suitable for: Flow Cyt, ICC/IF, WB, IHC-P
  • Knockout validated
  • Reacts with: Mouse, Rat, Human

Overview

  • Product name

    Anti-Bad antibody [Y208] - BSA and Azide free
    See all Bad primary antibodies

  • Description

    Rabbit monoclonal [Y208] to Bad - BSA and Azide free

  • Host species

    Rabbit

  • Tested applications

    Suitable for: Flow Cyt, ICC/IF, WB, IHC-Pmore details

  • Species reactivity

    Reacts with: Mouse, Rat, Human
    Predicted to work with: Dog

  • Immunogen

    This information is proprietary to Abcam and/or its suppliers.

  • Positive control

    • WB: HAP1, MCF7 and HeLa cell lysate. IHC-P: Human lymphoma. ICC/IF: RAW 264.7. Flow Cyt: MCF7.

  • General notes

    Ab220116 is the carrier-free version of ab32445. This format is designed for use in antibody labeling, including fluorochromes, metal isotopes, oligonucleotides, enzymes.

    Our carrier-free formats are supplied in a buffer free of BSA, sodium azide and glycerol for higher conjugation efficiency.

    Use our conjugation kits for antibody conjugates that are ready-to-use in as little as 20 minutes with

    ab220116 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    This product is a recombinant monoclonal antibody, which offers several advantages including:

    • - High batch-to-batch consistency and reproducibility
    • - Improved sensitivity and specificity
    • - Long-term security of supply
    • - Animal-free production
    For more information see here.

    Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.

Properties

Images

  • Western blot - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    Western blot - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    All lanes : Anti-Bad antibody [Y208] (ab32445) at 1/2000 dilution

    Lane 1 : Wild-type HeLa cell lysate
    Lane 2 : BAD knockout HeLa cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 18 kDa
    Observed band size: 23 kDa
    why is the actual band size different from the predicted?



    This data was developed using the same antibody clone in a different buffer formulation (ab32445).

      Lanes 1- 2: Merged signal (red and green). Green - ab32445 observed at 23 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

     ab32445 was shown to react with Bad in wild-type HeLa cells in western blot. Loss of signal was observed when knockout cell line ab264843 (knockout cell lysate ab256847) was used. Wild-type HeLa and BAD knockout HeLa cell lysates were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab32445 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 2000 dilution and a 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye®800CW) preadsorbed (ab216773) and Goat anti-Mouse IgG H&L (IRDye®680RD) preadsorbed (ab216776) secondary antibodies at 1 in 20000 dilution for 1 hour at room temperature before imaging.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)

    Immunohistochemical staining of paraffin-embedded human lymphoma using ab32445 at 1/100 dilution.

    Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32445).

  • Western blot - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    Western blot - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    All lanes : Anti-Bad antibody [Y208] (ab32445) at 1/2000 dilution

    Lane 1 : Wild-type HAP1 whole cell lysate
    Lane 2 : BAD knockout HAP1 whole cell lysate
    Lane 3 : HeLa whole cell lysate
    Lane 4 : MCF7 whole cell lysate

    Lysates/proteins at 20 µg per lane.

    Predicted band size: 18 kDa



    This data was developed using the same antibody clone in a different buffer formulation (ab32445).

    Lanes 1 - 4: Merged signal (red and green). Green - ab32445 observed at 23 kDa. Red - loading control, ab9484, observed at 37 kDa.

    ab32445 was shown to specifically recognise BAD in wild-type HAP1 cells along with additional cross reactive bands. No band was observed when BAD knockout cells were examined. Wild-type and BAD knockout samples were subjected to SDS-PAGE. Ab32445 and ab9484 (Mouse anti-GAPDH loading control) were incubated overnight at 4°C at 1/2000 dilution and 1/20,000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed ab216773 and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed ab216776 secondary antibodies at 1/20,000 dilution for 1 hour at room temperature before imaging.

  • Immunocytochemistry/ Immunofluorescence - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    Immunocytochemistry/ Immunofluorescence - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)

    Immunocytochemical analysis of 2% paraformaldehyde-fixed, 0.1% Triton-X100 permeabilized RAW 264.7 labeling Bad with ab32445 at 1/250 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 568) secondary antibody at 1/1000 dilution.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32445).

  • Flow Cytometry - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    Flow Cytometry - Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    Overlay histogram showing MCF-7 cells stained with ab32445 (red line). The cells were fixed with 4% paraformaldehyde (10 min) and then permeabilized with 0.1% PBS-Tween for 20 min. The cells were then incubated in 1x PBS / 10% normal goat serum / 0.3M glycine to block non-specific protein-protein interactions followed by the antibody (ab32445, 1/20 dilution) for 30 min at 22°C. The secondary antibody used was DyLight® 488 goat anti-rabbit IgG (H+L) (ab96899) at 1/500 dilution for 30 min at 22°C. Isotype control antibody (black line) was rabbit monoclonal IgG (1µg/1x106 cells) used under the same conditions. Acquisition of >5,000 events was performed. This antibody gave a decreased signal in MCF-7 cells fixed with methanol (5 min)/permeabilized in 0.1% PBS-Tween used under the same conditions.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32445).

  • Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)
    Anti-Bad antibody [Y208] - BSA and Azide free (ab220116)

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