Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-AIF antibody [E20] - BSA and Azide free (ab220215)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [E20] to AIF - BSA and Azide free
  • Suitable for: ICC/IF, IP, WB, IHC-P, Flow Cyt
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-AIF antibody [E20] - BSA and Azide free
    See all AIF primary antibodies

  • Description

    Rabbit monoclonal [E20] to AIF - BSA and Azide free

  • Host species

    Rabbit

  • Tested applications

    Suitable for: ICC/IF, IP, WB, IHC-P, Flow Cytmore details

  • Species reactivity

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

  • Immunogen

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

  • Positive control

    • WB: HEK-293T and K562 cell lysate. IHC-P: Human cervical carcinoma tissue.

  • General notes

    ab220215 is the carrier-free version of ab32516. 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

    ab220215 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.

    The Life Science industry has been in the grips of a reproducibility crisis for a number of years. Abcam is leading the way in addressing the problem with our range of recombinant monoclonal antibodies and knockout edited cell lines for gold-standard validation.

    One factor contributing to the crisis is the use of antibodies that are not suitable. This can lead to misleading results and the use of incorrect data informing project assumptions and direction. To help address this challenge, we have introduced an application and species grid on our primary antibody datasheets to make it easy to simplify identification of the right antibody for your needs.

    Learn more here.

Properties

Images

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)

    Ab32516, at a 1/500 dilution, staining AIF in paraffin embedded human cervical carcinoma tissue by Immunohistochemistry.

    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 (ab32516).

  • Immunocytochemistry/ Immunofluorescence - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)
    Immunocytochemistry/ Immunofluorescence - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)

    ICC/IF image of ab32516 stained HeLa cells. The cells were 4% PFA fixed (10 min) and then incubated in 1%BSA / 10% normal goat serum / 0.3M glycine in 0.1% PBS-Tween for 1h to permeabilise the cells and block non-specific protein-protein interactions. The cells were then incubated with the antibody (ab32516, 1µg/ml) overnight at +4°C. The secondary antibody (green) was Alexa Fluor® 488 goat anti-rabbit IgG (H+L) used at a 1/1000 dilution for 1h. Alexa Fluor® 594 WGA was used to label plasma membranes (red) at a 1/200 dilution for 1h. DAPI was used to stain the cell nuclei (blue) at a concentration of 1.43µM.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32516).

  • Western blot - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)
    Western blot - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)
    All lanes : Anti-AIF antibody [E20] - Mitochondrial Marker (ab32516) at 1/1000 dilution

    Lane 1 : Wild-type HEK-293T cell lysate
    Lane 2 : AIFM1 knockout HEK-293T cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 67 kDa
    Observed band size: 67 kDa



    This data was developed using the same antibody clone in a different buffer formulation (ab32516).

    Lanes 1- 2: Merged signal (red and green). Green - ab32516 observed at 67 kDa. Red - Anti-GAPDH antibody [6C5] - Loading Control (ab8245) observed at 37 kDa.

     ab32516 was shown to react with AIF in wild-type HEK-293T cells in western blot. Loss of signal was observed when knockout cell line ab266347 (knockout cell lysate ab256834) was used. Wild-type HEK-293T and AIFM1 knockout HEK-293T 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. ab32516 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) overnight at 4°C at a 1 in 1000 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.

  • Immunoprecipitation - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)
    Immunoprecipitation - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)
    AIF was immunoprecipitated using 0.5mg K562 whole cell extract, 5µg of Rabbit monoclonal to AIF and 50µl of protein G magnetic beads (+). No antibody was added to the control (-).
    The antibody was incubated under agitation with Protein G beads for 10min, K562 whole cell extract lysate diluted in RIPA buffer was added to each sample and incubated for a further 10min under agitation.
    Proteins were eluted by addition of 40µl SDS loading buffer and incubated for 10min at 70oC; 10µl of each sample was separated on a SDS PAGE gel, transferred to a nitrocellulose membrane, blocked with 5% BSA and probed with ab32516.
    Secondary: Mouse monoclonal [SB62a] Secondary Antibody to Rabbit IgG light chain (HRP) (ab99697).
    Band: 67kDa: AIF

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab32516).

  • Flow Cytometry - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)
    Flow Cytometry - Anti-AIF antibody [E20] - BSA and Azide free (ab220215)

    Overlay histogram showing K562 cells stained with ab32516 (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 (ab32516, 1/50 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 positive signal in K562 cells fixed with methanol (5 min)/permeabilized with 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 (ab32516).

  • Anti-AIF antibody [E20] - BSA and Azide free (ab220215)
    Anti-AIF antibody [E20] - BSA and Azide free (ab220215)

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