Astana Biomed Group, an authorized Abcam distributor in Central Asia

Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

Key features and details

  • Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
  • Rabbit monoclonal [EPR19759] to PD-L1 - BSA and Azide free
  • Suitable for: ICC, IHC-P, IP, WB, Flow Cyt
  • Knockout validated
  • Reacts with: Human

Overview

  • Product name

    Anti-PD-L1 antibody [EPR19759] - BSA and Azide free
    See all PD-L1 primary antibodies

  • Description

    Rabbit monoclonal [EPR19759] to PD-L1 - BSA and Azide free

  • Host species

    Rabbit

  • Tested Applications & Species

    Application Species
    Flow Cyt
    Chinese hamster
    IHC-P
    Human
    IP
    Human
    WB
    Human
    See all applications and species data

  • Immunogen

    This information is proprietary to Abcam and/or its suppliers.
    (Peptide available as ab150077)

  • Positive control

    • WB: Wild-type A549 treated with 100 ng/mL IFN gamma (ab259377) for 48 h cell lysate; Chinese hamster ovary cell lysate overexpressing PD-L1; NCI-H1975 whole cell lysate. IHC-P: Human tonsil, placenta and stomach cancer tissues. ICC/IF: CHO-PDL1 and NCI-H1975 cells. IP: NCI-H1975 whole cell lysate.

  • General notes

    Ab221612 is the carrier-free version of ab213524. 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

    ab221612 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-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    Anti-PD-L1 antibody [EPR19759] (ab213524)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) analysis of human tonsil tissue labelling PD-L1 with ab213524 at a dilution of 1:250. Heat mediated antigen retrieval was performed using AR9 antigen retrieval solution, and microwave treatment for 15 min at 20% power. Anti-Rabbit/Mouse HRP polymer (PerkinElmer Opal Polymer HRP Ms Plus Rb) was used as secondary antibody. Opal tyramide amplification was performed using Opal 520 fluorophore. Counterstained with DAPI stain. Image scanned with Vectra 3.0 and analyzed via Phenochart software.
    This image was courteously provided by Dr. Houssein Abdul Sater, Georgia Cancer Center.

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    This IHC data was generated using the same anti-PDL1 antibody clone, EPR19759, in a different buffer formulation (cat# ab213524).

    Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling PD-L1 with ab213524 at 1/250 dilution, followed by Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Membrane staining on the human tonsil crypt epithelium is observed.

    Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.

  • Western blot - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Western blot - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    All lanes : Anti-PD-L1 antibody [EPR19759] (ab213524) at 1/1000 dilution

    Lane 1 : Wild-type A549 treated with 100 ng/mL IFN gamma (ab259377) for 48 h cell lysate
    Lane 2 : CD274 knockout A549 treated with 100 ng/mL IFN gamma (ab259377) for 48 h cell lysate
    Lane 3 : U-87 MG cell lysate
    Lane 4 : MCF7 cell lysate
    Lane 5 : Wild-type A549 untreated cell lysate
    Lane 6 : CD274 knockout A549 untreated cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 33 kDa
    Observed band size: 50 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 (ab213524).

    Lanes 1 - 6: Merged signal (red and green). Green - ab213524 observed at 50 kDa. Red - loading control, ab8245 observed at 37 kDa.

    ab213524 Recombinant Anti-PD-L1 antibody [EPR19759] was shown to specifically react with PD-L1 in wild-type A549 treated with 100 ng/mL IFN gamma for 48 h cells in western blot. Loss of signal was observed when both treated and untreated knockout cell line ab267055 (treated and untreated knockout cell lysates ab256866) were used. Wild-type and PD-L1 knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab213524 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 and 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.

  • Flow Cytometry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Flow Cytometry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    Flow Cytometry analysis of CHO-PD-L1 (red) and CHO-S (blue) cells labelling PD-L1 with ab213524 at 1/500. Cells were fixed with 4% paraformaldehyde, permeabilized with 0.1% tween-20-PBS and blocked with 10% goat serum. An Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1/2000) was used as the secondary antibody.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213524).

  • Immunocytochemistry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunocytochemistry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    Clone EPR19759 (ab221612) has been successfully conjugated by Abcam. This image was generated using Anti-PD-L1 antibody [EPR19759] (Alexa Fluor® 647). Please refer to ab215251 for protocol details.

    ab215251 staining PDL1 in CHO-PDL1 cells. The lower panels demonstrate that ab215251 does not cross react with un-transfected CHO cells.

    The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab215251 at 1/200 dilution (shown in red) and ab195887, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 488), at 1/250 dilution (shown in green). Nuclear DNA was labelled with DAPI (shown in blue).

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • Immunocytochemistry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunocytochemistry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    Clone EPR19759 (ab221612) has been successfully conjugated by Abcam. This image was generated using Anti-PD-L1 antibody [EPR19759] (Alexa Fluor® 488). Please refer to ab214958 for protocol details.

    ab214958 staining PDL1 in CHO-PDL1 cells. The lower panels demonstrate that ab214958 does not cross react with un-transfected CHO cells.

    The cells were fixed with 4% formaldehyde (10 min), permeabilized with 0.1% Triton X-100 for 5 minutes and then blocked with 1% BSA/10% normal goat serum/0.3M glycine in 0.1% PBS-Tween for 1h. The cells were then incubated overnight at +4°C with ab214958 at 1/200 dilution (shown in green) and ab195889, Mouse monoclonal to alpha Tubulin (Alexa Fluor® 594), at 1/250 dilution (shown in red). Nuclear DNA was labelled with DAPI (shown in blue).

    Image was taken with a confocal microscope (Leica-Microsystems, TCS SP8).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    Immunohistochemical analysis of paraffin-embedded human placenta tissue labeling PD-L1 with ab213524 at 1/250 dilution, followed by Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Membrane staining on the human placenta is observed.

    Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is rabbit specific IHC polymer detection kit HRP/DAB (ab209101)

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213524).

    Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.

  • Immunocytochemistry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunocytochemistry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized CHO (Chinese hamster ovary cell line) cells labeling PD-L1 with ab213524 at 1/100 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

    Confocal image showing membrane and cytoplasmic staining on CHO-PDL1 cells.

    The nuclear counterstain is DAPI (blue).

    Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

    The negative controls are as follows:-

    -ve control 1: ab213524 at 1/100 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution.

    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) 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 (ab213524).

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    Immunohistochemical analysis of paraffin-embedded human stomach cancer tissue labeling PD-L1 with ab213524 at 1/250 dilution, followed by Rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    Membrane staining on the human stomach cancer is observed.

    Counter stained with Hematoxylin.

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is rabbit specific IHC polymer detection kit HRP/DAB (ab209101).

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213524).

    Heat mediated antigen retrieval was performed before commencing with IHC staining protocol.

  • Immunocytochemistry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunocytochemistry - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized NCI-H1975 (Human lung non small cell carcinoma cell line) cells labeling PD-L1 with ab213524 at 1/100 dilution, followed by Goat Anti-Rabbit IgG (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green).

    Confocal image showing weakly membrane and cytoplasmic staining on NCI-H1975 cells.

    The nuclear counterstain is DAPI (blue).

    Tubulin is detected with Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution, followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution (red).

    The negative controls are as follows:-

    -ve control 1: ab213524 at 1/100 dilution followed by Goat Anti-Mouse IgG H&L (Alexa Fluor® 594) (ab150120) secondary antibody at 1/1000 dilution.

    -ve control 2: Anti-alpha Tubulin mouse MAb (ab7291) at 1/1000 dilution followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) 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 (ab213524).

  • Western blot - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Western blot - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    All lanes : Anti-PD-L1 antibody [EPR19759] (ab213524) at 1/1000 dilution

    Lane 1 : Wild-type A549 treated with 100 ng/mL IFN gamma for 48 h cell lysate
    Lane 2 : CD274 knockout A549 treated with 100 ng/mL IFN gamma for 48 h cell lysate
    Lane 3 : U-87 MG cell lysate
    Lane 4 : MCF7 cell lysate
    Lane 5 : Wild-type A549 untreated cell lysate
    Lane 6 : CD274 knockout A549 untreated cell lysate

    Lysates/proteins at 20 µg per lane.

    Performed under reducing conditions.

    Predicted band size: 33 kDa
    Observed band size: 50 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 (ab213524).

    Lanes 1 - 6: Merged signal (red and green). Green - ab213524 observed at 50 kDa. Red - loading control, ab8245 observed at 37 kDa.

    ab213524 Recombinant Anti-PD-L1 antibody [EPR19759] was shown to specifically react with PD-L1 in wild-type A549 treated with 100 ng/mL IFN gamma for 48 h cells in western blot. Loss of signal was observed when both treated and untreated knockout cell line ab267054 (treated and untreated knockout cell lysates ab256831) were used. Wild-type and PD-L1 knockout samples were subjected to SDS-PAGE. Membrane was blocked for 1 hour at room temperature in 0.1% TBST with 3% non-fat dried milk. ab213524 and Anti-GAPDH antibody [6C5] - Loading Control (ab8245) were incubated overnight at 4°C at 1 in 1000 and 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-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Immunoprecipitation - Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

    PD-L1 was immunoprecipitated from 0.35 mg of NCI-H1975 (Human non-small cell lung cancer cell line) whole cell lysate with ab213524 at 1/30 dilution.

    Western blot was performed from the immunoprecipitate using ab213524 at 1/1000 dilution.

    VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

    Lane 1: NCI-H1975 whole cell lysate 10µg (Input).

    Lane 2: ab213524 IP in NCI-H1975 whole cell lysate.

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab213524 in NCI-H1975 whole cell lysate.

    Blocking and dilution buffer and concentration: 5% NFDM/TBST.

    Exposure time: 3 minutes.

    This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab213524).

  • Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)
    Anti-PD-L1 antibody [EPR19759] - BSA and Azide free (ab221612)

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