Recombinant

Recombinant Anti-LAG-3 antibody [CAL26] (ab237719)

Overview

  • Product name

    Anti-LAG-3 antibody [CAL26]
    See all LAG-3 primary antibodies
  • Description

    Rabbit monoclonal [CAL26] to LAG-3
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, Flow Cyt, IP, IHC-Pmore details
    Unsuitable for: ICC/IF
  • Species reactivity

    Reacts with: Human
  • Immunogen

    Synthetic peptide. This information is considered to be commercially sensitive.

  • Positive control

    • WB: LAG-3-GFP transfected HEK-293 whole cell lysate; human PBMC whole cell lysate. IHC-P: Human tonsil tissue. Flow cyt: Human PBMCs treated with 10µg/ml PHA for 48 hours. IP: LAG-3-GFP transfected HEK-203& whole cell lysate.
  • General notes

    This product was previously labelled as Lymphocyte Activation Gene 3.

Properties

Applications

Our Abpromise guarantee covers the use of ab237719 in the following tested applications.

The application notes include recommended starting dilutions; optimal dilutions/concentrations should be determined by the end user.

Application Abreviews Notes
WB 1/1000. Detects a band of approximately 16, 70 kDa (predicted molecular weight: 57 kDa).
Flow Cyt 1/250.
IP 1/20.
IHC-P Use a concentration of 0.3 µg/ml. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.
  • Application notes
    Is unsuitable for ICC/IF.
  • Target

    • Function

      Involved in lymphocyte activation. Binds to HLA class-II antigens.
    • Tissue specificity

      On cell surface of activated NK and T-lymphocytes.
    • Sequence similarities

      Contains 3 Ig-like C2-type (immunoglobulin-like) domains.
      Contains 1 Ig-like V-type (immunoglobulin-like) domain.
    • Cellular localization

      Membrane.
    • Information by UniProt
    • Database links

    • Alternative names

      • CD223 antibody
      • CD223 antigen antibody
      • FDC protein antibody
      • LAG-3 antibody
      • Lag3 antibody
      • LAG3_HUMAN antibody
      • Lymphocyte activating 3 antibody
      • Lymphocyte activation gene 3 antibody
      • Lymphocyte activation gene 3 protein antibody
      • Protein FDC antibody
      see all

    Images

    • Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling LAG-3 with ab237719 at 1/400 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) ready to use. Positive staining on the human tonsil is observed. Counter stained with hematoxylin.

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (HRP) ready to use.

      Heat mediated antigen retrieval using ab93684 (Tris/EDTA buffer, pH 9.0).

    • LAG-3 was immunoprecipitated from 0.35 mg of LAG-3-GFP transfected HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen)  whole cell lysate with ab237719 at 1/20 dilution. Western blot was performed from the immunoprecipitate using ab237719 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used as secondary antibody at 1/5000 dilution.

      Lane 1: LAG-3-GFP transfected HEK-293T whole cell lysate 10 μg (Input).
      Lane 2: ab237719 IP in LAG-3-GFP transfected HEK-293T whole cell lysate.
      Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab237719 in LAG-3-GFP transfected HEK-293T whole cell lysate.

      Blocking and dilution buffer and concentration: 5% NFDM/TBST.
      Exposure time: 30 seconds.

    • Flow cytometric analysis of 2% paraformaldehyde-fixed, 0.1% Tween-20 permeabilized human peripheral blood mononuclear cells ((PBMC) (treated with 10μg/ml PHA for 48 hours) cell line labeling LAG-3 with ab237719 at 1/250 dilution (Right) compared with Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (ab172730) (Left).

      Goat Anti-Rabbit IgG Fc (Alexa Fluor® 488) preadsorbed (ab150097), at 1/5000 dilution was used as the secondary antibody.

      Cells were surface stained with anti-CD3 conjugated to Alexa Fluor® 647. Then fixed with 2% PFA for 10min followed by intracellular staining rabbit IgG (Left) or ab237719 (Right).

    • All lanes : Anti-LAG-3 antibody [CAL26] (ab237719) at 1/1000 dilution

      Lane 1 : Untransfected HEK-293T (human epithelial cell line from embryonic kidney transformed with large T antigen) whole cell lysate
      Lane 2 : LAG-3-GFP overexpression HEK-293T whole cell lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 57 kDa
      Observed band size: 16,70 kDa why is the actual band size different from the predicted?


      Exposure time: 3 minutes


      Blocking and dilution buffer: 5% NFDM/TBST.

      This antibody also detects a 16kDa cleaved fragment. (PMID: 15557174).

    • All lanes : Anti-LAG-3 antibody [CAL26] (ab237719) at 1/1000 dilution

      Lane 1 : Human peripheral blood mononuclear cell (PBMC), whole cell lysate
      Lane 2 : Human peripheral blood mononuclear cell (PBMC) treated with 10µg/ml PHA for 48 hours, whole cell lysate
      Lane 3 : Human lymphoma lysate

      Lysates/proteins at 20 µg per lane.

      Secondary
      All lanes : Goat Anti-Rabbit IgG H&L (HRP) (ab97051) at 1/100000 dilution

      Predicted band size: 57 kDa
      Observed band size: 16,70 kDa why is the actual band size different from the predicted?


      Exposure time: 3 minutes


      Blocking and dilution buffer: 5% NFDM/TBST.

      This antibody also detects a 16kDa cleaved fragment. (PMID: 15557174).

       

    • Formalin-fixed, paraffin-embedded human tonsil tissue stained for LAG-3 using ab237719 at 0.3 μg/ml in immunohistochemical analysis.

    References

    ab237719 has not yet been referenced specifically in any publications.

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