Recombinant
RabMAb

Recombinant Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)

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Overview

  • Product name

    Anti-LAG-3 antibody [EPR20261] - BSA and Azide free
    See all LAG-3 primary antibodies

  • Description

    Rabbit monoclonal [EPR20261] to LAG-3 - BSA and Azide free

  • Host species

    Rabbit

  • Tested applications

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

  • Species reactivity

    Reacts with: Human

  • Immunogen

    Recombinant fragment aa 1-300. The exact sequence is proprietary.
    Database link: P18627

  • Positive control

    • WB: Human LAG-3 Fc chimera recombinant protein (aa23-450). IHC-P: Human tonsil and Hodgkin’s lymphoma tissues. ICC/IF: HEK-293T cells transfected with a GFP-tagged LAG3 expression construct. Flow Cyt: HEK-293T transfected with a GFP-tagged human LAG-3 construct. IP: HEK-293T transfected with a GFP-tagged human LAG-3 construct whole cell lysate.

  • General notes

    ab227579 is the carrier-free version of ab209236 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 <1 minute hands-on-time and 100% antibody recovery: available for fluorescent dyes, HRP, biotin and gold.

    Ab227579 is compatible with the Maxpar® Antibody Labeling Kit from Fluidigm.

    Maxpar® is a trademark of Fluidigm Canada Inc.

    This product was previously labelled as Lymphocyte Activation Gene 3.

Properties

Applications

Our Abpromise guarantee covers the use of ab227579 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
IHC-P Use at an assay dependent concentration. Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

Different mRNA expression levels of LAG3 in brain have been reported in the literature (PMID: 1692078; PMID: 12825348). In IHC, under our experimental conditions, this antibody showed no positive staining on human cerebral cortex.
WB Use at an assay dependent concentration. Detects a band of approximately 90 kDa (predicted molecular weight: 57 kDa).
ICC/IF Use at an assay dependent concentration.
IP Use at an assay dependent concentration.
Flow Cyt Use at an assay dependent concentration.

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

    • Flow Cytometry - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)
      Flow Cytometry - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)

      Flow cytometric analysis of Human peripheral blood mononuclear cells treated with 1 μg/mL PHA for 3 days cells with ab209236 at 1/50 dilution (right) compared with a rabbit monoclonal IgG isotype control (ab172730; left). ab150077 at 1/2000 dilution was used as the secondary antibody.

      Only the CD3+ population are also positive for LAG-3. Gated on total viable cells.

       

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab209236).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)

      Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling LAG-3 with ab209236 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Sporadic cytoplasmic staining on immunocytes of human tonsil [PMID: 11527700; PMID: 16757686].

      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.

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab209236).

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    • Immunocytochemistry/ Immunofluorescence - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)
      Immunocytochemistry/ Immunofluorescence - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)

      Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized HEK-293T (Human epithelial cell line from embryonic kidney) cells transfected with GFP-tagged LAG3 expression construct or GFP only, labeling LAG-3 with ab209236 at 1/100 dilution, followed by Goat anti-rabbit IgG (Alexa Fluor® 647) (ab150079) secondary antibody at 1/1000 dilution (green).

      Confocal image showing positive staining on HEK-293T cells transfected with a GFP-tagged LAG-3 expression construct.

      The nuclear counter stain is DAPI (blue). Tubulin is detected with ab195889 (Anti-alpha Tubulin antibody [DM1A] - Microtubule Marker (Alexa Fluor® 594)) at 1/200 dilution (red).

      Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat anti-rabbit IgG (Alexa Fluor® 647) (ab150079) 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 (ab209236).

    • Flow Cytometry - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)
      Flow Cytometry - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)

      Flow cytometric analysis of HEK-293T (Human epithelial cell line from embryonic kidney) cells transfected with a GFP-tagged human LAG3 construct labeling LAG-3 with ab209236 at 1/500 dilution (right) compared with a rabbit monoclonal IgG isotype control (ab172730; left). Goat anti rabbit IgG (Alexa Fluor® 647) ab150079 at 1/2000 dilution was used as the secondary antibody.

      Note: Fresh cells without fixation and permeabilization were used to perform FC testing. Only GFP positive population results in LAG3 positive staining (Q2, right panel).

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab209236).

    • Immunoprecipitation - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)
      Immunoprecipitation - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)

      LAG-3 was immunoprecipitated from 0.35 mg of HEK-293T (Human epithelial cell line from embryonic kidney) transfected with a GFP-tagged human LAG3 construct whole cell lysate with ab209236 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab209236 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/10000 dilution.

      Lane 1: HEK-293T transfected with a GFP-tagged human LAG3 construct whole cell lysate 10 μg (Input).

      Lane 2: ab209236 IP in HEK-293T transfected with a GFP-tagged human LAG3 construct whole cell lysate.

      Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab209236 in HEK-293T transfected with a GFP-tagged human LAG3 construct whole cell lysate.

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

      Exposure time: 1 second.

       

      This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (ab209236).

    • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)
      Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [EPR20261] - BSA and Azide free (ab227579)

      This IHC data was generated using the same anti-LAG-3 antibody clone, EPR20261, in a different buffer formulation (cat# ab209236).

      Immunohistochemical analysis of paraffin-embedded human tonsil Hodgkin’s lymphoma labeling LAG-3 with ab209236 at 1/1000 dilution, followed by Goat Anti-Rabbit IgG H&L (HRP) Ready to use. Cytoplasmic staining on immunocytes of the human Hodgkin’s lymphoma [PMID: 11527700; PMID: 16757686].

      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.

      Perform heat mediated antigen retrieval with Tris/EDTA buffer pH 9.0 before commencing with IHC staining protocol.

    References

    ab227579 has not yet been referenced specifically in any publications.

    Publishing research using ab227579? Please let us know so that we can cite the reference in this datasheet.

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