Recombinant
RabMAb

Recombinant Anti-LAG-3 antibody [EPR20294-77] (ab209238)

Overview

  • Product name

    Anti-LAG-3 antibody [EPR20294-77]
    See all LAG-3 primary antibodies
  • Description

    Rabbit monoclonal [EPR20294-77] to LAG-3
  • Host species

    Rabbit
  • Tested applications

    Suitable for: WB, ICC/IF, IPmore details
  • Species reactivity

    Reacts with: Mouse
  • Immunogen

    Recombinant fragment within Mouse LAG-3 aa 1-250. The exact sequence is proprietary.
    Database link: Q61790

  • Positive control

    • WB: Mouse LAG-3 recombinant protein (aa24-442) tagged with murine IgG2A at the C-terminus; mouse splenocytes treated with 5 ug/ml concanavalin A for 72 hours, whole cell lysate. ICC/IF: Mouse splenocytes. IP: Mouse splenocytes treated with 5 µg/ml concanavalin A for 72 hours, whole cell lysate.
  • General notes

    This product was previously labelled as Lymphocyte Activation Gene 3.

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

    This product is a recombinant rabbit monoclonal antibody.

Properties

Applications

Our Abpromise guarantee covers the use of ab209238 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 54, 70 kDa (predicted molecular weight: 57 kDa).
ICC/IF 1/100.
IP 1/30.

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

  • Immunofluorescent analysis of 4% paraformaldehyde-fixed, 0.1% Triton X-100 permeabilized untreated and concanavalin A (2.5 μg/ml for 72 hours) treated mouse splenocytes labeling LAG-3 with ab209238 at 1/100 dilution, followed by Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) secondary antibody at 1/1000 dilution (green). Confocal image showing increased membranous staining following treatment of mouse splenocytes with concanavalin A (2.5 μg/ml for 72 hours) (PMID 20391435; PMID 26318293; PMID 11527700; PMID 19631993).

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

    Secondary antibody only control: Used PBS instead of primary antibody, secondary antibody is Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) (ab150077) at 1/1000 dilution

  • Anti-LAG-3 antibody [EPR20294-77] (ab209238) at 1/10000 dilution + Mouse LAG-3 recombinant protein (aa24-442) tagged with murine IgG2A at the C-terminus, 10 ng

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

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 57 kDa
    Observed band size: 100 kDa
    why is the actual band size different from the predicted?


    Exposure time: 1 second


    Blocking/Dilution Buffer: 5% NFDM/TBST

     

  • All lanes : Anti-LAG-3 antibody [EPR20294-77] (ab209238) at 1/1000 dilution

    Lane 1 : Untreated mouse splenocytes whole cell lysate
    Lane 2 : Mouse splenocytes treated with 5 ug/ml concanavalin A for 72 hours whole cell lysate

    Lysates/proteins at 10 µg per lane.

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

    Developed using the ECL technique.

    Performed under reducing conditions.

    Predicted band size: 57 kDa
    Observed band size: 54, 70 kDa why is the actual band size different from the predicted?


    Exposure time: 10 seconds


    Blocking/Dilution Buffer: 5% NFDM/TBST

    The blot was developed on a BIO-RAD® ChemiDoc™ MP instrument.

    LAG3 expression can be elevated in activated splenocytes (PMID 19631993).

    The molecular mass observed is consistent with the literature (PMID 15971272; PMID 15557174).

  • LAG-3 was immunoprecipitated from 0.35 mg of mouse splenocytes (treated with 5 μg/ml concanavalin A for 72 hours) whole cell lysate with ab209238 at 1/30 dilution. Western blot was performed from the immunoprecipitate using ab209238 at 1/500 dilution. VeriBlot for IP Detection Reagent (HRP) (ab131366), was used for detection at 1/1000 dilution.

    Lane 1: Mouse splenocytes (treated with 5 μg/ml concanavalin A for 72 hours), whole cell lysate 10μg (Input). 

    Lane 2: ab209238 IP in mouse splenocytes (treated with 5 μg/ml concanavalin A for 72 hours), whole cell lysate. 

    Lane 3: Rabbit monoclonal IgG (ab172730) instead of ab209238 in mouse splenocytes (treated with 5 μg/ml concanavalin A for 72 hours), whole cell lysate.

    Blocking and dilution buffer: 5% NFDM/TBST.

    Exposure time: 1 second.

References

ab209238 has not yet been referenced specifically in any publications.

Customer reviews and Q&As

1-2 of 2 Abreviews or Q&A

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (tumor)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: Citrate
Permeabilization
No
Specification
tumor
Blocking step
Dako Protein Block as blocking agent for 30 minute(s) · Concentration: 100% · Temperature: 25°C
Fixative
Formaldehyde

Mr. Quentin LECOCQ

Verified customer

Submitted Jun 11 2019

Application
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections)
Sample
Mouse Tissue sections (kidney, spleen, lymph node)
Antigen retrieval step
Heat mediated - Buffer/Enzyme Used: citrate buffer
Permeabilization
Yes - 80% methanol
Specification
kidney, spleen, lymph node
Blocking step
Dako blocking solution as blocking agent for 15 minute(s) · Concentration: 100% · Temperature: 20°C
Fixative
Methanol

Nina Dvorina

Verified customer

Submitted Jan 10 2018

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