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Rabbit Recombinant Monoclonal LAG-3 antibody. Suitable for IP, Flow Cyt (Intra), WB, IHC-P and reacts with Human samples.

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Images

Immunoprecipitation - Anti-LAG-3 antibody [CAL25] (AB237718), expandable thumbnail
  • Western blot - Anti-LAG-3 antibody [CAL25] (AB237718), expandable thumbnail
  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [CAL25] (AB237718), expandable thumbnail
  • Flow Cytometry (Intracellular) - Anti-LAG-3 antibody [CAL25] (AB237718), expandable thumbnail
  • Western blot - Anti-LAG-3 antibody [CAL25] (AB237718), expandable thumbnail

Key facts

Isotype
IgG
Host species
Rabbit
Storage buffer

pH: 7.5
Preservative: 0.05% Sodium azide
Constituents: PBS

Form
Liquid
Clonality
Monoclonal

Immunogen

  • The exact immunogen used to generate this antibody is proprietary information.

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Reactivity data

Select an application
Product promiseTestedExpectedPredictedNot recommended
IPFlow Cyt (Intra)WBICC/IFIHC-P
Human
Tested
Tested
Tested
Not recommended
Tested

Tested
Tested

Species
Human
Dilution info
1/30
Notes

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

Tested
Tested

Species
Human
Dilution info
1/50
Notes

-

Tested
Tested

Species
Human
Dilution info
1/1000
Notes

-

Not recommended
Not recommended

Species
Human
Dilution info
-
Notes

-

Tested
Tested

Species
Human
Dilution info
0.3 µg/mL
Notes

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

Associated Products

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Target data

Function

Lymphocyte activation gene 3 protein: Inhibitory receptor on antigen activated T-cells (PubMed:20421648, PubMed:7805750, PubMed:8647185). Delivers inhibitory signals upon binding to ligands, such as FGL1 (By similarity). FGL1 constitutes a major ligand of LAG3 and is responsible for LAG3 T-cell inhibitory function (By similarity). Following TCR engagement, LAG3 associates with CD3-TCR in the immunological synapse and directly inhibits T-cell activation (By similarity). May inhibit antigen-specific T-cell activation in synergy with PDCD1/PD-1, possibly by acting as a coreceptor for PDCD1/PD-1 (By similarity). Negatively regulates the proliferation, activation, effector function and homeostasis of both CD8(+) and CD4(+) T-cells (PubMed:20421648, PubMed:7805750, PubMed:8647185). Also mediates immune tolerance: constitutively expressed on a subset of regulatory T-cells (Tregs) and contributes to their suppressive function (By similarity). Also acts as a negative regulator of plasmacytoid dendritic cell (pDCs) activation (By similarity). Binds MHC class II (MHC-II); the precise role of MHC-II-binding is however unclear (PubMed:8647185). Secreted lymphocyte activation gene 3 protein. May function as a ligand for MHC class II (MHC-II) on antigen-presenting cells (APC), promoting APC activation/maturation and driving Th1 immune response.

Alternative names

CD223, FDC, LAG3, Lymphocyte activation gene 3 protein, LAG-3

Recommended products

Rabbit Recombinant Monoclonal LAG-3 antibody. Suitable for IP, Flow Cyt (Intra), WB, IHC-P and reacts with Human samples.

Key facts

Isotype
IgG
Form
Liquid
Clonality
Monoclonal
Immunogen
  • The exact immunogen used to generate this antibody is proprietary information.
Clone number
CAL25
Purification technique
Affinity purification Protein A
Concentration
Loading...
Purification notes

Purity >99%.

Storage

Shipped at conditions
Blue Ice
Appropriate short-term storage duration
1-2 weeks
Appropriate short-term storage conditions
+4°C
Appropriate long-term storage conditions
-20°C
Aliquoting information
Upon delivery aliquot
Storage information
Avoid freeze / thaw cycle

Supplementary info

This supplementary information is collated from multiple sources and compiled automatically.
Activity summary

LAG-3 also known as CD223 is a transmembrane protein with a mass of approximately 70 kDa. It acts as a checkpoint molecule similar to CTLA-4 and PD-1 in the immune system. LAG-3 is expressed on activated T cells natural killer cells and some dendritic cells. It interacts with its ligand MHC class II leading to inhibition of cellular proliferation and cytokine secretion an important part of immune response regulation.

Biological function summary

LAG-3 is involved in maintaining immune homeostasis and preventing autoimmunity. It is not part of a complex but works in synergy with other immune checkpoint molecules. When interacting with MHC class II LAG-3 transmits inhibitory signals that attenuate immune cell responses. Its role in immune regulation helps to balance immune activation and tolerance which is essential for preventing tissue damage while protecting against pathogens.

Pathways

The inhibitory role of LAG-3 integrates into the T cell receptor (TCR) signaling pathway and is also linked to the PD-1 signaling pathway. In TCR signaling LAG-3 negatively regulates T cell activation. LAG-3 is functionally related to proteins like PD-1 and CTLA-4 sharing similar roles in immune checkpoint pathways that are critical for maintaining immune balance.

Associated diseases and disorders

LAG-3 has links to cancer and autoimmune diseases. In cancer LAG-3 expression on tumor-infiltrating lymphocytes correlates with immune evasion by tumors. This relationship appears in various cancers including melanoma and renal cell carcinoma. In autoimmune diseases LAG-3's expression on regulatory T cells helps control conditions such as multiple sclerosis. It collaborates with CTLA-4 in modulating immune responses that prevent excessive immune activation and tissue damage associated with autoimmunity.

Product promise

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7 product images

  • Immunoprecipitation - Anti-LAG-3 antibody [CAL25] (ab237718), expandable thumbnail

    Immunoprecipitation - Anti-LAG-3 antibody [CAL25] (ab237718)

    LAG-3 was immunoprecipitated from 0.35 mg HEK-293T (human embryonic kidney epithelial cell) transfected with GFP-tagged LAG-3 expression vector whole cell lysate using ab237718 at 1/30 dilution. Western blot was performed on the immunoprecipitate using ab237718 at 1/1000 dilution. VeriBlot for IP Detection Reagent (HRP) (VeriBlot for IP Detection Reagent (HRP) ab131366) was used as the secondary antibody at 1/5000 dilution.

    Lane 1: HEK-293T transfected with GFP-tagged LAG-3 expression vector whole cell lysate 10 μg (input).
    Lane 2: ab237718 IP in HEK-293T GFP-tagged LAG-3 expression vector whole cell lysate.
    Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of ab237718 in 293T transfected with GFP-tagged LAG-3 expression vector whole cell lysate.

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

    Exposure time: 30 seconds.

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

    All lanes: Immunoprecipitation - Anti-LAG-3 antibody [CAL25] (ab237718)

    Predicted band size: 57 kDa

    Observed band size: 16 kDa, 70 kDa

  • Western blot - Anti-LAG-3 antibody [CAL25] (ab237718), expandable thumbnail

    Western blot - Anti-LAG-3 antibody [CAL25] (ab237718)

    False colour image of Western blot: Anti-LAG-3 antibody [CAL25] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (Anti-GAPDH antibody [6C5] - Loading Control ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab237718 was shown to bind specifically to LAG-3. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in fluorescent western blot (TBS-based) blocking solution before incubation with primary antibodies overnight at 4°C. Blots were washed four times in TBS-T, incubated with secondary antibodies for 1 h at room temperature, washed again four times then imaged. Secondary antibodies used were Goat anti-Rabbit IgG H&L (IRDye® 800CW) preabsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) at 1/20000 dilution.

    All lanes: Western blot - Anti-LAG-3 antibody [CAL25] (ab237718) at 1/1000 dilution

    Lane 1: HDLM-2 cell lysate at 20 µg

    Lane 2: Jurkat cell lysate at 20 µg

    Performed under reducing conditions.

    Predicted band size: 57 kDa

    Observed band size: 57 kDa, 70 kDa

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [CAL25] (ab237718), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [CAL25] (ab237718)

    Immunohistochemical analysis of paraffin-embedded human tonsil tissue labeling LAG-3 using ab237718 at 1/800 dilution, followed by a ready to use Goat Anti-Rabbit IgG H&L (HRP). Positive staining on the human tonsil is observed. The section was incubated with ab237718 for 15 mins at room temperature. The immunostaining was performed on a Leica Biosystems BOND® RX instrument. Counterstained with Hematoxylin.

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

  • Flow Cytometry (Intracellular) - Anti-LAG-3 antibody [CAL25] (ab237718), expandable thumbnail

    Flow Cytometry (Intracellular) - Anti-LAG-3 antibody [CAL25] (ab237718)

    Flow cytometric analysis of 2% paraformaldehyde-fixed 0.1% Tween-20 permeabilized Human peripheral blood mononuclear cells (PBMC) treated with 10μg/ml PHA for 48h labeling LAG-3 with ab237718 at 1/50 dilution (Right) compared to a Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730, Left). Goat anti rabbit IgG (Alexa Fluor® 488, Goat Anti-Rabbit IgG Fc (Alexa Fluor® 488) preadsorbed ab150097) was used as the secondary at 1/5000 dilution.

    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 ab237718 (Right).

  • Western blot - Anti-LAG-3 antibody [CAL25] (ab237718), expandable thumbnail

    Western blot - Anti-LAG-3 antibody [CAL25] (ab237718)

    Blocking/diluting buffer and concentration: 5% NFDM/TBST

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

    All lanes: Western blot - Anti-LAG-3 antibody [CAL25] (ab237718) at 1/1000 dilution

    Lane 1: HEK-293T (human embryonic kidney epithelial cell) whole cell lysate at 20 µg

    Lane 2: HEK-293T transfected with GFP-tagged LAG-3 expression vector whole cell lysate at 20 µg

    Secondary

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

    Predicted band size: 57 kDa

    Observed band size: 16 kDa, 70 kDa

    Exposure time: 3min

  • Western blot - Anti-LAG-3 antibody [CAL25] (ab237718), expandable thumbnail

    Western blot - Anti-LAG-3 antibody [CAL25] (ab237718)

    Blocking buffer and concentration: 5% NFDM/TBST

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

    All lanes: Western blot - Anti-LAG-3 antibody [CAL25] (ab237718) at 1/1000 dilution

    Lane 1: Human peripheral blood mononuclear cell (PBMC) whole cell lysate at 20 µg

    Lane 2: Human peripheral blood mononuclear cell (PBMC) treated with 10μg/ml PHA for 48h whole cell lysate at 20 µg

    Secondary

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

    Predicted band size: 57 kDa

    Observed band size: 16 kDa, 70 kDa

    Exposure time: 3min

  • Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [CAL25] (ab237718), expandable thumbnail

    Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [CAL25] (ab237718)

    Formalin-fixed, paraffin-embedded human tonsil tissue stained for LAG-3 using ab237718 at 0.3 μg/ml in immunohistochemical analysis.

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