Recombinant Anti-LAG-3 antibody [CAL26] (ab237719)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [CAL26] to LAG-3
- Suitable for: WB, Flow Cyt, IP, IHC-P
- Reacts with: Human
Related conjugates and formulations
Overview
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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 proprietary to Abcam and/or its suppliers.
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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.
Properties
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Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C short term (1-2 weeks). Upon delivery aliquot. Store at -20°C long term. Avoid freeze / thaw cycle. -
Storage buffer
pH: 7.50
Preservative: 0.05% Sodium azide
Constituent: PBS -
Concentration information loading...
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Purity
Protein A purified -
Purification notes
Purity >99%. -
Clonality
Monoclonal -
Clone number
CAL26 -
Isotype
IgG -
Research areas
Associated products
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Alternative Versions
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Compatible Secondaries
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Immunohistochemistry reagents
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Isotype control
Applications
The Abpromise guarantee
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 |
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WB |
1/1000. Detects a band of approximately 16, 70 kDa (predicted molecular weight: 57 kDa).
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Flow Cyt |
1/250.
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IP |
1/20.
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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.
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Notes |
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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. |
Target
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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
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Database links
- Entrez Gene: 3902 Human
- Omim: 153337 Human
- SwissProt: P18627 Human
- Unigene: 409523 Human
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Alternative names
- CD223 antibody
- CD223 antigen antibody
- FDC protein antibody
see all
Images
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All lanes : Anti-LAG-3 antibody [CAL26] (ab237719) at 1/1000 dilution
Lane 1 : HDLM-2 cell lysate
Lane 2 : Jurkat cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 57 kDa
Observed band size: 57,70 kDa why is the actual band size different from the predicted?False colour image of Western blot: Anti-LAG-3 antibody [CAL26] staining at 1/1000 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, ab237719 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 (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (ab216776) at 1/20000 dilution.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [CAL26] (ab237719)
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).
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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).
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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 minutesBlocking and dilution buffer: 5% NFDM/TBST.
This antibody also detects a 16kDa cleaved fragment. (PMID: 15557174).
This blot was developed using a higher sensitivity ECL substrate.
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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 minutesBlocking and dilution buffer: 5% NFDM/TBST.
This antibody also detects a 16kDa cleaved fragment. (PMID: 15557174).
This blot was developed using a higher sensitivity ECL substrate.
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Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-LAG-3 antibody [CAL26] (ab237719)
Formalin-fixed, paraffin-embedded human tonsil tissue stained for LAG-3 using ab237719 at 0.3 μg/ml in immunohistochemical analysis.
Protocols
To our knowledge, customised protocols are not required for this product. Please try the standard protocols listed below and let us know how you get on.
Datasheets and documents
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Datasheet download
References (0)
ab237719 has not yet been referenced specifically in any publications.