Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)

Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176) is part of the multiplex kits range. Abcam offers high-quality biological reagents and tools including antibodies, proteins, assays, cell lines and lysates.

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Images

Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (AB278176), expandable thumbnail

Key facts

Reactive species: Human
Target: CD40LG
(See target data below)

Target data

See full target information CD40LG

Function

Cytokine that acts as a ligand to CD40/TNFRSF5 (PubMed:1280226, PubMed:31331973). Costimulates T-cell proliferation and cytokine production (PubMed:8617933). Its cross-linking on T-cells generates a costimulatory signal which enhances the production of IL4 and IL10 in conjunction with the TCR/CD3 ligation and CD28 costimulation (PubMed:8617933). Induces the activation of NF-kappa-B (PubMed:15067037, PubMed:31331973). Induces the activation of kinases MAPK8 and PAK2 in T-cells (PubMed:15067037). Induces tyrosine phosphorylation of isoform 3 of CD28 (PubMed:15067037). Mediates B-cell proliferation in the absence of co-stimulus as well as IgE production in the presence of IL4 (By similarity). Involved in immunoglobulin class switching (By similarity). CD40 ligand, soluble form. Acts as a ligand for integrins, specifically ITGA5:ITGB1 and ITGAV:ITGB3; both integrins and the CD40 receptor are required for activation of CD40-CD40LG signaling, which have cell-type dependent effects, such as B-cell activation, NF-kappa-B signaling and anti-apoptotic signaling.

Additional Targets

TNFSF4, TNFRSF9, ICOSLG, CD40, ICOS, TNFSF9, TNFRSF4

Alternative names

CD154, CD40L, TNFSF5, TRAP, CD40LG, CD40 ligand, CD40-L, T-cell antigen Gp39, TNF-related activation protein, Tumor necrosis factor ligand superfamily member 5

What's included?

1 Kit

Components

ab254385

Anti-4-1BBL antibody [EPR21995-147]

2 x 10 µL

ab264466

Anti-CD134 / OX40L receptor antibody [EPR23000-42]

2 x 10 µL

ab209256

Anti-CD137 antibody [EPR20238]

2 x 10 µL

ab224639

Anti-CD40 antibody [EPR20735]

2 x 10 µL

ab209262

Anti-ICOS Ligand/ICOSL antibody [EPR20240]

2 x 10 µL

ab224644

Anti-ICOS antibody [EPR20560]

2 x 10 µL

ab76130

Anti-OX40L/TNFSF4 antibody [EP1168Y]

2 x 10 µL

ab52750

Anti-TRAP/CD40L antibody [EP462E]

2 x 10 µL

Key facts

Reactive species: Human
Target: CD40LG

Storage

Shipped at conditions: Blue Ice
Appropriate short-term storage conditions: -20°C
Appropriate long-term storage conditions: -20°C
Storage information: -20°C

Notes

Stimulatory Immune Checkpoint panel 1 - Human WB ab278176 contains multiple trial-sized versions of anti-human antibody clones against OX40L/TNFSF4, CD134/OX40L, ICOS, ICOS Ligand/ICOSL, CD137, 4-1BBL, CD40 and TRAP/CD40L specifically selected for high performance in various applications. They are provided as a sampler panel to allow you to easily evaluate each antibody.

For guidelines on how to use each antibody within the panel, please consult the individual datasheet for each antibody.

Panel contains:

- Rabbit monoclonal [EP1168Y] to OX40L/TNFSF4 (20 μL) ab76130

- Rabbit monoclonal [EPR23000-42] to CD134 / OX40L receptor (20 μL) ab264466

- Rabbit monoclonal [EPR20560] to ICOS (20 μL) ab224644

- Rabbit monoclonal [EPR20240] to ICOS Ligand/ICOSL (20 μL) ab209262

- Rabbit monoclonal [EPR20238] to CD137 (20 μL) ab209256

- Rabbit monoclonal [EPR21995-147] to 4-1BBL (20 μL) ab254385

- Rabbit monoclonal [EPR20735] to CD40 (20 μL) ab224639

- Rabbit monoclonal [EP462E] to TRAP/CD40L (20 μL) ab52750

Explore our range of antibody sample panels designed to provide you with a variety of trial-size antibodies in a convenient and cost-effective format.

Carrier-free formulations of our recombinant antibodies are also available for easy conjugation to labels of your choice and for multiplex applications. Use our intuitive search and select carrier-free or your label of choice. For bespoke conjugations or large volumes email bespoke@abcam.com.

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Full details and terms and conditions can be found here:
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16 product images

Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-TRAP/CD40L antibody [EP462E] (Anti-TRAP/CD40L antibody [EP462E] ab52750) at 1/5000 dilution.
Lane 1: MOLT-4 (human lymphoblastic leukemia cell line) cell lysate, 10 ug
Secondary (all lanes): Goat anti-rabbit HRP labeled at 1/2000 dilution.
Predicted MW: 29 kDa.
Observed MW: 36 kDa.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-TRAP/CD40L antibody [EP462E] (Anti-TRAP/CD40L antibody [EP462E] ab52750) at 1μg/ml.
Lane 1: Human lymph node tissue lysate, 20 μg
Lane 2: Mouse lymph node tissue lysate, 20 μg
Secondary (all lanes): Goat Anti-Rabbit IgG (HRP), specific to the non-reduced form of IgG at 1/2000 dilution.
Predicted MW: 29 kDa.
Observed MW: 36 kDa.
Blocking and diluting buffer: 5% NFDM/TBST.

Exposure time: Lane 1: 90 seconds
Lane 2: 20 seconds
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-CD40 antibody [EPR20735] (Anti-CD40 antibody [EPR20735] ab224639) at 1/1000 dilution.
Lane 1: Human lymph node tissue lysate, 20 μg
Lane 2:Human lymphoma tissue lysate, 20 μg

Lane 3: Raji (human Burkitt's lymphoma cell line) whole cell lysate, 20 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/20000 dilution.
Predicted MW: 30 kDa.
Observed MW: 42 kDa
Blocking/Dilution buffer: 5% NFDM/TBST.
Exposure: 15 seconds
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-4-1BBL antibody [EPR21995-147] (Anti-4-1BBL antibody [EPR21995-147] ab254385) at 1/1000 dilution.
Lane 1: Raji (human Burkitts lymphoma cell line) whole cell lysate, 30 μg
Lane 2: K562 (Human chronic myelogenous leukemia cell line from bone marrow) whole cell lysate, 60 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/20000 dilution.
Predicted MW: 27 kDa.
Observed MW: 26 kDa
Blocking and diluting buffer and concentration: 5% NFDM/TBST.
Low expression control: K562 (PMID: 20670353).
Exposure time: 3 minutes.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-CD137 antibody [EPR20238] (Anti-CD137 antibody [EPR20238] ab209256) at 1/1000 dilution.
Lane 1: Human lymphoma lysate, 20 μg
Lane 2:Human fetal spleen lysate, 20 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/50000 dilution.
Predicted MW: 28 kDa.
Observed MW: 36 kDa.
Blocking/Dilution buffer: 5% NFDM/TBST.
The MW observed is consistent with the literature: PMID 10528184; PMID 17560335.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-CD137 antibody [EPR20238] (Anti-CD137 antibody [EPR20238] ab209256) at 1/1000 dilution.
Lane 1: H9 (Human cutaneous T Lymphocyte cell line) whole cell lysate, 20 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/100000 dilution.
Predicted MW: 28 kDa.
Observed MW: 36, 40 kDa.
Blocking/Dilution buffer: 5% NFDM/TBST.
The MW observed is consistent with the literature: PMID 10528184; PMID 17560335.
Exposure time: 3 mins.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-CD137 antibody [EPR20238] (Anti-CD137 antibody [EPR20238] ab209256) at 1/1000 dilution.
Lane 1: Whole cell lysate from untreated Jurkat (Human T cell leukemia cell line from peripheral blood), 10 μg
Lane 2:Whole cell lysate from Jurkat cells treated with 10ng/ml Phorbol-12-myristate-13-acetate (TPA) and 1μM Ionomycin (Ionomycin Ca2+ Salt, Ca2+ ionophore ab120116) for 48 hours, 10 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/100000 dilution.
Predicted MW: 28 kDa.
Observed MW: 36, 40 kDa.
Blocking/Dilution buffer: 5% NFDM/TBST.
The expression level of CD137 can be induced by TPA and Ionomycin [PMID 14648705].
The MW observed is consistent with the literature: PMID 10528184; PMID 17560335.
Exposure time: 3 mins.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-ICOS Ligand/ICOSL antibody [EPR20240] (Anti-ICOS Ligand/ICOSL antibody [EPR20240] ab209262) at 1/20000 dilution.
Lane 1: His-tagged recombinant human ICOS ligand/ICOSL protein, 0.01 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/100000 dilution.
Predicted MW: 33 kDa.
Observed MW: 50 kDa.
Blocking/Dilution buffer: 5% NFDM/TBST.
Exposure time: 10 secs.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-ICOS Ligand/ICOSL antibody [EPR20240] (Anti-ICOS Ligand/ICOSL antibody [EPR20240] ab209262) at 1/1000 dilution.
Lane 1: Untreated JAR (Human placenta choriocarcinoma cell line) whole cell lysate, 20 μg
Lane 2: JAR whole cell lysate treated with PNGase F, 20 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/20000 dilution.
Predicted MW: 33 kDa.
Observed MW: 60 kDa (glycosylated form); 35 kDa (non-glycosylated form).
Blocking/Dilution buffer: 5% NFDM/TBST.
ICOS ligand/ICOSL is highly glycosylated (PMID 11983910).
Exposure time: 30 secs.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-ICOS Ligand/ICOSL antibody [EPR20240] (Anti-ICOS Ligand/ICOSL antibody [EPR20240] ab209262) at 1/1000 dilution.
Lane 1: Untreated HUVEC (Human umbilical vein endothelial cell line) whole cell lysate, 20 μg
Lane 2: HUVEC whole cell lysate treated with tumor necrosis factor-α (TNF-alpha, Recombinant human TNF alpha protein ab9642) 200 unit/ml for 24 hours, 20 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/20000 dilution.
Predicted MW: 33 kDa.
Observed MW: 60kDa.
Blocking/Dilution buffer: 5% NFDM/TBST.
The level of ICOS Ligand/ICOSL expression can be elevated by TNF-alpha treatment (PMID 11983910).
Exposure time: 30 secs.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-ICOS antibody [EPR20560] (Anti-ICOS antibody [EPR20560] ab224644) at 1/1000 dilution.
Lane 1: Untreated human tonsil tissue lysate, 20 μg
Lane 2: Human tonsil tissue lysate treated with PNGase F, 20 μg

Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/20000 dilution.
Predicted MW: 22 kDa.
Observed MW: 22, 27-29 kDa.
Blocking and dilution buffer: 5% NFDM/TBST.
When untreated two distinct bands of 27kDa and 29kDa were observed. Treatment with PNGase F resulted in a single band with a molecular of 22kDa.
The expression profile observed is consistent with what has been described in the literature (PMID: 11169414 ).
Exposure time: 10 secs.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-ICOS antibody [EPR20560] (Anti-ICOS antibody [EPR20560] ab224644) at 1/1000 dilution.
Lane 1: Human tymus tissue lysate, 20 μg
Lane 2: HuT-78 (human Sezary syndrome cutaneous T lymphocyte) whole cell lysate, 20 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/20000 dilution.
Predicted MW: 22 kDa.
Observed MW: 27-29 kDa.
Blocking and dilution buffer: 5% NFDM/TBST.
The expression profile observed is consistent with what has been described in the literature (PMID: 11169414).
Exposure time: 10 secs.
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-ICOS antibody [EPR20560] (Anti-ICOS antibody [EPR20560] ab224644) at 1/1000 dilution.
Lane 1: His-tagged human ICOS (aa21-140-GGGGS-aa162-199) recombinant protein, 10 ng
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/100000 dilution.
Predicted MW: 22 kDa.
Observed MW: 17 kDa.
Blocking and dilution buffer: 5% NFDM/TBST
Exposure time: 1 second
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-CD134 / OX40L receptor antibody [EPR23000-42] (Anti-CD134 / OX40L receptor antibody [EPR23000-42] ab264466) at 1/1000 dilution.
Lane 1: Human thymus tissue lysate, 10 μg
Lane 2: Human lymph node, 10 μg

Secondary (all lanes): VeriBlot for IP Detection Reagent (HRP) ab131366 at 1/1000 dilution.
Predicted MW: 29 kDa.
Observed MW: 50 kDa.
Blocking and dilution buffer: 5% NFDM/TBST.
Exposure time: 3 minutes.
This blot was developed using a higher sensitivity ECL substrate.
The molecular weight observed is consistent with what has been described in the literature (PMID:9108415, 23897980).
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-CD134 / OX40L receptor antibody [EPR23000-42] (Anti-CD134 / OX40L receptor antibody [EPR23000-42] ab264466) at 1/1000 dilution.
Lane 1: Untreated human PBMC, whole cell lysate, 20 μg
Lane 2: Human PBMC treated with 10 μg/ml phytohaemagglutinin (PHA) for 2 days, whole cell lysate, 20 μg

Lane 3: Jurkat (human T cell leukemia T lymphocyte), whole cell lysate, 20 μg

Secondary (all lanes): VeriBlot for IP Detection Reagent (HRP) ab131366 at 1/500 dilution.
Predicted MW: 29 kDa.
Observed MW: 50, 37 kDa.
Blocking and dilution buffer: 5% NFDM/TBST.
Exposure time: 37 seconds.
The expression profile and molecular weight observed is consistent with what has been described in the literature (PMID:9108415,23897980).
Negative control: Jurkat (PMID:9108415).
Western blot - Stimulatory Immune Checkpoint panel 1 - Human WB (ab278176)
All lanes: Anti-OX40L/TNFSF4 antibody [EP1168Y] (Anti-OX40L/TNFSF4 antibody [EP1168Y] ab76130) at 1/1000 dilution.
Lane 1: HUVEC (human umbilical vein endothelial cell) whole cell lysate, 20 μg
Lane 2: A-172 (human brain glioblastoma) whole cell lysate, 20 μg

Lane 3: Raji (human burkitt's lymphoma b lymphocyte) whole cell lysate, 20 μg
Secondary (all lanes): Goat Anti-Rabbit IgG H&L (HRP) ab97051 at 1/100000 dilution.
Predicted MW: 21 kDa.
Observed MW: 30kDa.
OX40L/TNFSF4 is a glycoprotein. The expression profile and molecular weight observed is consistent with what has been described in the literature (PMID: 8642328)
Negative control: Raji (PMID: 8642328)
Exposure times: 6 secs.