Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Rabbit Recombinant Monoclonal ZAP70 antibody. Carrier free. Suitable for IP, WB, IHC-P, Flow Cyt (Intra) and reacts with Human samples.

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Immunoprecipitation - Anti-ZAP70 antibody [YE291] - BSA and Azide free (AB247258), expandable thumbnail

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Constituents: PBS
Form: Liquid
Clonality: Monoclonal

Immunogen

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

Reactivity data

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Product promiseTestedExpectedPredictedNot recommended

	IP	WB	IHC-P	Flow Cyt (Intra)
Human	Tested	Tested	Tested	Tested

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes Perform heat-mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.

Tested
Tested

Species	Dilution info	Notes
Species Human	Dilution info -	Notes -

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Target data

See full target information ZAP70

Function

Tyrosine kinase that plays an essential role in regulation of the adaptive immune response. Regulates motility, adhesion and cytokine expression of mature T-cells, as well as thymocyte development. Contributes also to the development and activation of primary B-lymphocytes. When antigen presenting cells (APC) activate T-cell receptor (TCR), a serie of phosphorylations lead to the recruitment of ZAP70 to the doubly phosphorylated TCR component CD247/CD3Z through ITAM motif at the plasma membrane. This recruitment serves to localization to the stimulated TCR and to relieve its autoinhibited conformation. Release of ZAP70 active conformation is further stabilized by phosphorylation mediated by LCK. Subsequently, ZAP70 phosphorylates at least 2 essential adapter proteins: LAT and LCP2. In turn, a large number of signaling molecules are recruited and ultimately lead to lymphokine production, T-cell proliferation and differentiation. Furthermore, ZAP70 controls cytoskeleton modifications, adhesion and mobility of T-lymphocytes, thus ensuring correct delivery of effectors to the APC. ZAP70 is also required for TCR-CD247/CD3Z internalization and degradation through interaction with the E3 ubiquitin-protein ligase CBL and adapter proteins SLA and SLA2. Thus, ZAP70 regulates both T-cell activation switch on and switch off by modulating TCR expression at the T-cell surface. During thymocyte development, ZAP70 promotes survival and cell-cycle progression of developing thymocytes before positive selection (when cells are still CD4/CD8 double negative). Additionally, ZAP70-dependent signaling pathway may also contribute to primary B-cells formation and activation through B-cell receptor (BCR).

Alternative names

SRK, ZAP70, Tyrosine-protein kinase ZAP-70, 70 kDa zeta-chain associated protein, Syk-related tyrosine kinase

Recommended products

Rabbit Recombinant Monoclonal ZAP70 antibody. Carrier free. Suitable for IP, WB, IHC-P, Flow Cyt (Intra) and reacts with Human samples.

Key facts

Isotype: IgG
Host species: Rabbit
Storage buffer: pH: 7.2 - 7.4
Constituents: PBS
Form: Liquid
Clonality: Monoclonal
Immunogen: The exact immunogen used to generate this antibody is proprietary information.
Carrier free: Yes
Clone number: YE291
Purification technique: Affinity purification Protein A
Concentration

Storage

Shipped at conditions: Blue Ice
Appropriate short-term storage conditions: +4°C
Appropriate long-term storage conditions: +4°C
Storage information: Do Not Freeze

Notes

ab247258 is the carrier-free version of Anti-ZAP70 antibody [YE291] ab32429.

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 monoclonal antibody, which offers several advantages including:

- High batch-to-batch consistency and reproducibility
- Improved sensitivity and specificity
- Long-term security of supply
- Animal-free batch production

For more information, read more on recombinant antibodies.

Our carrier-free antibodies are typically supplied in a PBS-only formulation, purified and free of BSA, sodium azide and glycerol. The carrier-free buffer and high concentration allow for increased conjugation efficiency.

This conjugation-ready format is designed for use with fluorochromes, metal isotopes, oligonucleotides, and enzymes, which makes them ideal for antibody labelling, functional and cell-based assays, flow-based assays (e.g. mass cytometry) and Multiplex Imaging applications.

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.

This product is compatible with the Maxpar^® Antibody Labeling Kit from Fluidigm, without the need for antibody preparation. Maxpar^® is a trademark of Fluidigm Canada Inc.

Supplementary info

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

Activity summary

ZAP70 also known as Zeta-chain-associated protein kinase 70 is a tyrosine kinase that plays an important role in T-cell receptor signaling. The protein has a molecular mass of approximately 70 kDa. ZAP70 is mainly expressed in T cells and natural killer (NK) cells. It gets activated upon T-cell receptor engagement initiating downstream signaling cascades essential for T-cell function and adaptive immune response.

Biological function summary

ZAP70 is instrumental in the signaling cascade that activates T cells in response to antigen recognition. It forms part of a larger signaling complex following the engagement of the T-cell receptor with antigens. This complex transmits critical activation signals leading to further cellular responses such as cytokine production and cell proliferation. ZAP70's function ensures appropriate immune responses and helps maintain immune system balance.

Pathways

ZAP70 plays an integral role in T-cell receptor signaling pathways. This protein interacts with other signaling molecules like Lck and LAT facilitating the transmission of activation signals within the cell. As part of the T-cell activation pathway ZAP70 helps to bridge receptor-ligand interactions with cellular responses enabling the immune cells to combat infections effectively.

Associated diseases and disorders

ZAP70 mutations or dysregulations can lead to severe immunodeficiencies or autoimmune diseases. For example ZAP70 deficiency results in severe combined immunodeficiency (SCID) which severely impairs immune function. Additionally aberrant ZAP70 activity has links to certain autoimmune disorders like rheumatoid arthritis where it influences abnormal immune signaling. Understanding the biochemical pathways that involve ZAP70 provides insight into managing these diseases and developing therapeutic strategies.

Product promise

We are dedicated to supporting your work with high quality reagents and we are here for you every step of the way should you need us.

In the unlikely event of one of our products not working as expected, you are covered by our product promise.

Full details and terms and conditions can be found here:
Terms & Conditions.

12 product images

Immunoprecipitation - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using Anti-ZAP70 antibody [YE291] ab32429, the same antibody clone in a different buffer formulation.ZAP70 was immunoprecipitated from 0.35 mg Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 µg with Anti-ZAP70 antibody [YE291] ab32429 at 1/50 dilution (2µg). VeriBlot for IP Detection Reagent (HRP)(VeriBlot for IP Detection Reagent (HRP) ab131366) was used at 1/5000 dilution.
Lane 1: Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate 10 µg
Lane 2: abab32429 IP in Jurkat whole cell lysate
Lane 3: Rabbit monoclonal IgG (Rabbit IgG, monoclonal [EPR25A] - Isotype Control ab172730) instead of Anti-ZAP70 antibody [YE291] ab32429 in Jurkat whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
All lanes: Immunoprecipitation - Anti-ZAP70 antibody [YE291] (Anti-ZAP70 antibody [YE291] ab32429)
Predicted band size: 69 kDa
Observed band size: 70 kDa
Western blot - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
False colour image of Western blot: Anti-ZAP70 antibody [YE291] staining at 1/500 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, Anti-ZAP70 antibody [YE291] ab32429 was shown to bind specifically to ZAP70. A band was observed at 70 kDa in wild-type Jurkat cell lysates with no signal observed at this size in ZAP70 CRISPR-Cas9 edited cell line Human ZAP70 knockout Jurkat cell line ab273841 (CRISPR-Cas9 edited cell lysate Human ZAP70 knockout Jurkat cell lysate ab273795). The band observed in the CRISPR-Cas9 edited lysate lane below 70 kDa is likely to represent a truncated form of ZAP70. This has not been investigated further and the functional properties of the gene product have not been determined. To generate this image, wild-type and ZAP70 CRISPR-Cas9 edited Jurkat cell lysates were analysed. First, samples were run on an SDS-PAGE gel then transferred onto a nitrocellulose membrane. Membranes were blocked in 3 % milk in TBS-0.1 % Tween® 20 (TBS-T) 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.
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
All lanes: Western blot - Anti-ZAP70 antibody [YE291] (Anti-ZAP70 antibody [YE291] ab32429) at 1/500 dilution
Lane 1: Wild-type Jurkat cell lysate at 20 µg
Lane 2: ZAP70 CRISPR-Cas9 edited Jurkat cell lysate at 20 µg
Lane 2: Western blot - Human ZAP70 knockout Jurkat cell line (Human ZAP70 knockout Jurkat cell line ab273841)
Lane 3: MOLT-4 cell lysate at 20 µg
Lane 4: Raji cell lysate at 20 µg
Performed under reducing conditions.
Predicted band size: 69 kDa
Observed band size: 70 kDa
Western blot - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
Anti-GAPDH antibody, Anti-GAPDH antibody [6C5] - Loading Control ab8245 (1/20000) was used as a primary antibody for the loading control and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed, Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776 (1/10000) was used as a loading control secondary antibody.
Lanes 1-2: Merged signal (red and green). Green – Anti-ZAP70 antibody [YE291] ab32429 observed at 70 kDa. Red - loading control Anti-GAPDH antibody [6C5] - Loading Control ab8245 observed at 36 kDa.
Anti-ZAP70 antibody [YE291] ab32429 and Anti-GAPDH antibody [6C5] - Loading Control (Anti-GAPDH antibody [6C5] - Loading Control ab8245) were incubated overnight at 4°C at 1 in 1000 dilution and 1 in 20000 dilution respectively. Blots were developed with Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed ab216776) secondary antibodies at 1 in 10000 dilution for 1 hour at room temperature before imaging.
The expression profile observed in Raji is consistent with the literature (PMID: 25275600).
Negative control: Raji (PMID: 25275600)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
All lanes: Western blot - Anti-ZAP70 antibody [YE291] (Anti-ZAP70 antibody [YE291] ab32429) at 1/1000 dilution
Lane 1: Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate at 20 µg
Lane 2: Raji (Human Burkitt's lymphoma B lymphocyte) whole cell lysate at 20 µg
Secondary
All lanes: Western blot - Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed ab216773) at 1/10000 dilution
Predicted band size: 69 kDa
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
Anti-ZAP70 antibody [YE291] ab32429, at a 1/100 dilution, staining ZAP70 in paraffin embedded human lymph node tissue by immunohistochemistry.
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Flow Cytometry (Intracellular) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
Intracellular Flow Cytometry analysis of Jurkat cells labeling ZAP70 with Anti-ZAP70 antibody [YE291] ab32429 at 1/80 dilution (red) or rabbit IgG as negative control (green).
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
Anti-ZAP70 antibody [YE291] ab32429 showing positive staining in Normal tonsil tissue.
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
Anti-ZAP70 antibody [YE291] ab32429 showing negative staining in Normal liver tissue.
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
Anti-ZAP70 antibody [YE291] ab32429 showing negative staining in Normal heart tissue.
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
Anti-ZAP70 antibody [YE291] ab32429 showing negative staining in Normal kidney tissue.
Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
Flow Cytometry (Intracellular) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using Anti-ZAP70 antibody [YE291] ab32429, the same antibody clone in a different buffer formulation.
Flow cytometry staining of human peripheral blood mononuclear cells (PBMCs) with Anti-ZAP70 antibody [YE291] ab32429 (right) or Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (left). Cells were fixed and permeabilised with BD Cytofix/Cytoperm™ for 20 min. PBMCs were incubated for 30 min on ice in 1x PBS containing 10 µg/ml human IgG and 10 % normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody Anti-ZAP70 antibody [YE291] ab32429 or Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (1x 10⁶in 100 µl at 0.04 µg/ml (1/12500)) for 30 min at 22°C . The cells were simultaneously stained with CD3.
The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C
Acquisition of >30000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter. Events were gated on live cells.
Flow Cytometry (Intracellular) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
Flow cytometry staining of human peripheral blood mononuclear cells (PBMCs) with Anti-ZAP70 antibody [YE291] ab32429 (right) or Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (left). PBMCs were incubated for 30 min on ice in 1x PBS containing 10 µg/ml human IgG and 10 % normal goat serum to block FC receptors and non-specific protein-protein interaction after which staining with CD3 occurred. The cells were then fixed and permeabilised with BD Cytofix/Cytoperm™ for 20 min. Cells were then stained with the antibody Anti-ZAP70 antibody [YE291] ab32429 or Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (1x 10⁶ in 100 µl at 0.04 μg/ml (1/50,000 dilution)) for 30 min at 22°C.
The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C.
Acquisition of >30000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter. Events were gated on live cells.
Flow Cytometry (Intracellular) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
This data was developed using the same antibody clone in a different buffer formulation containing PBS, BSA, glycerol, and sodium azide (Anti-ZAP70 antibody [YE291] ab32429).
Flow cytometry staining of human peripheral blood mononuclear cells (PBMCs) with Anti-ZAP70 antibody [YE291] ab32429 (right) or Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (left). Cells were fixed and permeabilised with BD Cytofix/Cytoperm™ for 20 min. PBMCs were incubated for 30 min on ice in 1x PBS containing 10 µg/ml human IgG and 10 % normal goat serum to block FC receptors and non-specific protein-protein interaction followed by the antibody Anti-ZAP70 antibody [YE291] ab32429 or Recombinant Rabbit IgG, monoclonal [EPR25A] - Isotype Control (1x 10⁶ in 100 µl at 0.04 μg/ml (1/50000)) for 30 min at 22°C . The cells were simultaneously stained with CD19.
The secondary antibody Goat Anti-Rabbit IgG H&L (Alexa Fluor^® 488) preadsorbed was incubated at 1/4000 for 30min at 22°C
Acquisition of >30000 events were collected using a 50 mW Blue laser (488nm) and 525/40 bandpass filter. Events were gated on live cells.

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Anti-ZAP70 antibody [YE291] - BSA and Azide free

Images

Key facts

Immunogen

Reactivity data

IP

TestedTested

WB

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IHC-P

TestedTested

Flow Cyt (Intra)

TestedTested

Associated Products

1 product for Alternative Product

2 products for Alternative Version

Target data

Function

Alternative names

Recommended products

Key facts

Storage

Notes

Supplementary info

Product promise

Tested

Expected

Predicted

Not recommended

12 product images

Immunoprecipitation - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Western blot - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Western blot - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Flow Cytometry (Intracellular) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Immunohistochemistry (Formalin/PFA-fixed paraffin-embedded sections) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Flow Cytometry (Intracellular) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Flow Cytometry (Intracellular) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Flow Cytometry (Intracellular) - Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)

Downloads

Product protocols

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