Recombinant Anti-ZAP70 antibody [YE291] - BSA and Azide free (ab247258)
Key features and details
- Produced recombinantly (animal-free) for high batch-to-batch consistency and long term security of supply
- Rabbit monoclonal [YE291] to ZAP70 - BSA and Azide free
- Suitable for: IHC-P, Flow Cyt (Intra), WB, IP, ICC/IF
- Reacts with: Human
Related conjugates and formulations
Overview
-
Product name
Anti-ZAP70 antibody [YE291] - BSA and Azide free
See all ZAP70 primary antibodies -
Description
Rabbit monoclonal [YE291] to ZAP70 - BSA and Azide free -
Host species
Rabbit -
Tested applications
Suitable for: IHC-P, Flow Cyt (Intra), WB, IP, ICC/IFmore details -
Species reactivity
Reacts with: Human -
Immunogen
Synthetic peptide. This information is proprietary to Abcam and/or its suppliers.
-
Positive control
- WB: Jurkat cell lysate. IHC-P: Human lymph node tissue. Flow Cyt (intra): Jurkat cells IP: Jurkat lysate.
-
General notes
ab247258 is the carrier-free version of ab32429.
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.
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 production
Our RabMAb® technology is a patented hybridoma-based technology for making rabbit monoclonal antibodies. For details on our patents, please refer to RabMAb® patents.
Properties
-
Form
Liquid -
Storage instructions
Shipped at 4°C. Store at +4°C. Do Not Freeze. -
Storage buffer
pH: 7.2
Constituent: PBS -
Carrier free
Yes -
Concentration information loading...
-
Purity
Protein A purified -
Clonality
Monoclonal -
Clone number
YE291 -
Isotype
IgG -
Research areas
Associated products
-
Alternative Versions
-
Conjugation kits
-
Isotype control
-
Recombinant Protein
Applications
The Abpromise guarantee
Our Abpromise guarantee covers the use of ab247258 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 |
---|---|---|
IHC-P |
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol.
|
|
Flow Cyt (Intra) |
Use at an assay dependent concentration.
|
|
WB |
Use at an assay dependent concentration. Detects a band of approximately 70 kDa (predicted molecular weight: 70 kDa).
|
|
IP |
Use at an assay dependent concentration.
|
|
ICC/IF |
Use at an assay dependent concentration.
|
Notes |
---|
IHC-P
Use at an assay dependent concentration. Perform heat mediated antigen retrieval with citrate buffer pH 6 before commencing with IHC staining protocol. |
Flow Cyt (Intra)
Use at an assay dependent concentration. |
WB
Use at an assay dependent concentration. Detects a band of approximately 70 kDa (predicted molecular weight: 70 kDa). |
IP
Use at an assay dependent concentration. |
ICC/IF
Use at an assay dependent concentration. |
Target
-
Function
Plays a role in T-cell development and lymphocyte activation. Essential for TCR-mediated IL-2 production. Isoform 1 induces TCR-mediated signal transduction, isoform 2 does not. -
Tissue specificity
Expressed in T- and natural killer cells. -
Involvement in disease
Defects in ZAP70 are the cause of selective T-cell defect (STD) [MIM:176947]. STD is an autosomal recessive form of severe combined immunodeficiency characterized by a selective absence of CD8-type T-cells. -
Sequence similarities
Belongs to the protein kinase superfamily. Tyr protein kinase family. SYK/ZAP-70 subfamily.
Contains 1 protein kinase domain.
Contains 2 SH2 domains. -
Domain
The SH2 domains bind to the phosphorylated tyrosine-based activation motif (TAM) of CD3Z and the non-canonical phosphorylated tyrosine-based activation motif (TAM) of RHOH. -
Post-translational
modificationsPhosphorylated on tyrosine residues upon T-cell antigen receptor (TCR) stimulation. Tyr-319 phosphorylation is essential for full activity. -
Cellular localization
Cytoplasm. Cell membrane. After antigen stimulation, isoform 1 concentrates at the immunological synapse and isoform 2 remains cytoplasmic. Co-localizes together with RHOH in the immunological synapse. RHOH is required for its proper localization to the cell membrane and cytoskeleton fractions in the thymocytes. - Information by UniProt
-
Database links
- Entrez Gene: 7535 Human
- Omim: 176947 Human
- SwissProt: P43403 Human
- Unigene: 234569 Human
-
Alternative names
- 70 kDa zeta associated protein antibody
- 70 kDa zeta-associated protein antibody
- EC 2.7.10.2 antibody
see all
Images
-
All lanes : Anti-ZAP70 antibody [YE291] (ab32429) at 1/500 dilution
Lane 1 : Wild-type Jurkat cell lysate
Lane 2 : ZAP70 CRISPR-Cas9 edited Jurkat cell lysate
Lane 3 : MOLT-4 cell lysate
Lane 4 : Raji cell lysate
Lysates/proteins at 20 µg per lane.
Performed under reducing conditions.
Predicted band size: 70 kDa
Observed band size: 70 kDaFalse colour image of Western blot: Anti-ZAP70 antibody [YE291] staining at 1/500 dilution, shown in green; Mouse anti-GAPDH antibody [6C5] (ab8245) loading control staining at 1/20000 dilution, shown in red. In Western blot, 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 ab273841 (CRISPR-Cas9 edited 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 (ab216773) and Goat anti-Mouse IgG H&L (IRDye® 680RD) preabsorbed (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 (ab32429).
-
All lanes : Anti-ZAP70 antibody [YE291] (ab32429) at 1/1000 dilution
Lane 1 : Jurkat (Human T cell leukemia T lymphocyte) whole cell lysate
Lane 2 : Raji (Human Burkitt's lymphoma B lymphocyte) whole cell lysate
Lysates/proteins at 20 µg per lane.
Secondary
All lanes : Goat anti-Rabbit IgG H&L (IRDye® 800CW) preadsorbed (ab216773) at 1/10000 dilution (Goat anti-Mouse IgG H&L (IRDye® 680RD) preadsorbed)
Predicted band size: 70 kDaAnti-GAPDH antibody, ab8245 (1/20000) was used as a primary antibody for the loading control and 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 – ab32429 observed at 70 kDa. Red - loading control ab8245 observed at 36 kDa.
ab32429 and 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 (ab216773) and 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 (ab32429).
-
This data was developed using 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 ab32429 at 1/50 dilution (2µg). 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 (ab172730) instead of ab32429 in Jurkat whole cell lysate
Blocking and dilution buffer and concentration: 5% NFDM/TBST.
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
-
Datasheet download
Certificate of Compliance
References (0)
ab247258 has not yet been referenced specifically in any publications.