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Monoclonal Antibody Negative Regulation of Negative Chemotaxis

$303
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse

Application Methods: Western Blotting

Background: Vascular endothelial growth factor receptor 2 (VEGFR2, KDR, Flk-1) is a major receptor for VEGF-induced signaling in endothelial cells. Upon ligand binding, VEGFR2 undergoes autophosphorylation and becomes activated (1). Major autophosphorylation sites of VEGFR2 are located in the kinase insert domain (Tyr951/996) and in the tyrosine kinase catalytic domain (Tyr1054/1059) (2). Activation of the receptor leads to rapid recruitment of adaptor proteins, including Shc, GRB2, PI3 kinase, NCK, and the protein tyrosine phosphatases SHP-1 and SHP-2 (3). Phosphorylation at Tyr1212 provides a docking site for GRB2 binding and phospho-Tyr1175 binds the p85 subunit of PI3 kinase and PLCγ, as well as Shb (1,4,5). Signaling from VEGFR2 is necessary for the execution of VEGF-stimulated proliferation, chemotaxis and sprouting, as well as survival of cultured endothelial cells in vitro and angiogenesis in vivo (6-8).

$260
100 µl
APPLICATIONS
REACTIVITY
Mouse

Application Methods: Western Blotting

Background: Monocyte chemotactic protein-1 (MCP-1), also known as CCL2, monocyte chemotactic activating factor (MCAF) or glioma-derived chemotactic factor-2 (GDCF-2), is the product of the human JE gene and a member of the family of C-C (or β) chemokines (1-4). The predicted molecular weight of MCP-1 protein is 11-13 kDa, but it may migrate at 20-30 kDa due to glycosylation. MCP-1 is secreted by a variety of cell types in response to pro-inflammatory stimuli and was originally described for its chemotactic activity on monocytes. This activity has led to studies demonstrating its role in diseases characterized by monocyte infiltrates such as psoriasis (5), rheumatoid arthritis (6) and atherosclerosis (7). MCP-1 may also contribute to tumor progression and angiogenesis (8). Signaling by MCP-1 is mediated by the G-protein coupled receptor CCR2 (9).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Flow Cytometry, Immunofluorescence (Immunocytochemistry), Western Blotting

Background: Activation of protein kinase C (PKC) is one of the earliest events in a cascade that controls a variety of cellular responses, including secretion, gene expression, proliferation, and muscle contraction (1,2). PKC isoforms belong to three groups based on calcium dependency and activators. Classical PKCs are calcium-dependent via their C2 domains and are activated by phosphatidylserine (PS), diacylglycerol (DAG), and phorbol esters (TPA, PMA) through their cysteine-rich C1 domains. Both novel and atypical PKCs are calcium-independent, but only novel PKCs are activated by PS, DAG, and phorbol esters (3-5). Members of these three PKC groups contain a pseudo-substrate or autoinhibitory domain that binds to substrate-binding sites in the catalytic domain to prevent activation in the absence of cofactors or activators. Control of PKC activity is regulated through three distinct phosphorylation events. Phosphorylation occurs in vivo at Thr500 in the activation loop, at Thr641 through autophosphorylation, and at the carboxy-terminal hydrophobic site Ser660 (2). Atypical PKC isoforms lack hydrophobic region phosphorylation, which correlates with the presence of glutamic acid rather than the serine or threonine residues found in more typical PKC isoforms. The enzyme PDK1 or a close relative is responsible for PKC activation. A recent addition to the PKC superfamily is PKCμ (PKD), which is regulated by DAG and TPA through its C1 domain. PKD is distinguished by the presence of a PH domain and by its unique substrate recognition and Golgi localization (6). PKC-related kinases (PRK) lack the C1 domain and do not respond to DAG or phorbol esters. Phosphatidylinositol lipids activate PRKs, and small Rho-family GTPases bind to the homology region 1 (HR1) to regulate PRK kinase activity (7).

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Western Blotting

Background: ApoAI (Apolipoprotein A1) is a major component of high density lipoprotein (HDL, the “good cholesterol”) in plasma. It is produced in the liver and small intestine. ApoA1 is a cofactor for lecithin cholesterolacyltransferase (LCAT) that is responsible for the formation of plasma cholesteryl esters and promotes cholesterol efflux from tissues to the liver for excretion. Defects in ApoA1 are associated with high density lipoprotein deficiency (HDLD) and systemic non-neuropathic amyloidosis (1-3).

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Western Blotting

Background: Transforming growth factor-β (TGF-β) superfamily members are critical regulators of cell proliferation and differentiation, developmental patterning and morphogenesis, and disease pathogenesis (1-4). TGF-β elicits signaling through three cell surface receptors: type I (RI), type II (RII), and type III (RIII). Type I and type II receptors are serine/threonine kinases that form a heteromeric complex. In response to ligand binding, the type II receptors form a stable complex with the type I receptors allowing phosphorylation and activation of type I receptor kinases (5). The type III receptor, also known as betaglycan, is a transmembrane proteoglycan with a large extracellular domain that binds TGF-β with high affinity but lacks a cytoplasmic signaling domain (6,7). Expression of the type III receptor can regulate TGF-β signaling through presentation of the ligand to the signaling complex. The only known direct TGF-β signaling effectors are the Smad family proteins, which transduce signals from the cell surface directly to the nucleus to regulate target gene transcription (8,9).

$293
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Flow Cytometry, Western Blotting

Background: Interleukin-10 (IL-10) is an anti-inflammatory cytokine that is produced by T cells, NK cells, and macrophages (1,2). IL-10 initiates signal transduction by binding to a cell surface receptor complex consisting of IL-10 RI and IL-10 RII (1), leading to the activation of Jak1 and Tyk2 and phosphorylation of Stat3 (1,3). The anti-inflammatory activity of IL-10 is due to its ability to block signaling through other cytokine receptors, notably IFN-γ receptor, by upregulating expression of SOCS1 (1,3). In addition, IL-10 promotes T cell tolerance by inhibiting tyrosine phosphorylation of CD28 (4,5). IL-10 is an important negative regulator of the immune response, which allows for maintenance of pregnancy (1). In contrast, increased IL-10 levels contribute to persistent Leishmania major infections (6).

$305
50 tests
100 µl
This Cell Signaling Technology antibody is conjugated to phycoerythrin (PE) and tested in-house for direct flow cytometry analysis in human cells. This antibody is expected to exhibit the same species cross-reactivity as the unconjugated PKCα (D7E6E) Rabbit mAb #59754.
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Flow Cytometry

Background: Activation of protein kinase C (PKC) is one of the earliest events in a cascade that controls a variety of cellular responses, including secretion, gene expression, proliferation, and muscle contraction (1,2). PKC isoforms belong to three groups based on calcium dependency and activators. Classical PKCs are calcium-dependent via their C2 domains and are activated by phosphatidylserine (PS), diacylglycerol (DAG), and phorbol esters (TPA, PMA) through their cysteine-rich C1 domains. Both novel and atypical PKCs are calcium-independent, but only novel PKCs are activated by PS, DAG, and phorbol esters (3-5). Members of these three PKC groups contain a pseudo-substrate or autoinhibitory domain that binds to substrate-binding sites in the catalytic domain to prevent activation in the absence of cofactors or activators. Control of PKC activity is regulated through three distinct phosphorylation events. Phosphorylation occurs in vivo at Thr500 in the activation loop, at Thr641 through autophosphorylation, and at the carboxy-terminal hydrophobic site Ser660 (2). Atypical PKC isoforms lack hydrophobic region phosphorylation, which correlates with the presence of glutamic acid rather than the serine or threonine residues found in more typical PKC isoforms. The enzyme PDK1 or a close relative is responsible for PKC activation. A recent addition to the PKC superfamily is PKCμ (PKD), which is regulated by DAG and TPA through its C1 domain. PKD is distinguished by the presence of a PH domain and by its unique substrate recognition and Golgi localization (6). PKC-related kinases (PRK) lack the C1 domain and do not respond to DAG or phorbol esters. Phosphatidylinositol lipids activate PRKs, and small Rho-family GTPases bind to the homology region 1 (HR1) to regulate PRK kinase activity (7).

$260
100 µl
APPLICATIONS
REACTIVITY
Mouse

Application Methods: Western Blotting

Background: Interleukin-1β (IL-1β) is a proinflammatory cytokine produced predominantly by activated monocytes and epithelial cells (1). Precursor IL-1β is cleaved by caspase-1 and mature IL-1β is then secreted (1-3). Target cells include macrophages and many other cell types. Signaling by IL-1β involves IL-1β binding to IL-1 accessory protein (IL-1-AcP); the complex then binds to IL-1RI (1,2). Signaling occurs through activation of MAP kinase and NF-κB pathways (1,2). IL-1β also binds to IL-1RII, which lacks an intracellular signaling domain and thereby serves as a high affinity decoy receptor. IL-1β binding to IL-1RI is inhibited by the negative regulator, IL-1R antagonist (IL-1Ra). IL-1Ra binding to IL-1RI does not signal and serves to block IL-1β signaling. IL-1β plays critical roles in the acute phase response and sepsis (1-3).

$260
100 µl
APPLICATIONS
REACTIVITY
Mouse

Application Methods: Western Blotting

Background: Interleukin-1β (IL-1β) is a proinflammatory cytokine produced predominantly by activated monocytes and epithelial cells (1). Precursor IL-1β is cleaved by caspase-1 and mature IL-1β is then secreted (1-3). Target cells include macrophages and many other cell types. Signaling by IL-1β involves IL-1β binding to IL-1 accessory protein (IL-1-AcP); the complex then binds to IL-1RI (1,2). Signaling occurs through activation of MAP kinase and NF-κB pathways (1,2). IL-1β also binds to IL-1RII, which lacks an intracellular signaling domain and thereby serves as a high affinity decoy receptor. IL-1β binding to IL-1RI is inhibited by the negative regulator, IL-1R antagonist (IL-1Ra). IL-1Ra binding to IL-1RI does not signal and serves to block IL-1β signaling. IL-1β plays critical roles in the acute phase response and sepsis (1-3).

$260
100 µl
APPLICATIONS
REACTIVITY
Mouse

Application Methods: Flow Cytometry, Western Blotting

Background: Interleukin-1β (IL-1β), one of the major caspase-1 targets, is a multifunctional cytokine that is involved in a host of immune and proinflammatory responses (1). It is produced primarily by activated monocytes and macrophages. It signals through various adaptor proteins and kinases that lead to activation of numerous downstream targets (2-6). Human IL-1β is synthesized as a 31 kDa precursor. To gain activity, the precursor must be cleaved by caspase-1 between Asp116 and Ala117 to yield a 17 kDa mature form (7,8). Detection of the 17 kDa mature form of IL-1β is a good indicator of caspase-1 activity.

$303
100 µl
APPLICATIONS
REACTIVITY
Mouse

Application Methods: Immunoprecipitation, Western Blotting

Background: Interleukin-1β (IL-1β), one of the major caspase-1 targets, is a multifunctional cytokine that is involved in a host of immune and proinflammatory responses (1). It is produced primarily by activated monocytes and macrophages. It signals through various adaptor proteins and kinases that lead to activation of numerous downstream targets (2-6). Human IL-1β is synthesized as a 31 kDa precursor. To gain activity, the precursor must be cleaved by caspase-1 between Asp116 and Ala117 to yield a 17 kDa mature form (7,8). Detection of the 17 kDa mature form of IL-1β is a good indicator of caspase-1 activity.

$345
100 µg
Neutralizing antibodies can be used to inhibit normal biological function through their binding to biological molecules. These reagents can be used to determine the effects that a particular molecule has in biological systems. Human IL-4 Neutralizing (D20H1) Rabbit mAb has been shown to neutralize the proliferation of TF-1 cells in vitro with an ND50 in the range of 3-19 ng/ml.
REACTIVITY
Human
$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Western Blotting

Background: Acute phase response is induced by interleukin-6 (IL-6) produced by T cells, macrophages, fibroblasts, endothelial and other cells (1,2). IL-6 induces proliferation or differentiation in many cell types including B cells, thymocytes and T cells. IL-6, in concert with TGF-β, is important for developing Th17 responses. IL-6 binds to IL-6Rα and through this association induces gp130 homodimerization (1). gp130 homodimerization triggers the Jak/Stat cascade and the SHP-2/Erk MAP kinase cascade (1,3,4). IL-6 also forms a complex with an IL-6Rα splice variant that is nonmembrane-associated (3). The IL-6/soluble IL-6Rα complex can then activate the gp130 signaling pathway in cells that express gp130 but not IL-6Rα (3). Research studies have shown that IL-6, through increasing expression of proangiogenic VEGF, may also contribute to metastatic breast cancer (5).

$260
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: The adhesive glycoprotein thrombospondin-1 (THBS1, TSP1) localizes to the extracellular matrix (ECM) and mediates interactions between cells and the ECM and among cells. Thrombospondin-1 is a multi-domain, glycosylated protein that interacts with a wide variety of extracellular targets, including matrix metalloproteinases (MMPs), collagens, cell receptors, growth factors, and cytokines (1). The protein structure of THBS1 includes an amino-terminal laminin G-like domain, a von Willebrand factor-binding domain, and multiple thrombospondin (TSP) repeated sequences designated as type I, type II, or type III repeats. Each thrombospondin domain interacts with a distinct type of cell surface ligands or protein targets. The amino-terminal domain interacts with aggrecan, heparin, and integrin proteins. Type I TSP repeats interact with MMPs and CD36, while carboxy-terminal repeats bind the thrombospondin receptor CD47 (1). Through these interactions, THBS1 exerts diverse effects on different signaling pathways, such as VEGF receptor/NO signaling, TGFβ signaling, and the NF-κB pathway (2-5). Thrombospondin-1 is an important regulator of many biological processes, including cell adhesion/migration, apoptosis, angiogenesis, inflammation, vascular function, and cancer development (2-5). The activity of thrombospondin-1 is mainly regulated by extracellular proteases. The metalloproteinase ADAMTS1 cleaves thrombospondin, resulting in the release of peptides with anti-angiogenic properties. Elastase and plasmin proteases degrade the THBS1 protein and down regulate its activity (6). As THBS1 is an important protein inhibitor of angiogenesis, the development of thrombospondin-based compounds and their use in therapeutic studies may provide a beneficial approach to the treatment of cancer (7,8).

$303
100 µl
APPLICATIONS
REACTIVITY
Human, Monkey, Mouse, Rat

Application Methods: Western Blotting

Background: Activation of protein kinase C (PKC) is one of the earliest events in a cascade that controls a variety of cellular responses, including secretion, gene expression, proliferation, and muscle contraction (1,2). PKC isoforms belong to three groups based on calcium dependency and activators. Classical PKCs are calcium-dependent via their C2 domains and are activated by phosphatidylserine (PS), diacylglycerol (DAG), and phorbol esters (TPA, PMA) through their cysteine-rich C1 domains. Both novel and atypical PKCs are calcium-independent, but only novel PKCs are activated by PS, DAG, and phorbol esters (3-5). Members of these three PKC groups contain a pseudo-substrate or autoinhibitory domain that binds to substrate-binding sites in the catalytic domain to prevent activation in the absence of cofactors or activators. Control of PKC activity is regulated through three distinct phosphorylation events. Phosphorylation occurs in vivo at Thr500 in the activation loop, at Thr641 through autophosphorylation, and at the carboxy-terminal hydrophobic site Ser660 (2). Atypical PKC isoforms lack hydrophobic region phosphorylation, which correlates with the presence of glutamic acid rather than the serine or threonine residues found in more typical PKC isoforms. The enzyme PDK1 or a close relative is responsible for PKC activation. A recent addition to the PKC superfamily is PKCμ (PKD), which is regulated by DAG and TPA through its C1 domain. PKD is distinguished by the presence of a PH domain and by its unique substrate recognition and Golgi localization (6). PKC-related kinases (PRK) lack the C1 domain and do not respond to DAG or phorbol esters. Phosphatidylinositol lipids activate PRKs, and small Rho-family GTPases bind to the homology region 1 (HR1) to regulate PRK kinase activity (7).

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Western Blotting

Background: The receptor for advanced glycation end products (RAGE) is member of the immunoglobulin (Ig) superfamily. It can be expressed as full-length, membrane-bound RAGE isoform 1 or as a secreted sRAGE protein that lacks a transmembrane domain (1). RAGE is detected during early developmental stages and in the lung under normal physiological conditions (2) and is upregulated at sites of inflammation (3). Advanced glycation end products (AGEs) and a variety of other ligands interact with this receptor (1). Ligand binding activates full-length RAGE and initiates downstream signaling pathways that include activation of NF-κB, which leads to production of pro-inflammatory cytokines and inflammation (4). Activation of these pathways has been implicated in various disease states including Alzheimer disease, diabetes, arthritis, and atherosclerosis (4). Soluble RAGE can competitively bind RAGE ligands in the extracellular environment, which prevents ligand interaction with full-length RAGE at the cell surface (1).

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Flow Cytometry, Western Blotting

Background: Interleukin-4 (IL-4) is a cytokine secreted by activated T cells, basophils, and mast cells (1,2). While it contributes to many immunomodulatory responses, it is mainly recognized as the cytokine responsible for eliciting differentiation of naive T cells into Th2 lineage cells that are defined by their secretion of IL-4, IL-5, and IL-10 (3). In addition, IL-4 contributes to immunoglobulin class switching by inducing the production of IgE from B cells (4,5). IL-4 acts through the IL-4 receptor, leading to tyrosine phosphorylation and activation of the Stat6 transcription factor (6).

$303
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse, Rat

Application Methods: Immunoprecipitation, Western Blotting

Background: Activation of protein kinase C (PKC) is one of the earliest events in a cascade that controls a variety of cellular responses, including secretion, gene expression, proliferation, and muscle contraction (1,2). PKC isoforms belong to three groups based on calcium dependency and activators. Classical PKCs are calcium-dependent via their C2 domains and are activated by phosphatidylserine (PS), diacylglycerol (DAG), and phorbol esters (TPA, PMA) through their cysteine-rich C1 domains. Both novel and atypical PKCs are calcium-independent, but only novel PKCs are activated by PS, DAG, and phorbol esters (3-5). Members of these three PKC groups contain a pseudo-substrate or autoinhibitory domain that binds to substrate-binding sites in the catalytic domain to prevent activation in the absence of cofactors or activators. Control of PKC activity is regulated through three distinct phosphorylation events. Phosphorylation occurs in vivo at Thr500 in the activation loop, at Thr641 through autophosphorylation, and at the carboxy-terminal hydrophobic site Ser660 (2). Atypical PKC isoforms lack hydrophobic region phosphorylation, which correlates with the presence of glutamic acid rather than the serine or threonine residues found in more typical PKC isoforms. The enzyme PDK1 or a close relative is responsible for PKC activation. A recent addition to the PKC superfamily is PKCμ (PKD), which is regulated by DAG and TPA through its C1 domain. PKD is distinguished by the presence of a PH domain and by its unique substrate recognition and Golgi localization (6). PKC-related kinases (PRK) lack the C1 domain and do not respond to DAG or phorbol esters. Phosphatidylinositol lipids activate PRKs, and small Rho-family GTPases bind to the homology region 1 (HR1) to regulate PRK kinase activity (7).

$303
100 µl
APPLICATIONS
REACTIVITY
Human, Mouse

Application Methods: Western Blotting

Background: Vascular endothelial growth factor receptor 2 (VEGFR2, KDR, Flk-1) is a major receptor for VEGF-induced signaling in endothelial cells. Upon ligand binding, VEGFR2 undergoes autophosphorylation and becomes activated (1). Major autophosphorylation sites of VEGFR2 are located in the kinase insert domain (Tyr951/996) and in the tyrosine kinase catalytic domain (Tyr1054/1059) (2). Activation of the receptor leads to rapid recruitment of adaptor proteins, including Shc, GRB2, PI3 kinase, NCK, and the protein tyrosine phosphatases SHP-1 and SHP-2 (3). Phosphorylation at Tyr1212 provides a docking site for GRB2 binding and phospho-Tyr1175 binds the p85 subunit of PI3 kinase and PLCγ, as well as Shb (1,4,5). Signaling from VEGFR2 is necessary for the execution of VEGF-stimulated proliferation, chemotaxis and sprouting, as well as survival of cultured endothelial cells in vitro and angiogenesis in vivo (6-8).

$260
100 µl
APPLICATIONS
REACTIVITY
Human

Application Methods: Immunoprecipitation, Peptide ELISA (DELFIA), Western Blotting

Background: IFN-γ plays key roles in both the innate and adaptive immune response. IFN-γ activates the cytotoxic activity of innate immune cells, such as macrophages and NK cells (1,2). IFN-γ production by NK cells and antigen presenting cells (APCs) promotes cell-mediated adaptive immunity by inducing IFN-γ production by T lymphocytes, increasing class I and class II MHC expression, and enhancing peptide antigen presentation (1). The anti-viral activity of IFN-γ is due to its induction of PKR and other regulatory proteins. Binding of IFN-γ to the IFNGR1/IFNGR2 complex promotes dimerization of the receptor complexes to form the (IFNGR1/IFNGR2)2 -IFN-γ dimer. Binding induces a conformational change in receptor intracellular domains and signaling involves Jak1, Jak2, and Stat1 (3). The critical role of IFN-γ in amplification of immune surveillance and function is supported by increased susceptibility to pathogen infection by IFN-γ or IFNGR knockout mice and in humans with inactivating mutations in IFNGR1 or IFNGR2. IFN-γ also appears to have a role in atherosclerosis (4).