Revision 2

#3640

Store at -20C

Notch Isoform Antibody Sampler Kit

1 Kit

(4 x 20 microliters)

CST Logo
Orders:

877-616-CELL (2355)

[email protected]

Support:

877-678-TECH (8324)

3 Trask Lane | Danvers | Massachusetts | 01923 | USA

For Research Use Only. Not for Use in Diagnostic Procedures.

Product Includes Product # Quantity Mol. Wt Isotype/Source
Cleaved Notch1 (Val1744) (D3B8) Rabbit Monoclonal Antibody414720 µl110 kDaRabbit IgG
Notch1 (D1E11) Rabbit Monoclonal Antibody360820 µl120, 300 kDaRabbit IgG
Notch3 (D11B8) Rabbit Monoclonal Antibody527620 µl90, 270 kDaRabbit IgG
Notch2 (D76A6) Rabbit Monoclonal Antibody573220 µl110, 300 kDaRabbit
Anti-rabbit IgG, HRP-linked Antibody7074100 µlGoat

Please visit cellsignal.com for individual component applications, species cross-reactivity, dilutions, protocols, and additional product information.

Description

The Notch Isoform Antibody Sampler Kit provides an economical means to investigate Notch Signaling. The kit contains primary and secondary antibodies to perform two western mini-blots with each antibody.

Storage

Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA, 50% glycerol and less than 0.02% sodium azide. Store at –20°C. Do not aliquot the antibody.

Background

Notch proteins (Notch1-4) are a family of transmembrane receptors that play important roles in development and the determination of cell fate (1). Mature Notch receptors are processed and assembled as heterodimeric proteins, with each dimer composed of a large extracellular ligand-binding domain, a single-pass transmembrane domain, and a smaller cytoplasmic subunit (Notch intracellular domain, NICD) (2). Binding of Notch receptors to ligands of the Delta-Serrate-Lag2 (DSL) family triggers heterodimer dissociation, exposing the receptors to proteolytic cleavages; these result in release of the NICD, which translocates to the nucleus and activates transcription of downstream target genes (3,4).

Constitutively activated Notch1 signaling is associated with the majority of cases of T cell acute lymphoblastic leukemia (T-ALL). The activation is either due to mutations in Notch1 itself or in the components of ubiquitin ligase complex, namely FBW7 (5-6). Notch2 is a member of the Notch family and mutation in Notch2 is associated with Alagille syndrome (7). Notch3 is a member of the Notch family and is processed similar to Notch1 (8). It is expressed primarily in arterial smooth muscle cells (SMC). Mutations altering the number of cysteine residues in the notch3 extracellular region are associated with cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), a hereditary angiopathy leading to strokes and dementia in adults (9-11). Recent studies indicate that Notch3 is overexpressed in many types of cancer (12-14).

Trademarks and Patents

Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.

All other trademarks are the property of their respective owners. Visit cellsignal.com/trademarks for more information.

Limited Uses

Except as otherwise expressly agreed in a writing signed by a legally authorized representative of CST, the following terms apply to Products provided by CST, its affiliates or its distributors. Any Customer's terms and conditions that are in addition to, or different from, those contained herein, unless separately accepted in writing by a legally authorized representative of CST, are rejected and are of no force or effect.

Products are labeled with For Research Use Only or a similar labeling statement and have not been approved, cleared, or licensed by the FDA or other regulatory foreign or domestic entity, for any purpose. Customer shall not use any Product for any diagnostic or therapeutic purpose, or otherwise in any manner that conflicts with its labeling statement. Products sold or licensed by CST are provided for Customer as the end-user and solely for research and development uses. Any use of Product for diagnostic, prophylactic or therapeutic purposes, or any purchase of Product for resale (alone or as a component) or other commercial purpose, requires a separate license from CST. Customer shall (a) not sell, license, loan, donate or otherwise transfer or make available any Product to any third party, whether alone or in combination with other materials, or use the Products to manufacture any commercial products, (b) not copy, modify, reverse engineer, decompile, disassemble or otherwise attempt to discover the underlying structure or technology of the Products, or use the Products for the purpose of developing any products or services that would compete with CST products or services, (c) not alter or remove from the Products any trademarks, trade names, logos, patent or copyright notices or markings, (d) use the Products solely in accordance with CST Product Terms of Sale and any applicable documentation, and (e) comply with any license, terms of service or similar agreement with respect to any third party products or services used by Customer in connection with the Products.

Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Western blot analysis of total cell extract from various cell types using Notch1 (D1E11) XP® Rabbit mAb. The full-length (FL) Notch protein and the cleaved transmembrane/intracellular region (NTM) are indicated.
undefined Image 1: Notch Isoform Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using Cleaved Notch1 (Val1744) (D3B8) Rabbit mAb (upper) or Notch1 (D1E11) XP® Rabbit mAb #3608 (lower).
undefined Image 2: Notch Isoform Antibody Sampler Kit
Western blot analysis of extracts from various cell lines using Notch3 (D11B8) Rabbit mAb.
undefined Image 3: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Western blot analysis of extracts from 293 and SK-MEL-5 cells using Notch2 (D76A6) XP® Rabbit mAb. The full-length (FL) Notch2 protein and the transmembrane/intracellular region (NTM) are indicated.
undefined Image 4: Notch Isoform Antibody Sampler Kit
Confocal immunofluorescent analysis of SNB19 cells using Notch2 (D76A6) XP® Rabbit mAb (green). Blue pseudocolor = DRAQ5® #4084 (fluorescent DNA dye).
undefined Image 5: Notch Isoform Antibody Sampler Kit
Flow cytometric analysis of fixed and permeabilized SW620 cells (red, low expression) and 293 cells (blue, positive) cells using Notch2 (D76A6) XP® Rabbit mAb. Anti-rabbit IgG (H+L), F(ab')2 Fragment (Alexa Fluor® 488 Conjugate) #4412 was used as a secondary antibody.
undefined Image 6: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin-embedded human non-Hodgkin lymphoma using Notch2 (D76A6) XP® Rabbit mAb performed on the Leica BOND RX.

undefined Image 7: Notch Isoform Antibody Sampler Kit
After the primary antibody is bound to the target protein, a complex with HRP-linked secondary antibody is formed. The LumiGLO® is added and emits light during enzyme catalyzed decomposition.
undefined Image 8: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human inflammatory granulation tissue using Notch1 (D1E11) XP® Rabbit mAb.
undefined Image 9: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
CUTLL1 cells were cultured in media with γ-secretase inhibitor (1μM) for 3 days and then either harvested immediately (left panel) or washed and cultured in fresh media for 3h (right panel). Chromatin immunoprecipitations were performed with cross-linked chromatin from cells and Cleaved Notch1 (Val1744) (D3B8) Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using human HES1 promoter primers, SimpleChIP® Human HES4 Promoter Primers #7273, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
undefined Image 10: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded normal human placenta using Notch2 (D76A6) XP® Rabbit mAb performed on the Leica BOND RX.
undefined Image 11: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human breast carcinoma using Notch1 (D1E11) XP® Rabbit mAb.
undefined Image 12: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin-embedded human serous papillary carcinoma of the ovary using Notch2 (D76A6) XP® Rabbit mAb performed on the Leica BOND RX.
undefined Image 13: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded A2780 (left), Jurkat (center) and RL (right) cell pellets using Notch1 (D1E11) XP® Rabbit mAb. Both A2780 and Jurkat express Notch1, but only Jurkat cells have cleaved Notch1, while RL cells express very low or no Notch1.
undefined Image 14: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human papillary thyroid carcinoma using Notch2 (D76A6) XP® Rabbit mAb performed on the Leica BOND RX.
undefined Image 15: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin-embedded human stomach adjacent to MALT (mucosa-associated lymphoid tissue) lymphoma using Notch1 (D1E11) XP® Rabbit mAb.
undefined Image 16: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human serous papillary carcinoma of the ovary using Notch2 (D76A6) XP® Rabbit mAb (left) compared to concentration-matched Rabbit (DA1E) mAb IgG XP® Isotype Control #3900 (right).
undefined Image 17: Notch Isoform Antibody Sampler Kit
CUTLL1 cells were cultured in media with γ-secretase inhibitor (1 μM) for 3 days and then either harvested immediately (left panel) or washed and cultured in fresh media for 3 hr (right panel). Chromatin immunoprecipitations were performed with cross-linked chromatin from cells and Notch1 (D1E11) XP® Rabbit mAb or Normal Rabbit IgG #2729 using SimpleChIP® Enzymatic Chromatin IP Kit (Magnetic Beads) #9003. The enriched DNA was quantified by real-time PCR using human HES1 promoter primers, SimpleChIP® Human HES4 Promoter Primers #7273, and SimpleChIP® Human α Satellite Repeat Primers #4486. The amount of immunoprecipitated DNA in each sample is represented as signal relative to the total amount of input chromatin, which is equivalent to one.
undefined Image 18: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin-embedded human ovarian serous carcinoma using Notch2 (D76A6) XP® Rabbit mAb.
undefined Image 19: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human ductal breast carcinoma using Notch2 (D76A6) XP® Rabbit mAb.
undefined Image 20: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded 293T cell pellet (left, positive) or IMR-32 cell pellet (right, negative) using Notch2 (D76A6) XP® Rabbit mAb.
undefined Image 21: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin-embedded human colon carcinoma using Notch2 (D76A6) XP® Rabbit mAb.
undefined Image 22: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human B-cell non-Hodgkin's lymphoma using Notch2 (D76A6) XP® Rabbit mAb.
undefined Image 23: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human prostate adenocarcinoma (left) or endometrioid adenocarcinoma (right) using Notch2 (D76A6) XP® Rabbit mAb (top) or Notch2 Rabbit mAb (bottom). These two antibodies detect independent, unique epitopes on human Notch2. Both epitopes are contained within the intracellular domain and the nuclear staining represents active Notch2. The similar staining patterns obtained with both antibodies help to confirm the specificity of the staining.
undefined Image 24: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.

Revision 2

Cell Signaling Technology Logo
Immunohistochemical analysis of paraffin-embedded human placenta using Notch2 (D76A6) XP® Rabbit mAb.
undefined Image 25: Notch Isoform Antibody Sampler Kit
Immunohistochemical analysis of paraffin-embedded human testis using Notch2 (D76A6) XP® Rabbit mAb.
undefined Image 26: Notch Isoform Antibody Sampler Kit
Immunoprecipitation of Notch1 protein from Molt-4 cell extracts. Lane 1 is 10% input, lane 2 is Rabbit (DA1E) mAb IgG XP® Isotype Control #3900, and lane 3 is Notch1 (D1E11) XP® Rabbit mAb. Western blot analysis was performed using Notch1 (D1E11) XP® Rabbit mAb.
undefined Image 27: Notch Isoform Antibody Sampler Kit
Orders: 877-616-CELL (2355) [email protected] Support: 877-678-TECH (8324) [email protected] Web: cellsignal.com
For Research Use Only. Not for Use in Diagnostic Procedures.