Human Notch-1 Intracellular Domain Antibody

Name: Human Notch-1 Intracellular Domain Antibody
Brand: R&D Systems
SKU: AF3647
Rating: 4 (4 reviews)

R&D Systems, Inc. a Bio-Techne Brand

Catalog # AF3647

Citations (3)

(4)

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Species Reactivity:	Human
Immunogen:	E. coli-derived recombinant human Notch‑1 Gly2428-Lys2556 Accession # P46531
Applications:	Chromatin Immunoprecipitation, CyTOF-ready, Flow Cytometry, Immunocytochemistry/Immunofluorescence, Western Blot
Label:	Unconjugated
Source:	Polyclonal Sheep IgG
Specificity:	Detects the intracellular domain (ICD) of human Notch-1 in direct ELISAs and Western blots. In direct ELISAs and Western blots, less than 1% cross-reactivity with recombinant human (rh) Notch-2 ICD, rhNotch-3 ICD, and rhNotch-4 ICD is observed.

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Applications for Human Notch-1 Intracellular Domain Antibody

	Recommended Concentration	Sample
Western Blot	1 µg/mL	See below
Flow Cytometry	2.5 µg/10⁶ cells	U2OS human osteosarcoma cell line
Chromatin Immunoprecipitation (ChIP)	5 µg/5 x 10⁶ cells	See below
CyTOF-ready	Ready to be labeled using established conjugation methods. No BSA or other carrier proteins that could interfere with conjugation.
Immunocytochemistry	5-15 µg/mL	See below

Please Note: Optimal dilutions should be determined by each laboratory for each application.

Scientific Data Examples for Human Notch-1 Intracellular Domain Antibody

	Western Blot Detection of Human Notch-1. Western blot shows lysates of MOLT-4 human acute lymphoblastic leukemia cell line, Jurkat human acute T cell leukemia cell line, RPMI 8226 human multiple myeloma cell line, 293T human embryonic kidney cell line, and 293T human embryonic kidney cell line (1 µg per lane), transfected with full length human Notch-1. PVDF membrane was probed with 1 µg/mL of Sheep Anti-Human Notch-1 Intracellular Domain Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3647) followed by HRP-conjugated Anti-Sheep IgG Secondary Antibody (Catalog # HAF016). Specific bands were detected for Notch-1 intracellular domain (NICD) and full length Notch-1 (Notch-1 FL) at approximately 115 and 300 kDa (as indicated). This experiment was conducted under reducing conditions and using Immunoblot Buffer Group 1.
	Immunocytochemistry Notch‑1 in Saos-2 Human Cell Line. Notch-1 was detected in immersion fixed Saos-2 human osteosarcoma cell line using 10 µg/mL Sheep Anti-Human Notch-1 Intracellular Domain Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3647) for 3 hours at room temperature. Cells were stained with the NorthernLights™ 557-conjugated Anti-Sheep IgG Secondary Antibody (red; Catalog # NL010) and counterstained with DAPI (blue). View our protocol for Fluorescent ICC Staining of Cells on Coverslips.
	Chromatin Immunoprecipitation (ChIP) Detection of Notch‑1-regulated Genes by Chromatin Immunoprecipitation. Jurkat human acute T cell leukemia cell line treated with 50 ng/mL PMA and 200 ng/mL calcium ionomycin for 30 minutes was fixed using formaldehyde, resuspended in lysis buffer, and sonicated to shear chromatin. Notch-1/DNA complexes were immunoprecipitated using 5 µg Sheep Anti-Human Notch-1 Intracellular Domain Antigen Affinity-purified Polyclonal Antibody (Catalog # AF3647) or control antibody (Catalog # 5-001-A) for 15 minutes in an ultrasonic bath, followed by Biotinylated Anti-Sheep IgG Secondary Antibody (Catalog # BAF016). Immunocomplexes were captured using 50 µL of MagCellect Streptavidin Ferrofluid (Catalog # MAG999) and DNA was purified using chelating resin solution. Thec-mycpromoter was detected by standard PCR.

Formulation, Preparation and Storage

Purification	Antigen Affinity-purified
Reconstitution	Reconstitute at 0.2 mg/mL in sterile PBS. Reconstitution Buffer Available: Size / Price Qty Loading... RB01:
Formulation	Lyophilized from a 0.2 μm filtered solution in PBS with Trehalose. *Small pack size (SP) is supplied either lyophilized or as a 0.2 µm filtered solution in PBS.
Shipping	The product is shipped at ambient temperature. Upon receipt, store it immediately at the temperature recommended below. *Small pack size (SP) is shipped with polar packs. Upon receipt, store it immediately at -20 to -70 °C
Stability & Storage	Use a manual defrost freezer and avoid repeated freeze-thaw cycles. 12 months from date of receipt, -20 to -70 °C as supplied. 1 month, 2 to 8 °C under sterile conditions after reconstitution. 6 months, -20 to -70 °C under sterile conditions after reconstitution.

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Background: Notch-1

Notch-1 (so named for "notches" in fly wings; also TAN-1) is a 300 kDa member of the Notch family of glycoproteins. It is associated with gene activation in both embryo and adult. Human Notch-1 is a 2538 amino acid (aa) type I transmembrane glycoprotein. It undergoes Golgi processing to generate a heterodimer composed of a 180‑200 kDa disulfide-linked extracellular domain (aa 18‑1664) and a 120 kDa membrane-bound segment (aa 1665‑2556). Upon ligand binding, the 110 kDa segment undergoes two cleavages which generate an NICD (notch intracellular domain) (aa 1754‑2556), a nuclear transcription factor. One isoform shows a deletion of aa 248‑288. Over aa 2428‑2556, human Notch 1 is 83% and 89% aa identical to canine and mouse Notch-1, respectively.

Item	Value
Entrez Gene IDs	4851 (Human); 25496 (Rat)
Alternate Names	EC 2.1.2.11, EC 3.4.21.68, Notch (Drosophila) homolog 1 (translocation-associated), Notch homolog 1, translocation-associated, Notch-1, Notch1, TAN1, TAN1neurogenic locus notch homolog protein 1, Translocation-associated notch protein TAN-1, hN1, notch 1Notch homolog 1, translocation-associated (Drosophila)

Citations for Human Notch-1 Intracellular Domain Antibody (3)

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Citations are publications that use Bio-Techne products. Selected citations for Human Notch-1 Intracellular Domain Antibody include:

Species: Human
Sample Types: Cell Lysates
Applications: Western Blot

SC Hrstka et al. (2017), NOTCH1-dependent nitric oxide signaling deficiency in hypoplastic left heart syndrome revealed through patient-specific phenotypes detected in bioengineered cardiogenesis Stem Cells, 0(0).
PMID: 28142228
Species: Human
Sample Types: Cell Lysates
Applications: Western Blot

(2016), A dual molecular analogue tuner for dissecting protein function in mammalian cells Nat Commun, 7(0):11742.
PMID: 27230261
Gogoi D et al. (2014), Involvement of Notch in activation and effector functions of gammadelta T cells. J Immunol, 192(5):2054-62.
PMID: 24489102

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Customer Reviews for Human Notch-1 Intracellular Domain Antibody (4)

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Human Notch-1 Intracellular Domain Antibody

Name: Anonymous

Application: ICC/IF

Sample Tested: E12.5 mouse embryo fixed in 4% PFA

Species: Mouse

Verified Customer | Posted 06/23/2021

Fixed 4% PFA overnight.Blocked with 1% BSAPrimary antibody dilution - 1:20Secondary antibody - Invitrogen Alexa Fluor 488Secondary antibody dilution - 1:1000Stained on an E12.5 mouse right ventricle heart section
Human Notch-1 Intracellular Domain Antibody

Name: Anonymous

Application: WB

Sample Tested: iPS2 human induced pluripotent stem cells

Species: Human

Verified Customer | Posted 01/25/2018
Human Notch-1 Intracellular Domain Antibody

Name: Anonymous

Application: ICC/IF

Sample Tested: Bladder cancer tissue,Bladder tissue

Species: Human

Verified Customer | Posted 12/10/2017
Human Notch-1 Intracellular Domain Antibody

Name: Anonymous

Application: WB

Sample Tested: Breast cancer cells

Species: Human

Verified Customer | Posted 03/01/2017

Expression of cleaved Notch-1 (Intracellular Domain) in mammary tumor cell lines. Dilution: 1:200 in PBS with 5% BSA. Secondary Ab: anti-Sheep IgG 1:5,000.

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Product Documents for Human Notch-1 Intracellular Domain Antibody

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Human Notch-1 Intracellular Domain Antibody

R&D Systems, Inc. a Bio-Techne Brand

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Applications for Human Notch-1 Intracellular Domain Antibody

Scientific Data Examples for Human Notch-1 Intracellular Domain Antibody

Western Blot

Immunocytochemistry

Chromatin Immunoprecipitation (ChIP)

Formulation, Preparation and Storage

Plan Your Experiments

Reconstitution Calculator

Background: Notch-1

Citations for Human Notch-1 Intracellular Domain Antibody (3)

Customer Reviews for Human Notch-1 Intracellular Domain Antibody (4)

Customer Images

Human Notch-1 Intracellular Domain Antibody

Human Notch-1 Intracellular Domain Antibody

Human Notch-1 Intracellular Domain Antibody

Human Notch-1 Intracellular Domain Antibody

Product Documents for Human Notch-1 Intracellular Domain Antibody

Product Datasheets

COA

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