NOTCH1 Antibody

SKU: orb750485

Description

NOTCH1 antibody

Images & Validation

• Tested Applications: DOT, ELISA, IHC, WB
• Dilution Range: ELISA: 1:20,000 - 1:60,000, IHC: 1:200, WB: 1:500- 1:2,000
• Reactivity: Human, Mouse
• Application Notes:
  Anti-Notch 1 has been tested by ELISA, dot blot, western blot and immunohistochemistry. An 80 kDa band corresponding to Notch 1 was observed at a 1:500 dilution. Specific conditions for reactivity should be optimized by the end user.

Key Properties

• Antibody Type: Primary Antibody
• Host: Rabbit
• Clonality: Polyclonal
• Isotype: Antiserum
• Immunogen: This whole rabbit serum was prepared by repeated immunizations with a synthetic peptide corresponding to amino acid residues of human Notch 1 located near the N-terminal sequence of the cleaved N intracellular domain (NICD).
• Target: NOTCH1
• Purity: This antiserum is directed against human NOTCH 1. Based on the immunogen sequence, we expect this antibody to react as well with mouse and rat NOTCH 1 (100% sequence homology). This antibody reacts with mouse Notch constructs present in lysates of HEK 293 cells. Only the cleaved intracellular (activated) form (NICD) is detected. No reactivity is detected against mouse N2, N3 or N4. The immunogen epitope is only exposed after gamma secretase cleavage and is not accessible in the uncleaved form.
• Conjugation: Unconjugated

Storage & Handling

• Storage: Store vial at -20° C or below prior to opening. This vial contains a relatively low volume of reagent (25 µL). To minimize loss of volume dilute 1:10 by adding 225 µL of the buffer stated above directly to the vial. Recap, mix thoroughly and briefly centrifuge to collect the volume at the bottom of the vial. Use this intermediate dilution when calculating final dilutions as recommended below. Store the vial at -20°C or below after dilution. Avoid cycles of freezing and thawing.
• Form/Appearance: Liquid (sterile filtered)
• Buffer/Preservatives: 0.1% (w/v) Sodium Azide
• Concentration: 90 mg/mL
• Expiration Date: 12 months from date of receipt.
• Dry Ice Shipping: Please note: This product requires shipment on dry ice. A dry ice surcharge will apply.
• Disclaimer: For research use only

Alternative Names

rabbit anti-notch1 antibody, Neurogenic locus Notch homolog protein 1, hN1, Translocation-associated Notch protein TAN-1

Dot Blot of Rabbit anti-Notch 1 (Cleaved N Terminal) (Human Specific) Antibody. Antigen: Row 1 - Notch 1 Peptide (Cleaved N Terminal) Row 2 - Notch 1 (Intra) Peptide. Load: Lane 1 - 200 ng Lane 2 - 66.67 ng Lane 3 - 22.22 ng Lane 4 - 7.41 ng Lane 5 - 2.47 ng. Primary antibody: Rabbit anti-Notch 1 (Cleaved N Terminal) (Human Specific) Antibody at 1:1000 for 60 min at RT. Secondary antibody: HRP Rabbit Secondary at 1:40000 for 30 min at RT. Block: orb348637 for 1 HR at RT.

Immunohistochemistry of Rabbit anti-Notch1 antibody. Tissue: Exocrine glands of human pancreas. Fixation: FFPE. Primary antibody: Notch1 antibody at 1:200. Staining: moderate to strong membranous staining and faint to moderate cytoplasmic staining. Islets showed faint staining.

Rabbit anti-Human NOTCH 1 (Cleaved N Terminal) was used at a 1:500 dilution to detect mouse Notch 1 by Western blot. Equivalent amounts of lysates from transiently transfected 293 cells expressing recombinant myc-tagged mouse Notch constructs were electrophoresed and transferred to membrane using standard methods. A reaction with diluted primary antibody was followed by washing; reaction with a 1:10000 dilution of HRP conjugated Gt-a-Rabbit IgG (orb347654), and color development. Lane M: Mol wt markers. Lane 1: No transfection. Lane 2: N1 (mouse deleted extracellular domain)-myc. Lane 3: N1 (mouse intracellular domain)-myc. Lane 4: N2 (mouse deleted extracellular domain)-myc. Lane 5: N2 (mouse intracellular domain)-myc. Lane 6: N3 (mouse deleted extracellular domain)-myc. Lane 7: N3 (mouse intracellular domain)-myc. Lane 8: N4 (mouse deleted extracellular domain)-myc. Lane 9: N4 (mouse intracellular domain)-myc. Lane 10: N1 (mouse deleted extracellular domain)(V to G)-myc.

Systemic oestrogen signalling is mediated by EGFR and Notch. (A) Representative Western blot showing expression of cleaved (active) Notch1 (N1-ICD) following culture ± 1 nM 17β-estradiol ± 10 µm GSI. (Bi) Representative Western blot showing expression of Notch ligands in sorted MCF7 cells (left) and, where available, metastatic cells (right). (Bii) Densitometric analysis of three independent repeats of MCF7 sorting and of a single experiment for primary cells. Comparisons between population 1 (CSC enriched) and other populations are displayed. (C and D) Mammosphere formation was assessed following culture with 1 nM 17β-estradiol ± gamma secretase inhibitor (GSI) alone and in combination with gefitinib. Fold change is normalised to control, untreated cells represented as line. (E) Representative image of protein levels of ERK and phosphorylated (actived) ERK following culture for 48 hours in monolayer ± 10 µm GSI. Means plotted ± SEM, *P < 0.05, **P < 0.01, ***P < 0.001 compared to E2 treated. # P < 0.05 compared to control cells.

Western Blot of Rabbit anti-Notch1 antibody. Lane 1: MCF-7 control lysate. Lane 2: MCF-7 + 1 nM 17β-estradiol. Lane 3: MCF-7 + 10 µm gamma secretase inhibitor. Load: 35 µg per lane. Primary antibody: Notch1 antibody at 1:500 for overnight at 4°C. Secondary antibody: IRDye800™ rabbit secondary antibody at 1:10000 for 45 min at RT. Block: 5% BLOTTO overnight at 4°C. Predicted/Observed size: 80 kDa for Notch1.

Quick Database Links

Gene Symbol

NOTCH1

UniProt

P46531

RefSeq (Protein):NP_060087.3