Western blot analysis of extracts from various cell lines using Notch1 (C44H11) Rabbit mAb. The transmembrane/intracellular region (NTM) is indicated.
Western blot analysis of total extract from HPB-ALL cells using Notch1 (C44H11) Rabbit mAb. The full-length (FL) and transmembrane/intracellular region (NTM) are indicated.
|MW (kDa)||120, 300|
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.
For western blots, incubate membrane with diluted primary antibody in 5% w/v BSA, 1X TBS, 0.1% Tween® 20 at 4°C with gentle shaking, overnight.
NOTE: Please refer to primary antibody datasheet or product webpage for recommended antibody dilution.
From sample preparation to detection, the reagents you need for your Western Blot are now in one convenient kit: #12957 Western Blotting Application Solutions Kit
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalent grade water.
Load 20 µl onto SDS-PAGE gel (10 cm x 10 cm).
NOTE: Volumes are for 10 cm x 10 cm (100 cm2) of membrane; for different sized membranes, adjust volumes accordingly.
* Avoid repeated exposure to skin.
posted June 2005
revised November 2013
Reprobing of an existing membrane is a convenient means to immunoblot for multiple proteins independently when only a limited amount of sample is available. It should be noted that for the best possible results a fresh blot is always recommended. Reprobing can be a valuable method but with each reprobing of a blot there is potential for increased background signal. Additionally, it is recommended that you verify the removal of the first antibody complex prior to reprobing so that signal attributed to binding of the new antibody is not leftover signal from the first immunoblotting experiment. This can be done by re-exposing the blot to ECL reagents and making sure there is no signal prior to adding the next primary antibody.
NOTE: Prepare solutions with reverse osmosis deionized (RODI) or equivalently purified water.
posted June 2005
revised October 2016
Protocol Id: 10
Notch1 (C44H11) Rabbit mAb detects intracellular epitopes between 2400 and 2500 amino acids of human Notch1. It recognizes both the full-length (~300 KDa) and the NTM region (~120 KDa), which consists of a short extracellular juxtamembrane peptide, a transmembrane sequence and the intracellular domain (NICD). The antibody cannot detect the extracellular (ligand-binding) domain of Notch1 following cleavage at the S2 site by ADAM-type metalloproteases.Species Reactivity:
HumanSpecies predicted to react based on 100% sequence homology:
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro2439 of human Notch1.
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 comprised 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).
Explore pathways + proteins related to this product.
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc.
U.S. Patent No. 7,429,487, foreign equivalents, and child patents deriving therefrom.