|H M R Mk||Endogenous||78||Rabbit|
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Myc/DDK-tagged full-length human NUB1L protein (hNUB1L-Myc/DDK; +), using NUB1 Antibody.Learn more about how we get our images.
Western blot analysis of extracts from various cell lines using NUB1 Antibody (upper) and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower). As expected, U-2 OS cells express low levels of endogenous NUB1 protein (3).Learn more about how we get our images.
Western blot analysis of extracts from HeLa cells, untreated (-) or treated with Human Interferon-γ (hIFN-γ) #8901 (100 ng/ml, 72hr; +), using NUB1 Antibody (upper) and GAPDH (D16H11) XP® Rabbit mAb #5174 (lower).Learn more about how we get our images.
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
Supplied in 10 mM sodium HEPES (pH 7.5), 150 mM NaCl, 100 µg/ml BSA and 50% glycerol. Store at –20°C. Do not aliquot the antibody.
NUB1 Antibody recognizes endogenous levels of total NUB1 protein. Based upon sequence alignment, this antibody is predicted to detect both NUB1 and NUB1L.
Human, Mouse, Rat, Monkey
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues near the carboxy terminus of human NUB1 protein. Antibodies are purified by protein A and peptide affinity chromatography.
Both the NEDD8 ultimate buster 1 (NUB1) and the related NUB1L isoform are interferon-inducible adaptor proteins that negatively regulate ubiquitin-like protein NEDD8 (1,2). NUB1 protein contains an amino terminal ubiquitin-like (UBL) domain and multiple carboxy terminal ubiquitin-associated (UBA) domains. The NUB1L isoform is generated by alternative splicing and contains an extra UBA domain relative to NUB1 (2). Research studies indicate that NUB1 and NUB1L non-covalently bind NEDD8 and facilitate delivery of both NEDD8 monomers and NEDD8 conjugates to the proteasome for degradation (2-5). In addition, NUB1L binds and enhances the proteasomal degradation of the FAT10 ubiquitin-like protein (6). Additional research shows that NUB1 negatively regulates cell proliferation, likely due to inhibition of NEDD8 conjugation to SCF ubiquitin ligases, which leads to inhibition of p27 and cyclin E ubiquitination (3,7). NUB1 has been identified as a putative therapeutic target in Huntington's disease as NUB1 promotes a decrease in levels of mutant HTT protein (8).
Cell Signaling Technology is a trademark of Cell Signaling Technology, Inc. XP is a registered trademark of Cell Signaling Technology, Inc. Tween is a registered trademark of ICI Americas, Inc.
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|14810S||100 µl (10 western blots)||$ 255.0|