|H Mk||Endogenous||78||Rabbit IgG|
For western blots, incubate membrane with diluted primary antibody in 5% w/v nonfat dry milk, 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.
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 June 2016
Protocol Id: 263
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.
NO66 (D7C8E) Rabbit mAb recognizes endogenous levels of total NO66 protein.
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues near the amino terminus of human NO66 protein.
Nucleolor protein 66 (NO66), also known as Myc-associated protein with a jumonji C (JmjC) domain (MAPJD), or ribosomal oxygenase 1, belongs to a large family of JmjC-domain-containing oxygenase proteins. NO66 exhibits both ribosomal histidine hydroxylase and histone demethylase activities, and plays a key role in regulation of gene transcription, RNA processing, and translation. NO66-mediated hydroxylation of ribosomal protein L8 (Rpl8) may play a role in regulation of protein synthesis (1). NO66 also functions to repress transcription by demethylating histone H3 lys4 and lys36, two histone marks that are important for transcriptional activation (2). The interaction of NO66 with the transcription factor osterix (OSX) regulates osteoblast differentiation and bone formation through repression of OSX target genes (3,4). In embryonic stem cells, the PHF19 protein recruits NO66 along with polycomb repressor complex 2 (PRC2) to differentiation-specific target genes to repress transcription through demethylation of histone H3 lys36 and methylation of histone H3 lys27, the latter mark being associated with transcriptional repression (2). NO66 is overexpressed in non-small cell lung cancer and colorectal cancer, and is associated with poor prognosis (5,6).
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|20779S||100 µl (10 western blots)||$246.00.0|