Western blot analysis of extracts from U-2 OS cells treated with Nocodazole #2190 (10 nM, 16 hr) and released from mitosis into fresh medium for indicated times using CENP-E Antibody (upper) and α-Actinin (D6F6) XP® Rabbit mAb #6487 (lower).
Western blot analysis of extracts from HT-29 cells, untreated (-) or synchronized in mitosis by treatment with Nocodazole #2190 (10 nM, 16 hr; +), using CENP-E Antibody (upper) and α-Actinin (D6F6) XP® Rabbit mAb #6487 (lower).
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.
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 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 June 2020
Protocol Id: 263
CENP-E Antibody recognizes endogenous levels of total CENP-E protein.
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Val1897 of human CENP-E protein. Antibodies are purified by protein A and peptide affinity chromatography.
Centromere-associated protein E (CENP-E) is a kinesin-like motor protein and mitotic-checkpoint kinase BUB1B binding partner that is essential for establishing and maintaining stable attachments between mitotic chromosomes and spindle microtubules (1). CENP-E plays an important role as a motor protein in the alignment of chromosomes during prometaphase (2). Research studies indicate that CENP-E protein expression peaks in late G2 and M-phases of the cell cycle before the protein is degraded at mitotic exit (3). Additional studies show that the loss of CENP-E function results in cell cycle arrest in mitosis. Mutations in the corresponding CENPE gene can result in autosomal recessive primary microcephaly-13, a developmental disorder characterized by small head circumference, dysmorphic facial features, short stature, and delayed psychomotor development (4). Since CENP-E is essential for mitotic progression and is required for cellular proliferation, it has become an interesting target for cancer therapy (5-7).
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