Western blot analysis of extracts from various cell lines using EXPAND1/MUM1 (D1Z6Y) Rabbit mAb.
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Myc/DDK-tagged full-length human EXPAND1/MUM1 (hEXPAND1/MUM1-Myc/DDK; +) protein, using EXPAND1/MUM1 (D1Z6Y) Rabbit mAb (upper) or Myc-Tag (71D10) Rabbit mAb #2278 (lower).
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
EXPAND1/MUM1 (D1Z6Y) Rabbit mAb recognizes endogenous levels of total EXPAND1/MUM1 protein.Species Reactivity:
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Arg692 of human EXPAND1/MUM1 protein.
DNA double-strand breaks (DSBs) are potentially hazardous lesions that can be induced by ionizing radiation (IR), radiomimetic chemicals, or DNA replication inhibitors. Melanoma associated antigen (mutated) 1 (MUM1, EXPAND1) is a PWWP-domain containing chromatin binding protein involved in maintaining chromatin architecture of interphase chromosomes. In response to DNA damage, EXPAND1/MUM1 accumulates at sites of DNA double strand breaks through direct interaction with DNA repair factor 53BP1 (1). Accumulation of EXPAND1/MUM1 at damaged DNA sites is thought to modify the structure of the chromatin and allow access to other DNA repair factors (2). 53BP1 activates the checkpoint kinase ATM and promotes DNA double strand break repair via nonhomologous end joining (NHEJ) repair pathway (3).
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