Western blot analysis of extracts from various cell types using FANCB (D9W6S) Rabbit mAb.
Western blot analysis of extracts from 293T cells, mock transfected (-) or transfected with a construct expressing Myc/DDK-tagged full-length human FANCB (FANCB-Myc/DDK; +) protein, using FANCB (D9W6S) Rabbit mAb.
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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
FANCB (D9W6S) Rabbit mAb recognizes endogenous levels of total FANCB protein.Species Reactivity:
Human, Mouse, Rat
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro280 of human FANCB protein.
Fanconi anemia (FA) is an autosomal recessive genetic disorder resulting in symptoms that include chromosomal breakage, bone marrow failure, hypersensitivity to DNA cross-linking agents (such as mitomycin C), and a predisposition to cancer (1). FANCB is an X-linked member of the Fanconi Anemia nuclear complex (FANCA, FANCB, FANCC, FANCE, FANCF, FANCG, FANCM). In response to DNA damage, the FA nuclear complex induces mono-ubiquitination of FANCD2 and FANCI (2). FANCJ/BRIP1, FANCD1/BRCA2 and FANCN/PALB2 are then recruited to sites of DNA damage along with other DNA repair proteins. FA signaling is important in maintenance of chromosome stability and control of mitosis (3).
Studies of FANCB knockout embryonic stem cells suggest a role for FANCB in the formation of Rad51 and FANCD2 foci at chromosomal sites of DNA damage (4).
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