Western blot analysis of extracts from E6.5 and E10.5 stage mouse embryos using EOMES Antibody.
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 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
EOMES Antibody detects endogenous levels of total EOMES protein.Species Reactivity:
MouseSpecies predicted to react based on 100% sequence homology:
Human, Rat, Monkey
Polyclonal antibodies are produced by immunizing animals with a synthetic peptide corresponding to amino acid sequences at the amino terminus of human EOMES. Antibodies are purified by protein A and peptide affinity chromatography.
The T-box family of transcription factors is named for their shared homology with the DNA binding domain of the mouse brachyury (T) gene product. Members of this family bind DNA and are capable of transcriptional activation. They also have evolutionarily conserved expression patterns and roles in embryonic development, primarily mesoderm development (1). EOMES, or Tbr2 (T-box brain 2), is a master regulator of mesoderm formation that is also essential for trophoblast formation, gastrulation, neurogenesis and the differentiation of certain T cell subsets. Embryos from EOMES knock-out mice die soon after implantation due to their inability to develop a trophoblast (2,3). Conditional neural knock out mice show defects in development of a specific population of neural progenators known as Intermediate Progenator Cells (IPCs) that give rise only to neurons (4,5). These cells are formed from the radial glia in the ventricular and sub-ventricular zones of the cortex. Expression of EOMES increases as cells develop from radial glia to IPCs and then decreases as IPCs progress to neurons. Recent evidence suggests that EOMES and IPCs may also play a role neurogenesis in the adult hippocampal SGZ (5). EOMES is also a key transcription factor for memory T cells and for full effector differentiation of CD8+ T cells (6). Expression of EOMES is induced in CD8+ T cells following viral infection and bacterial infection where sufficient IL-12 has been produced to elicit acute host cell response (7).
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