Western blot analysis of recombinant Human Granulocyte Macrophage Colony Stimulating Factor (hGM-CSF) #8922 using GM-CSF (E8I1V) Rabbit mAb.
Western blot analysis of extracts from NK-92 cells, untreated (-) or treated (+) with TPA #4174 (80 nM, 5 hr), Ionomycin #9995 (3 μM, 5 hr), and Brefeldin A #9972 (300 ng/mL, last 4 hr of stimulation), using GM-CSF (E8I1V) Rabbit mAb (upper) or β-Actin (D6A8) Rabbit mAb #8457 (lower).
|MW (kDa)||15, 18, 22|
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 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 June 2020
Protocol Id: 10
GM-CSF (E8I1V) Rabbit mAb recognizes endogenous levels of total GM-CSF protein.
Monoclonal antibody is produced by immunizing animals with a synthetic peptide corresponding to residues surrounding Pro29 of human GM-CSF protein.
GM-CSF is produced by activated T cells, NK cells and macrophages (1,5). Target cells include granulocyte, monocyte precursors and subsets of differentiated myeloid cells (1,2,3). Many target cells require GM-CSF for survival. GM-CSF induces proliferation, is involved in hematopoietic differentiation of dendritic cells, and is a key factor in differentiation pathways leading from stem cells. GM-CSF activates effector functions of myeloid cells, thereby linking adaptive and innate immunity and in turn may boost anti-tumor immunity (4). GM-CSF receptor is composed of GM-CSFRα and the common β chain, βC, which is also utilized by IL-3 and IL-5 (1). Binding of GM-CSF initiates the Jak2, Stat5, and PI3K/Akt pathways (1).
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