Western analysis of STAT3 isoform expression and phosphorylation during granulocytic differentiation.
Whole cell extracts (20 μg) of CD34+-derived cells undergoing granulocytic differentiation were separated by sodium dodecyl sulfate–polyacrylamide gel electrophoresis, blotted onto polyvinylidene fluoride membranes, and probed with antibodies against (A) STAT3, (B) STAT3 pTyr 705, and (C) STAT3 pSer 727. The 4 culture conditions evaluated are designated as 1-4 and are as follows: 1 indicates 5% O2, pH 7.25, +IL-3; 2 indicates 20% O2, pH 7.25, +IL-3; 3 indicates 5% O2, pH 7.4, +IL-3; and 4 indicates 5% O2, pH 7.25, −IL-3. The relative positions of STAT3α, β, γ, and δ are indicated on the left. The results shown are representative of 3 sets of experiments from different mobilized peripheral blood samples.
