Quantification of the rate/extent of megakaryocyte differentiation and proplatelet production in vitro. Quantitative analysis of fetal liver cell and round megakaryocyte cultures was performed in ImageJ Version 1.45r software. Images were corrected for background illumination and thresholded based contrast. (A) Cells were categorized as hematopoietic stem cells (area < 330 μm2, blue line) or round megakaryocytes of increasing size (circularity ≥ 0.4, area ≥ 330 μm2, gray bars). Objects were normalized to initial (day 0) object counts and data from each time point was averaged across 4 separate regions. (B) Cells ≥ 330 μm2 in area were categorized as round megakaryocytes (circularity ≥ 0.4, blue line) or proplatelet-megakaryocytes (circularity < .0.4, black line). Objects were normalized to initial (day 4) object counts and are expressed as total number of objects (top right) and percentage of proplatelet-producing megakaryocytes (bottom right). Data from each time point were averaged across 4 separate regions.
