Human MKs express cFN isoforms. CD61⁺ MKs at 13 days of culture were separated using the immunomagnetic beads technique and total cellular RNA was extracted. (A) RT-PCR demonstrated expression of cFN (FN1 gene), the EDA⁺ FN and EDA⁻ spliced FN mRNA sequence, and the EDB⁺ FN and spliced EDB⁻ isoform. (B) MKs were permeabilized and stained in immunofluorescent dye with a polyclonal antibody against cFN (red) and (C) a monoclonal antibody against the EDA isoform (green). Scale bars = 20 μm and 10 μm, respectively. (D) cFN relocated to the MK plasma membrane upon activation with thrombin (1 U/mL; top panels) or omitting thrombin (bottom panels), as revealed by immunofluorescence with polyclonal antibody against cFN (red). Nuclei were stained with Hoechst 33288 (blue). Scale bar = 10 μm.