Fig. 2

Histological analysis. The thrombus was stained for H&E, MSB, DNA, platelets, VWF, NETs (H3Cit), leukocytes and microcalcifications. H&E staining (a) was used to visualize the overall thrombus composition and organization, indicating the presence of an RBC-rich area in red (b) and a large DNA-rich area in purple (c). Feulgen staining, a specific DNA stain, confirmed the presence of extracellular DNA with few intact nuclei in this area (d, pink). MSB staining and the platelets staining revealed the presence of fibrin (e, red) and platelets (f, purple) on the surface of the thrombus, whereas VWF (G, purple) was present throughout the DNA-rich area. H3Cit staining (purple) showed the presence of extracellular H3Cit positive smears (h, purple) and leukocyte staining showed a large smear with little to no intact leukocytes present (i, purple), indicating that extracellular DNA originated from leukocytes and to some extent from neutrophils. Von Kossa staining showed abundant microcalcifications (j, black). A representative image of an isotype control for the VWF staining is shown in panel K. Scale bars are 500 μm for A and 125 μm for B-K