Figure 3.

Flow cytometry and electron microscopy analysis of isolated resMØ-aggregates

(A) Harvesting of resMØ-aggregates by making an incision in the peritoneal wall, after injecting 8 mL PBS intraperitoneally and performing a peritoneal massage.

(B) Sedimentation of resMØ-aggregates harvested at 4 h after E. coli infection; the time after starting of the sedimentation process is indicated.

(C) Phase contrast microscopy image of resMØ-aggregates isolated at 4 h after E. coli infection, and transferred into a well of a flat-bottom 96-well plate.

(D) Flow cytometry analysis of resMØ-aggregates isolated at 4 h after E. coli infection, after immunofluorescent staining with PE-Cy7-conjugated anti-CD11b, APC-Cy7-conjugated anti-F4/80, APC-conjugated anti-MHC II, PE-conjugated anti-Ly6G, PE-conjugated anti-CD19, Pacific Blue-conjugated anti-B220 and PE-conjugated anti-CD90. Data were acquired on a Becton Dickinson LSR II flow cytometer and analyzed using FlowJo X software.

(E) WMI/confocal microscopy image of the peritoneal wall after DAPI staining, allowing the detection of resMØ-aggregates at 4 h after E. coli infection.

(F) Semithin section of the peritoneal wall at 4 h after E. coli infection, showing a resMØ-aggregate. Toluidine blue staining.

(G) Electron microscopy image of the area marked by a black lined square in (F). Figures F and G reprinted with permission from Vega-Pérez at al.¹