The dynamics of intracellular calcium storage in biofilm cells and its subsequent secretion
(A) Intracellular calcium levels of bacterial cells (n = 150) stained with calcein-AM, a calcium-specific fluorescent dye. Cells were isolated from B. subtilis biofilm colonies (day 3 and 6) grown with calcium. Inset – a representative image of stained cells isolated from a 6-day-old biofilm. Green – calcein-AM, red – NileRed membrane stain, gray – DIC. Scale bar – 2 μm.
(B and C) G-STED images B. subtilis cells isolated from 6-day-old biofilm colonies grown with calcium. Green – calcein-AM, red – NileRed membrane stain, gray – DIC. Scale bar – 2 μm.
(D) 30-nm thick virtual slice through a CSTET 3D reconstruction. Arrows indicate calcium-rich deposits associated with a membrane from the outside of the cell. Scale bar – 400 nm.