Figure 3
Correlation of Lifeact-eGFP MDCK cell monolayers grown on mesh grid with carbon + formvar film facing downwards. (A) A mesh grid was inserted into a SEM carrousel holder with already identifiable areas at low magnification. (B) Monolayer cells have grown on C + F film areas with a metal mesh on top. In grid centre, landmarks of metal mesh with small notches or cropped edges in corners are visible (white arrowheads). On an area of interest (red square), prior LCI of STM infection was performed. (C) Surface imaging was done by the InLens SE detector with EHT of 4 kV. Topographic differences are clearly visible at borders of holes, which should be omitted for correlative imaging. The growth behaviour of monolayer cells in the centre of holes was comparable to growth on the bottom of cell culture dishes regarding differentiation state and homogeneity. (D) Using the SESI detector with EHT of 6 kV, cell outlines become detectable more easily. (E) Overlay of images from InLens and SESI detectors. (F) Additional overlay with the fluorescence modality from LCI. Cell outlines in LCI modality, visible due to Lifeact-eGFP, matching cell outlines visualized by SESI detector. Scale bars, (A) 1 mm, (B) 100 µm, (C–F) 20 µm.
