Floromount g mounting medium

For fluorescence staining of SFB, purified mSFB from monocolonized mice were stained for 5 min with DAPI (0.2 mg/ml) and the membrane dye FM4-64FX (5 mg/ml) (Thermo Fisher F34653). Samples were washed twice in PBS and fixed in 4% PFA for 15 min at RT. Samples were again washed twice with PBS, spotted onto slides, mixed with Floromount-G mounting medium (Abcam ab104135), covered with a No. 1.5H cover slip and sealed with nail polish. For structured illumination microscopy acquisition, images (13 images/plane/channel) were taken on a Zeiss LSM780 Elyra PS1 microscope (Carl Zeiss, Germany) using a 63x/1.4 oil Plan Apo objective and an EMCCD Andor Ixon 887 1 K camera. Images were analyzed with ZEN and ImageJ software while the SIMcheck plugin in ImageJ was used to evaluate the acquisition and processing parameters. Brightness/contrast was applied to reduce the background signal.