Confocal laser scanning microscopy
This protocol is extracted from research article:
Fluorescent nanosensors reveal dynamic pH gradients during biofilm formation
NPJ Biofilms Microbiomes, Jun 17, 2021; DOI: 10.1038/s41522-021-00221-8

To generate more detailed images, a Zeiss LSM 700 compact confocal laser scanning microscope was used with a Zeiss alpha-Plan-Apochromat ×63/1.46 oil objective lens. The biofilms were imaged using a Zeiss confocal laser scanning microscope with the ×63/1.46 oil objective lens and the appropriate excitation settings for the fluorescence channels (laser: DAPI = 405 nm, CellMask = 639 nm, OG-FAM = 488 nm, TAMRA = 555 nm; 2% laser power; emission filters: DAPI = 420 nm, CellMask = SP 640 nm, OG-FAM = LP 555 nm, TAMRA = SP 640 nm). Images were recorded and 3D rendered using ZEN software and analysed using Zen blue software.

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