Axioobserver z1 epifluorescence microscopy system

Images were acquired under a Zeiss AxioObserver Z1 epifluorescence microscopy system with 40×/1.3 oil Plan-Apochromat, 63×/1.4 oil Plan-Aprochromat and 100×/1.4 oil Plan-Aprochromat objectives and a Hamamatsu ORCA-Flash4.0 LT Plus camera. The system is calibrated and aligned by using 200 nm-diameter TetraSpeck microspheres (T7280, ThermoFisher). Ten to fifty z-stacking images were acquired at 200 nm intervals covering a range from 2 to 10 μm by using ZEN Blue software.
Deconvolution was carried out using Huygens Professional deconvolution software (SVI) with a measured point-spread-function generated by 200 nm diameter TetraSpeck microspheres. Classical maximum likelihood estimation method with iterations of 40–60 and signal-to-noise of 20–60 was applied.

Image acquisition was carried out under a Zeiss AxioObserver Z1 epifluorescence microscopy system with 40 × /1.3 oil Plan-Apochromat, 63 × /1.4 oil Plan-Aprochromat and 100 × /1.4 oil Plan-Aprochromat objectives and a Hamamatsu ORCA-Flash4.0 LT camera. The system is calibrated and aligned by using 200 nm-diameter TetraSpeck microspheres (ThermoFisher). Z-stack images were acquired at 0.2 μm intervals covering a range from 3–8 μm by using ZEN blue software.
Deconvolution was carried out using Huygens Professional deconvolution software (SVI) with a measured point-spread-function generated by 200 nm-diameter TetraSpeck microspheres. Classical maximum likelihood estimation method with iterations of 40–60 and signal-to-noise of 20–40 was applied.