Computational protocol: Mycobacterium canettii Infection of Adipose Tissues

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Protocol publication

[…] Ziehl-Neelsen staining (RAL diagnostics, Martillac, France) was performed on infected cells (MOI 1:1 and 5:1) at day 3 p.i. Observations were acquired using the slide scanner Axio Scan.Z1 (Zeiss) (Magnification 20X) with a color Hitachi tri-CCD (1,800 × 1,200 pixels) camera. The number of intracellular mycobacteria per one cell was recorded on image acquisition by ImageJ software.M. canettii mCherry was used to localize intracellular bacteria using confocal microscopy. Infected adipocytes (MOI 10) were fixed using 4% formaldehyde at day 3 p.i. Lipid droplets were stained with BODIPY 493/503 (Sigma). The images were acquired using a confocal microscope (Zeiss, Spinning disk with a Yokogawa head and an emCCD camera 512 × 512 pixels) at magnification 63X/1.4 oil objective. [...] Statistics were performed using the SigmaPlot13 software. The distribution of the variables was assessed statistically with the Shapiro-Wilk test. The equality of the variance was checked using the Brown-Forsythe test. All data were normally distributed and were expressed using means ± standard deviation. The statistical significance was performed using the Student T-test. The Chi-square test was used to compare rates and proportions with Yates correction. A p <0.05 was regarded as significant. […]

Pipeline specifications

Software tools ImageJ, SigmaPlot
Applications Miscellaneous, Laser scanning microscopy, Microscopic phenotype analysis
Organisms Mycobacterium canettii, Mus musculus, Corynebacteriales
Diseases Tuberculosis, Tuberculosis, Pulmonary