Lsm 5 image browser software

Apoptotic cells in the skin explant tissue were detected using the in situ Cell Death Detection Kit, Fluorescein (11684795910, Roche, Penzberg, Germany) according to the manufacturer’s protocol. TUNEL assay was performed on cryosections (5 μm thick). The coverslips were mounted with VECTASHIELD mounting medium containing DAPI. The samples were observed with a laser-scanning microscope (LSM 700, Carl Zeiss, Jena, Germany) and analyzed with LSM 5 image browser software (Carl Zeiss).

AGS cells were seeded (2.2–2.5 × 10⁵) 18–24 hrs prior to co-culture on collagen-coated coverslips and prepared for infection and inoculated as described above. Three, 6, and 9 hrs post-infection, the coverslips were washed twice with PBS to remove non-adherent bacteria, fixed for 10 min (2.5% paraformaldehyde pH 7.4, 60.75 mM Na₂HPO₄, pH 9, and 14.3 mM NaH₂PO₄, pH 4.1), washed with PBS, and mounted using Vectashield (Vector Laboratories, Inc., Burlingame, CA). Twenty-four random fields per strain were imaged at 63X using differential interference contrast (DIC) microscopy on a Zeiss LSM 5 Pascal (Carl Zeiss Microscopy, LLC.; Thornwood, NY) or a Leica AF6000 (Leica Microsystems, Buffalo Grove, IL). To determine the relative length/breadth ratio, a total of 200–715 cells/strain were measured from three to six independent experiments. For each cell, maximal length was measured along the longest cellular projection and maximal width was measured at the widest portion of the cell (adapted from^{60 (link)}) using the LSM 5 Image Browser software (Carl Zeiss Microscopy, LLC.) or the Leica LAS X software (Leica Microsystems, Inc.). Images were exported as TIFF files into Adobe Photoshop CC (Adobe Systems Inc., San Jose, CA) where they were cropped and adjusted using the levels function for contrast and brightness. Data and images represent three to six independent experiments.