Bone marrow derived MDSCs were separated into M-MDCSs and suppressive neutrophils by FACS, then plated in RPMI with 10% FBS in the presence of DMSO or AZD1208 for 4 hours on coverslips. Cells were stained with 40 nM Mitotracker CMX Rosamine (Thermo Fisher. MA) for 15 minutes, then fixed with 4% formaldehyde dissolved in PBS (Electron Microscopy Sciences, PA) for 20 minutes at room temperature, washed with PBS and stained with DAPI (4′,6-diamidino-2-phenylindole) (Thermo Fisher. MA). Coverslips were inverted and placed onto standard microscopy slides (Fisher Scientific, MA) with Pro-Long Diamond (Thermo Fisher. MA) as a mounting medium. Images were acquired on a 3I (Intelligent Imaging Innovations, CO) Vivo spinning disk confocal microscope using a 100X oil immersion lens. Images were cropped to isolate single cells using ImageJ program (Schneider et al., 2012 (link)). Single cells were analyzed with Mitograph (Viana et al., 2015 (link)) as previously described by Harwig et al., 2018 (link).
Standard microscopy slides
Manufactured by Thermo Fisher Scientific
Standard microscopy slides are thin, flat pieces of glass or plastic used as a support for specimens in microscopy. They provide a stable and transparent surface for mounting and observing samples under a microscope.
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Lab products found in correlation
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (4 mentions)
Phosphate buffered saline (pbs), by Thermo Fisher Scientific (4 mentions)
Mitotracker cmx rosamine, by Thermo Fisher Scientific (2 mentions)
Formaldehyde, by Electron Microscopy Sciences (2 mentions)
Prolong diamond, by Thermo Fisher Scientific (2 mentions)
2 protocols using standard microscopy slides
1
Imaging Mitochondrial Dynamics in MDSCs
2
Imaging Mitochondrial Dynamics in MDSCs
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Bone marrow derived MDSCs were separated into M-MDCSs and suppressive neutrophils by FACS, then plated in RPMI with 10% FBS in the presence of DMSO or AZD1208 for 4 hours on coverslips. Cells were stained with 40 nM Mitotracker CMX Rosamine (Thermo Fisher. MA) for 15 minutes, then fixed with 4% formaldehyde dissolved in PBS (Electron Microscopy Sciences, PA) for 20 minutes at room temperature, washed with PBS and stained with DAPI (4′,6-diamidino-2-phenylindole) (Thermo Fisher. MA). Coverslips were inverted and placed onto standard microscopy slides (Fisher Scientific, MA) with Pro-Long Diamond (Thermo Fisher. MA) as a mounting medium. Images were acquired on a 3I (Intelligent Imaging Innovations, CO) Vivo spinning disk confocal microscope using a 100X oil immersion lens. Images were cropped to isolate single cells using ImageJ program (Schneider et al., 2012 (link)). Single cells were analyzed with Mitograph (Viana et al., 2015 (link)) as previously described by Harwig et al., 2018 (link).
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