Cells were incubated with 200 nM MitoTracker Red CMXRos probe (Beyotime) at room temperature for 30 min to label mitochondria. Images were obtained with a confocal laser scanning microscope (Carl Zeiss).
Mitotracker red cmxros probe
Manufactured by Beyotime
Sourced in China
MitoTracker Red CMXRos is a fluorescent probe used for staining mitochondria in live cells. It selectively accumulates in active mitochondria, allowing visualization and analysis of mitochondrial morphology and function.
Automatically generated - may contain errors
Lab products found in correlation
Confocal laser scanning microscope, by Zeiss (1 mentions)
Atp assay kit, by Beyotime (1 mentions)
Confocal microscope, by Olympus (1 mentions)
Novocyte flow cytometer, by Agilent Technologies (1 mentions)
Luminometer counter, by PerkinElmer (1 mentions)
Dcfh da probe, by Beyotime (1 mentions)
Mitotracker red cmxros probe, by Thermo Fisher Scientific (1 mentions)
Hoechst 33342, by Beyotime (1 mentions)
Enspire microplate reader, by PerkinElmer (1 mentions)
5 protocols using mitotracker red cmxros probe
1
Mitochondrial Staining with MitoTracker
2
Mitochondrial Function Assays in Cardiomyocytes
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ATP was measured using an ATP Assay kit (Beyotime Biotechnology, China) as determined by the manufacturer’s instructions. The luminescence produced was measured with a luminometer counter (perkin-elmer, Waltham, MA and United States), and the concentration of ATP was calculated using an ATP standard curve.
Mitochondrial membrane and intracellular ROS were measured using the relevant assay kit (MedChemExpress,USA) as determined by the manufacturer’s instructions. Flow cytometry was performed on the NovoCyte Flow Cytometer (3,130; ACEA, China) and the data were analyzed by FlowJo software (Treestar, Ashland, OR, USA).
The active mitochondria in the primary cardiomyocytes were labeled with MitoTracker Red CMXRos probe (Beyotime Biotechnology, China) and imaged using a confocal laser-scanning microscope (Olympus + Confocal Microscope).
Mitochondrial membrane and intracellular ROS were measured using the relevant assay kit (MedChemExpress,USA) as determined by the manufacturer’s instructions. Flow cytometry was performed on the NovoCyte Flow Cytometer (3,130; ACEA, China) and the data were analyzed by FlowJo software (Treestar, Ashland, OR, USA).
The active mitochondria in the primary cardiomyocytes were labeled with MitoTracker Red CMXRos probe (Beyotime Biotechnology, China) and imaged using a confocal laser-scanning microscope (Olympus + Confocal Microscope).
3
Cellular Oxidative Injury Assessment
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Cellular oxidative injury was evaluated by detecting reactive oxygen species (ROS) generation and mitochondrial membrane potential (MMP). The generation of ROS was measured by the DCFH-DA probe (Beyotime, Cat#S0033S), and MMP was detected by the Mito-Tracker Red CMXRos probe (Beyotime, Cat#C1049) according to the manufacturers’ instruction and previous studies (Jurado-Campos et al., 2021 (link); Gu et al., 2022 (link)). Briefly, cells were cultured and treated in 24-well plates, and incubated with corresponding probes at 37°C in the dark for 30 min. After washing with PBS, the samples were examined under a fluorescence microscope.
4
Assessing Mitochondrial Membrane Potential in Parasites and Host Cells
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Extracellular parasites (1 × 105 per group) collected as described in Section “Parasites” were maintained in suspension with myrislignan (32 or 70 μg/ml) in DMEM or without any drug (positive or negative control) for 8 h at 37°C. The negative control samples were incubated for 20 min with 10 μM carbonyl cyanide 3-chlorophenylhydrazone (CCCP), which can cause ΔΨm depletion. After incubation, the tachyzoite samples were stained with the MitoTracker Red CMXRos probe (50 nM, Invitrogen, USA) for 20 min, as described previously with some modifications (Besteiro et al., 2011 (link)).
Vero cells were placed in cell culture dishes and incubated with myrislignan (70 μg/ml) or without any drug (as a positive control) in DMEM for 8 h at 37°C, stained with the MitoTracker Red CMXRos probe (250 nM) for 20 min, rinsed twice with PBS, fixed with 4% polyformaldehyde for 15 min, and washed twice with PBS to remove the polyformaldehyde, stained with Hoechst 33342 (C1027, Beyotime Institute of Biotechnology, China). The fluorescence intensity was observed by laser confocal microscopy. The relative fluorescence intensity of the cells was measured using an Enspire Microplate Reader (PerkinElmer, German).
Vero cells were placed in cell culture dishes and incubated with myrislignan (70 μg/ml) or without any drug (as a positive control) in DMEM for 8 h at 37°C, stained with the MitoTracker Red CMXRos probe (250 nM) for 20 min, rinsed twice with PBS, fixed with 4% polyformaldehyde for 15 min, and washed twice with PBS to remove the polyformaldehyde, stained with Hoechst 33342 (C1027, Beyotime Institute of Biotechnology, China). The fluorescence intensity was observed by laser confocal microscopy. The relative fluorescence intensity of the cells was measured using an Enspire Microplate Reader (PerkinElmer, German).
5
Mitochondrial Dysfunction in T. gondii
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T. gondii tachyzoites in Vero cell monolayers were incubated with DMEM with 1 or 5 μM HQNO for 8 or 24 h, washed with PBS, and coincubated with the MitoTracker Red CMXRos probe (50 nM, C1049B, Beyotime Biotechnology, China) for 20 min. The fluorescence intensity of the samples was observed by a multifunctional microplate reader (Zhang et al., 2021 ).
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