THP-1 cells infected with eGFP-expressing L. donovani were treated or not with 10 μM of naloxonazine, 10 μM of naloxone or 80 nM of concanamycin A for 24 h, or co-treated with either naloxonazine (10 μM) and concanamycin A (80 nM) or naloxone (10 μM) and concanamycin A (80 nM) for 24 h, then stained with 180 nM of the Lysotracker red DND-99 (Life Technologies) for 1 h at 37°C. For microscopy, cells were further stained with 500 nM of the nucleic acid stain Hoechst 33342 (Life Technologies) and images were acquired with an LSM 700 Zeiss confocal microscope. For flow cytometry, Lysotracker red DND-99 stained cells were first trypsinised with TrypLE Select (Invitrogen), washed with PBS and analysed with a BD FACSVerse flow cytometer and the BD FACSSuite software.
Lysotracker red dnd 99
Manufactured by Thermo Fisher Scientific
Sourced in United States, United Kingdom, Germany, Japan, France, China, Spain
LysoTracker Red DND-99 is a fluorescent dye that selectively stains acidic organelles, such as lysosomes, in live cells. It can be used to visualize and monitor the distribution and activity of lysosomes within the cellular environment.
Automatically generated - may contain errors
Lab products found in correlation
Hoechst 33342, by Thermo Fisher Scientific (91 mentions)
4 6 diamidino 2 phenylindole (dapi), by Thermo Fisher Scientific (40 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (38 mentions)
Mitotracker green fm, by Thermo Fisher Scientific (33 mentions)
Lsm 710 confocal microscope, by Zeiss (26 mentions)
Mitotracker red cmxros, by Thermo Fisher Scientific (22 mentions)
Lsm 700 confocal microscope, by Zeiss (19 mentions)
Lsm 780 confocal microscope, by Zeiss (19 mentions)
Bafilomycin a1, by Merck Group (16 mentions)
Penicillin streptomycin, by Thermo Fisher Scientific (15 mentions)
Lysotracker green dnd 26, by Thermo Fisher Scientific (15 mentions)
Lysosensor green dnd 189, by Thermo Fisher Scientific (15 mentions)
Dulbecco modified eagle medium (dmem), by Thermo Fisher Scientific (15 mentions)
Lysosensor yellow blue dnd 160, by Thermo Fisher Scientific (15 mentions)
Mitotracker deep red fm, by Thermo Fisher Scientific (15 mentions)
Hoechst 33342, by Merck Group (14 mentions)
4 6 diamidino 2 phenylindole (dapi), by Merck Group (13 mentions)
Mitotracker red, by Thermo Fisher Scientific (13 mentions)
Er tracker red, by Thermo Fisher Scientific (12 mentions)
Lipofectamine 2000, by Thermo Fisher Scientific (12 mentions)
961 protocols using lysotracker red dnd 99
1
Evaluating Lysosomal Activity in Leishmania-Infected Macrophages
2
Lysosome and Mitochondria Imaging
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Lysosome content per cell was measured by flow cytometry of cells stained with 50 nM LysoTracker® Red DND-99 (Life Technologies). MitoTracker® Green FM (Molecular Probes) and LysoTracker Red DND-99 were used for live cell imaging by confocal microscopy of mitochondria and lysosomes, respectively, with spot-area calculation (ImageJ) and image deconvolution (SVI, Huygens software).
3
Quantifying Endosomal Acidification Inhibition
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Inhibition of endosomal acidification was measured using LysoTracker Red DND-99 (ThermoFisher). Vero cells (ATCC) were seeded in 96-well clear CellBind plates (Sigma-Aldrich) 24 h prior to the experiment at a density of 40,000 cells/well in a 100 μL volume of complete DMEM supplemented with 10% inactive FBS and 1% of penicillin/streptomycin. On the day of the experiment, the medium was replaced with 100 μL of serum free DMEM. A volume of 0.4 μL of each compound from the Spectrum Collection library (MicroSource), consisting of 2,560 individual compounds formatted as 10 mM solutions in DMSO, was incubate with the cells at 37°C for 2 h. Following this, 0.1 μM final concentration of LysoTracker Red DND-99 (ThermoFisher) was added to each well and incubated for 30 min at 37°C. The cell medium was then replaced with 100 μL of FluoroBrite DMEM (ThermoFisher). Fluorescence at excitation/emission 574/594 nm was measured using an Envision plate reader (Perkin Elmer). Data were plotted using Prism 9 (GraphPad).
4
Lysotracker RedDND-99 Immunostaining Protocol
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The Lysotracker RedDND-99 (Life Technologies) immunostaining was performed as previously described (Yacobi-Sharon et al., 2013 (link); Yang and Yamashita, 2015 (link); Chiang et al., 2017 (link)). That is, the dissected testes were incubated in Lysotracker RedDND-99 (Life Technologies) in PBS (1:1000) for 30 min on a shaker before 4% paraformaldehyde fixation.
5
Lysosome Labeling in Testis
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The Lysotracker RedDND-99 (Life Technologies) immunostaining was performed as previously described (Yacobi-Sharon et al., 2013; (link)Yang and Yamashita, 2015; (link)Chiang et al., 2017) (link). That is, the dissected testes were incubated in Lysotracker RedDND-99 (Life Technologies) in PBS (1:1000) for 30 min on a shaker before 4% paraformaldehyde fixation.
6
Lysosome Visualization in Larval Zebrafish
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Larvae at 2 d.p.f, 5 d.p.f, and 10 d.p.f were incubated with 0.5% LysoTracker™ Red DND-99 (Invitrogen) in embryo medium for 45 min. 10 d.p.f larvae were treated from 1 d.p.f. with EM containing 0.03% phenylthiourea to prevent pigmentation. After staining, larvae were then anaesthetized, mounted in 1% low melting agarose in embryo medium and viewed using a Leica SP8 laser confocal microscope. Images were then processed with FIJI software (ImageJ).
7
Lipid Imaging Probes for Cellular Analysis
8
CD8+ T Cell Ligand Binding Assay
9
Lysosomal Activity in Podocytes
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WT or FcRn KO podocytes were treated with regular media or immune complexes for 4 hours and then stained with 50 nM Lysotracker Red DND99 (Invitrogen, cat. #L7528) at 37°C for 15 minutes and then rinsed and imaged immediately using a Zeiss 780 LSM confocal microscope. Corrected total cell fluorescence (CTCF) was measured using Image J.
10
Withaferin A Modulates Autophagic Flux in HCC Cells
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Hepatocellular carcinoma cells (Huh7, HepG2, MHCC97H and MHCC97L) were grown and cultured in DMEM (Corning, cellgro, Manassas, VA, USA; cat #10-013-CV) supplemented with 10% FBS and 1% antibiotic-antimycotic. MHCC97H and MHCC97L cells represent cells that exhibit similar malignancy but differ in metastatic potential representing high vs. low lung metastasis. All the cell lines are cultured and stored following instructions from the supplier and frozen stocks are used within low passage number. Withaferin A (WFA) was procured from Calbiochem EMD Millipore (Billerica, MA, USA). MAP1LC3B/LC3B (3868), ATG5 (12994), ATG7 (8558), BECN1 (3495), PARP1 (9532), cleaved-PARP1 (5625), and SQSTM1/p62 (5114) were obtained from Cell-Signaling Technology (Danvers, MA, USA). ACTB/β-actin (A5441) 3-Methyladenine (M9281), and Chloroquine (C6628) were purchased from Sigma-Aldrich (St. Louis, MO, USA). LysoTracker Red DND-99 (L7528) was procured from Invitrogen. Bafilomycin (11038) was purchased from Cayman Chemical (Ann Arbor, MI, USA). DQTM Green BSA assay (D12050), Earle’s Balanced Salt Solution (EBSS; 14155-063), Alexa Fluor 488 (A-11008) and Alexa Fluor 555 (A-21428) were purchased from Thermo Fisher Scientific. (Waltham, MA, USA)
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