Cell death was induced by harvesting logarithmic L. major cells by centrifugation at 600g for 10min and incubating cells at 107cells/mL in culture medium at 26°C with different concentrations of drugs. Miltefosine was sourced from Santa Cruz Biotechnology (USA) while other drugs came from Sigma-Aldrich Inc. (USA).
Miltefosine
Manufactured by Santa Cruz Biotechnology
Sourced in United States
Miltefosine is a lipid-based compound commonly used in laboratory research applications. It is a synthetic phospholipid analog that can be utilized as a tool compound for studying cellular membranes and signaling pathways. The core function of Miltefosine is to perturb and modulate membrane-related processes, making it a valuable reagent for various in vitro and ex vivo experiments.
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Lab products found in correlation
Curcumin, by Merck Group (4 mentions)
Amphotericin b, by Merck Group (2 mentions)
Pentamidine, by Merck Group (2 mentions)
Sephadex g 50, by Merck Group (1 mentions)
1 6 diphenyl 1 3 5 hexatriene dph, by Merck Group (1 mentions)
Eugenol, by Merck Group (1 mentions)
Calcein, by Merck Group (1 mentions)
Lh 20, by Merck Group (1 mentions)
Alamar blue, by Thermo Fisher Scientific (1 mentions)
Calcein am, by Thermo Fisher Scientific (1 mentions)
Propidium iodide, by Merck Group (1 mentions)
1 palmitoyl 2 oleoyl sn glycero 3 phosphocholine popc, by Avanti Polar Lipids (1 mentions)
Wortmannin, by Merck Group (1 mentions)
Staurosporine, by Merck Group (1 mentions)
6 protocols using miltefosine
1
Inducing Cell Death in Leishmania major
2
Lipid Membrane Composition Analysis
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Eugenol, amphotericin B, calcein, sephadex G-50, LH-20, ergosterol (Erg-), pentamidine isethionate, propidium iodide (PI), and 1,6-diphenyl-1,3,5-hexatriene (DPH) were obtained from Sigma-Aldrich (Bornem, Belgium). Miltefosine was obtained from Santa Cruz Biotechnology (Heidelberg, Germany). calcein-AM included in a live/dead Viability/Cytotoxicity Kit for mammalian cells, Alamar blue and goat anti-Guinea Pig IgG Secondary Antibody, Alexa Fluor™ 488 were obtained from Thermo Fisher Scientific (Merelbeke, Belgium). The 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphoethanolamine (POPE), l -α-phosphatidylinositol (PI—Liver, Bovine), and cholesterol (Chol—Ovine Wool) were purchased from Avanti Polar Lipids Inc. (Alabaster, AL, USA). All organic solvents used were from VWR (Leuven, Belgium).
3
Inducing Cell Death in Leishmania major
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Cell death was induced by harvesting logarithmic L. major cells by centrifugation at 600 × g for 10 min and incubating cells at 107 cells/ml in culture medium with 40 μM miltefosine (Santa Cruz Biotechnology, Dallas, TX, USA) for 24 h, 50 μM curcumin (Sigma-Aldrich, St. Louis, MO, USA) or 0.5 mM H2O2 (Sigma-Aldrich) for 5 h.
For nutrient deprivation, logarithmic L. major cells, after harvesting, were washed once with sterile PBS and incubated at 107 cells/ml in a serum-deprived medium for 4 days, possibly with 10 μM wortmannin (Sigma-Aldrich). Cell concentration was evaluated using a Thoma counting chamber.
For nutrient deprivation, logarithmic L. major cells, after harvesting, were washed once with sterile PBS and incubated at 107 cells/ml in a serum-deprived medium for 4 days, possibly with 10 μM wortmannin (Sigma-Aldrich). Cell concentration was evaluated using a Thoma counting chamber.
4
Inducing Cell Death in Leishmania major
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Cell death was induced by harvesting logarithmic L. major cells by centrifugation at 600xg for 10min and incubating cells at 107cells/mL in culture medium with 40μM miltefosine (Santa Cruz Biotechnology, Dallas, TX, USA) or 50μM curcumin (Sigma-Aldrich, Saint-Louis, MO, USA) for 24h.
For nutrient deprivation, logarithmic L. major cells, after harvesting, were washed once with sterile PBS and incubated at 107cells/mL in a serum-deprived medium. Cell concentration was evaluated using a Thoma counting chamber.
For nutrient deprivation, logarithmic L. major cells, after harvesting, were washed once with sterile PBS and incubated at 107cells/mL in a serum-deprived medium. Cell concentration was evaluated using a Thoma counting chamber.
5
Anti-leishmanial Compound Evaluation
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Logarithmic L. major cells were incubated with amphotericin B (Sigma-Aldrich, Saint-Louis, MO, USA), curcumin (Sigma-Aldrich), H2O2 (Sigma-Aldrich), miltefosine (Santa Cruz Biotechnology, Dallas, TX, USA), pentamidine (Sigma-Aldrich) or staurosporine (Sigma-Aldrich) for 24 h at 5 x 106 cells/mL for the IC50 and growth inhibition experiments, or at 107 cells/mL for all other experiments.
6
Treating Leishmania major with Curcumin, Miltefosine, and Pentamidine
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Logarithmic L. major cells were harvested by centrifugation and incubated at 106 cells/ml for the MTT assays or at 107 cells/ml for all other experiments, with 30 or 50 µM curcumin (Sigma-Aldrich, Saint-Louis, MO, USA), 40 µM miltefosine (Santa Cruz Biotechnology, Dallas, TX, USA) or 50 or 100 µM pentamidine (Sigma-Aldrich), drug concentrations previously shown as inducing L. major apoptosis [16 (link)].
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