To analyze the cell viability in response to the metabolites, a resazurin-based assay with the reagent PrestoBlue (ThermoFisher; Walthman, MA, USA) was used (19 (link)). The assay was performed according to the manufacture's protocol. Briefly, cells were seeded with a density of 10,000 cells/well in 96-well-plates (Corning 3610; Corning, NY, USA). Cells were treated with different concentrations of the metabolites (1 μM−1 mM) for 24 h at 37°C (with 5% CO2). After the treatment, cells were incubated with media containing 1× PrestoBlue for 1 h at 37°C (with 5% CO2) and fluorescence intensity was measured at Ex/Em 560/590 nm (Tecan infinite M200; Tecan Group Ltd., Männedorf, Switzerland).
Prestoblue
Manufactured by Tecan
Sourced in Switzerland
PrestoBlue is a cell viability reagent that can be used to measure the proliferation and cytotoxicity of cells in culture. It works by detecting the metabolic activity of cells, which is proportional to the number of viable cells.
Automatically generated - may contain errors
Lab products found in correlation
Prestoblue, by Thermo Fisher Scientific (10 mentions)
Infinite m200, by Tecan (2 mentions)
Prestoblue hs, by Thermo Fisher Scientific (2 mentions)
S8068, by Selleck Chemicals (2 mentions)
Infinite m200 photometer, by Tecan (1 mentions)
Prestoblue reagent, by Thermo Fisher Scientific (1 mentions)
Infinite f200, by Tecan (1 mentions)
Sunrise plate reader, by Tecan (1 mentions)
Infinite 200, by Tecan (1 mentions)
Prism 9, by GraphPad (1 mentions)
Spark multimode microplate reader, by Tecan (1 mentions)
Cck 8, by Thermo Fisher Scientific (1 mentions)
Spark 10m, by Tecan (1 mentions)
Cyquant assay, by Thermo Fisher Scientific (1 mentions)
Infinite m200 pro, by Tecan (1 mentions)
Infinite f500, by Tecan (1 mentions)
48 well plate, by Corning (1 mentions)
15 protocols using prestoblue
1
Cell Viability Assay with Metabolites
2
Curcumin Effects on Rhabdomyosarcoma Cells
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Cells were seeded into 96-well plates at a density of 5000 cells/well, in 100 µL of cell culture medium, and incubated for 24 h to allow cell adherence. To test the effects of curcumin, growth media was exchanged, and the rhabdomyosarcoma cell lines were cultured for 24, 48, or 72 h, either in the presence of the vehicle (DMSO 0.1%) or increasing concentrations of curcumin (0.8–50 µM). Cell growth was determined using resazurin-based PrestoBlue reagent (Invitrogen, Monza, Italy), according to the manufacturer’s instructions. Briefly, the PrestoBlue solution (10 µL) was added to each well, and the plates were then incubated for an additional 2 h. The plates were then directly read on an Infinite M200 photometer (Tecan Group Ltd., Mannedorf, Switzerland) at a wavelength of 600 nm.
3
Helichrysum Infusion Cytotoxicity Assay
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Cell viability after exposure to Helichrysum infusions was determined using PrestoBlue™ reagent (Invitrogen™, Carlsbad, CA, USA) according to the manufacturer’s instructions. Cells were treated with infusions diluted in cell culture media for 24 h. PrestoBlue™ was added and after 30 min, fluorescence was measured on a spectrophotometer Infinite F200 (Tecan Group Ltd., Zürich, Switzerland) at excitation/emission (ex/em) of 535/595 nm.
4
PrestoBlue Cell Viability Assay
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Cells were seeded in 24-well plates (30,000 cells per well) and allowed to attach. One day later, the cells were placed in growth media at 42 °C for 1 h, followed by an incubation at 37 °C for 2 h to recover. Cells were incubated for 1 h with growth media containing 10% PrestoBlue (Invitrogen). The reduction of PrestoBlue reagent was measured on a Tecan Sunrise Plate Reader with the following parameters: bottom-read fluorescence (excitation = 560 nm, emission = 590 nm).
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5
Cell Viability Assay for Mesenchymal Stem Cells
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Before performing the cell experiments, all scaffolds were sterilized by immersion in 75% EtOH and exposure to UV light for 20 min. The human marrow mesenchymal stem cells (hMSCs) were obtained from Sciencell Research Laboratories (Sciencell, Carlsbad, CA, USA) and grown in commercial mesenchymal stem cell medium (Sciencell) at passage 3–6. After different culturing times, cell viability was determined by the PrestoBlue® (Thermo Fisher Scientific Inc., Waltham, MA, USA) assay. At the end of the culture period, the medium was discarded and the wells were washed twice with cold PBS. Each well was filled with a 1:9 ratio of PrestoBlue® in fresh DMEM and incubated at 37 °C for 1.5 h. The solution in each well was then transferred to a new 96-well plate and Tecan Infinite 200® PRO microplate reader (Tecan, Männedorf, Switzerland) was used at 570 nm with a reference wavelength of 600 nm. Cells cultured on tissue culture plates without materials were used as a control (Ctl). The results were obtained in triplicate from three separate experiments in terms of optical density (OD).
6
Ex vivo drug susceptibility in human macrophages
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Drug susceptibility ex vivo was determined in differentiated human THP-1 macrophages (ATCC TIB-202). THP-1 monocytes (1X105 cells/well) were differentiated overnight using 100 ng/mL PMA and seeded in a 96 well-plate. The next day, differentiated macrophages were infected with a mid-logarithmic phase culture of M. tb H37Rv (OD 0.4–0.8) at an MOI 5. Infection was allowed for 5h. Extracellular bacteria were removed by washing cells twice with pre-warmed (37°C) PBS 1X. RPMI media containing two-fold serial dilutions of compounds was added to the infected cells and incubated for 48h at 37°C with 5% CO2. THP-1 viability was determined using PrestoBlue (ThermoFisher). Cells were incubated for 1h in the presence of PrestoBlue and fluorescence monitored by a TECAN F420. The percentage of cell viability was determined by subtracting values for non-treated infected cells from the non-infected cells, and results were plotted using GraphPad Prism version 7. Experiments were performed in two biological replicates.
7
Metabolic Activity Measurement via PrestoBlue
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To assess viability, metabolic activity was measured on days 1, 8 and 15 using PrestoBlue® (ThermoFisher Scientific, United Kingdom). Briefly, culture media was replaced with 200 µL of PrestoBlue® working solution (10% PrestoBlue® in BM) and incubated for 1 h. The solution was then transferred to a black 96-well plate and read at λex: 560 nm, λem: 590 nm in a plate reader (Tecan Infinite 200, Switzerland), where fluorescence correlates with metabolic activity, and fresh OIM was added to the discs. Each group consisted of five samples (n = 5).
8
Hemocyte Viability Assay with PrestoBlue
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A total of 105 hemocytes/well were placed in a polystyrene 96-well flat bottom microtiter plate (Corning Costar) with 200 µL of Grace’s supplemented insect medium. Once the cells had attached to the well surface, the media was removed, and a solution containing 10 µL of PrestoBlue (Invitrogen) and 90 µL of media was added to each well. After a 2-h incubation, the fluorescence was measured according to the PrestoBlue Protocol in a Spark multimode microplate reader (TECAN). The gain was adjusted to the optimal value of 65. The Z-position was established as 30,000 μm, and the integration time was set to 40 μs. This procedure was performed on different days to measure cell viability over time. The results were plotted with GraphPad Prism 9.0 software (San Diego, CA, USA).
9
Viability Assay of Cultured Cells
10
Quantifying Cell Proliferation with CCK8/PrestoBlue
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Cell proliferation was determined using cell counting kit 8 (CCK8; Dojindo or PrestoBlue; Invitrogen) according to the manufacturer’s protocol. Briefly, 5 × 104 of each stable cell lines were plated in 96-well plates in DMEM medium supplemented with 1% heat-inactivated fetal bovine serum (FBS). 10 µl of substract solution were added daily to each well, followed by incubation for 4 h. The cell viability in each well was determined by reading the optical density at 450 nm for CCK8 or by measuring the fluorescence intensity (RFU) with PrestoBlue at 560/590 nm on a TECAN SPARK 10 M (TECAN).
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