A total of 104 cells were treated with 91 and 182 μΜ for the T98 cell line, and 98 and 196 μΜ for the U87 cell line of linearol. Untreated cells were used as negative control having less than 1% of DMSO. At least three independent experiments were performed, and all samples were run in triplicates. Cells were treated with linearol at its IC50 and 2IC50 values. Flow cytometric analysis was performed 72 h post-treatment with linearol. For the DNA cell cycle, cells were treated with trypsin, centrifuged, washed with PBS twice and then incubated with PI (Propidium Iodide) working solution (50 µg/mL PI, 20 mg/mL RNase A, and 0.1% Triton X-100, Sigma-Aldrich, St. Louis, MO, USA) for 15 min at 37 °C in the dark. With the use of a flow cytometer (Omnicyt Flow Cytometer, Cytognos, Athens, Greece), the PI fluorescence of 104 individual nuclei was determined. Then, the fractions of cells in G0/G1, S, G2/M and sub-G0/G1 phase were analyzed.
Omnicyt flow cytometer
Manufactured by Cytognos
Sourced in Spain, Greece
The Omnicyt is a high-performance flow cytometer designed for advanced research and clinical applications. It utilizes advanced optical and fluidic systems to enable accurate and reliable cell analysis. The Omnicyt provides researchers with the necessary tools to conduct in-depth cellular studies, without any additional interpretation or extrapolation.
Automatically generated - may contain errors
Lab products found in correlation
Rnase a, by Merck Group (2 mentions)
Triton x 100, by Merck Group (2 mentions)
Facsdiva software, by BD (1 mentions)
Facscanto 2 flow cytometer, by BD (1 mentions)
Infinicyt 2, by Cytognos (1 mentions)
Facscanto 2 cytometer, by BD (1 mentions)
Intracellular glutathione detection assay kit, by Abcam (1 mentions)
5 protocols using omnicyt flow cytometer
1
Linearol's Cell Cycle Effects
2
Flow Cytometric Analysis of Cell Cycle
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For flow cytometry, 104 cells were treated with increased concentrations (34 and 68 µM for the T98 cell line and 30 and 60 µM for the U87 cell line) of 9″-lithospermic acid methyl ester. As a negative control, untreated cells containing less than 1% DMSO were employed. All samples were run in triplicate, and at least three separate experiments were conducted. Flow cytometric analysis was performed 72 h post-treatment with 9″-lithospermic acid methyl ester. For the DNA cell cycle, cells were treated with trypsin, centrifuged, washed with PBS twice, and then incubated with PI (Propidium Iodide) working solution (50 µg/mL PI, 20 mg/mL RNase A, and 0.1% Triton X-100, Sigma-Aldrich, St. Louis, MO, USA) for 15 min at 37 °C in the dark. With the use of a flow cytometer (Omnicyt Flow Cytometer, Cytognos, Athens, Greece), the PI fluorescence of 104 individual nuclei was determined. Subsequently, the fractions of cells in G0/G1, S, G2/M, and sub-G0/G1 phases were analyzed [18 (link)].
3
Multiparameter Flow Cytometry Analysis
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Cytometric experiments were carried out with a FACSCanto II flow cytometer (BD, Franklin Lakes, NJ, USA) equipped with an argon laser (Blue, Excitation 488 nm), a helium-neon laser (Red, Excitation 633 nm), a solid-state diode laser (Violet, Excitation 405 nm), and Omnicyt flow cytometer (Cytognos SL, Salamanca, Spain). Analyses were performed with the FACSDivaTM software (BD) and the Infinicyt 2.0 software (Cytognos SL). Approximately 10,000 cell events were acquired for each sample.
4
Absolute Cell Counting by Flow Cytometry
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Absolute cell counting was performed by using Dako CytoCountTM beads (Thermo Fisher Scientific, Waltham, MA, USA). A total of 200 µL of the sample was carefully dispensed at the bottom of the tube and 50 µL beads were added. Samples were acquired by using a FACSCanto II cytometer (BD, USA) within 60 min. Approximately 20,000 cell events were collected. Setup and calibration procedures were optimised for the absolute counting protocols [112 (link)]. Additionally, absolute counting was performed by an Omnicyt flow cytometer (Cytognos SL, Salamanca, Spain).
5
Hemolymph Cell Viability Assay
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Aliquots (200 μl) of whole hemolymph (each containing about 1–2 × 106 cells/ml) were incubated with the OlyA6/PlyB mixture (25 and 50 μg/ml) for 30 min at 16°C. Control samples were run in parallel. Total hemocyte count was carried out using an Omnicyt flow cytometer (Cytognos SL, Salamanca, Spain). Samples were pelleted by centrifugation (100 × g for 10 min) and stained with different fluorophores. Annexin V-FITC (ANX)/propidium iodide (PI) and TMRE staining was carried out as previously described (Ciacci et al., 2012 (link); Canesi et al., 2015 (link)); MitoSOX and CM-H2DCFDA staining was performed as described above for confocal scanning laser microscopy. Samples were also analysed by the Intracellular Glutathione Detection Assay Kit (Abcam) (at ex: 490, em: 520 nm). Sample acquisition and analyses were performed by means of a FACS Canto II flow cytometer and analysed with DiVa™ software collecting at least 10,000 events for each sample. Data, representing the mean ± standard deviation (SD) of at least three experiments, are expressed as Mean Fluorescence Intensities reported as percent changes with respect to controls, except for ANX/PI staining, where data are expressed as percent positive cells with respect to controls.
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