Cell cycle analysis was performed as previously described [34 (link)]. NB cells were seeded into six-well plates to adhere overnight and treated with GNE987 at the indicated concentrations for another 24 h. The cells were collected after being washed by cold PBS, then fixed in 70% ethanol overnight and punched by 0.5% TritonX-100. Then the samples were treated with 25 µg/ml RNase A (#7013, CST, MA, USA) and 1.5 µmol/l propidium iodide (PI) (P4170, Sigma, Darmstadt, Germany) following the manufacturer’s instructions. Finally, samples were detected by Beckman Gallios™ Flow Cytometer (Immunotech Beckman Coulter, CA, USA) and analyzed using MultiCycle AV DNA analysis software (version 328, Verity Software House).
Beckman gallios flow cytometer
Manufactured by Beckman Coulter
Sourced in United States, Germany
The Beckman Gallios™ Flow Cytometer is a multi-color flow cytometry system designed for high-performance cell analysis. It utilizes laser-based technology to detect and analyze the physical and fluorescent characteristics of cells or particles suspended in a fluid stream.
Automatically generated - may contain errors
Lab products found in correlation
Multicycle av dna analysis software, by Verity Software House (6 mentions)
Annexin 5 fitc apoptosis detection kit, by Keygen Biotech (2 mentions)
Anti mouse cd3 efluor450, by Thermo Fisher Scientific (2 mentions)
Fetal bovine serum (fbs), by Thermo Fisher Scientific (1 mentions)
Kaluza analysis 2, by Beckman Coulter (1 mentions)
Fitc conjugated anti human cd14, by BD (1 mentions)
Fitc conjugated anti human cd16, by BioLegend (1 mentions)
Fitc conjugated mouse igg1, by BioLegend (1 mentions)
Annexin 5 fitc apoptosis kit, by BD (1 mentions)
Annexin 5 fitc antibody, by BD (1 mentions)
Kaluza software, by Beckman Coulter (1 mentions)
Pe conjugated cd11c, by Thermo Fisher Scientific (1 mentions)
Pi rnase staining buffer, by BD (1 mentions)
Fitc annexin 5 apoptosis detection kit, by BD (1 mentions)
Fcs express software, by Beckman Coulter (1 mentions)
Kaluza 1, by Beckman Coulter (1 mentions)
Kaluza version 1, by Beckman Coulter (1 mentions)
Beckman gallios, by Beckman Coulter (1 mentions)
Model 680, by Bio-Rad (1 mentions)
Propidium iodide, by Merck Group (1 mentions)
32 protocols using beckman gallios flow cytometer
1
Cell Cycle Analysis of NB Cells
2
Apoptosis Assessment by Annexin-V/PI Staining
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Annexin-V/PI staining was performed to evaluate cell apoptotic rates. The BGC-823 cells were cultured in a 6-well plate (1 × 106 cells/well) and exposed to indicated conditioned medium or 10 μg/mL 5-FU for 24 h. Subsequently, the cells were resuspended in the Annexin-V-FITC and PI binding buffer (Beyotime, Shanghai, China) for a quarter of an hour with no light exposure. Thus, the stained cells could be detected by Beckman Gallios Flow Cytometer (Beckman Coulter Inc., Brea, CA, USA). In addition, 488 nm was the excitation wavelength of FITC and PI, while the emission wavelengths of FITC and PI were 530 nm and 630 nm, respectively.
3
Quantifying Cell Apoptosis in Leukemia
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Cell apoptosis was analyzed as previously described [33 (link)]. Briefly, NB4, MV4-11, Kasumi, and THP-1 cells were treated with dBET1 at indicated concentrations. After 24 h incubation, cells were harvested and washed with cold 1 × PBS buffer. Cells were then suspended in the 1× binding buffer and stained with FITC-Annexin V antibody and PI solution according to the manual of the FITC-Annexin V apoptosis kit (cat. 556420; BD Biosciences, Franklin Lakes, NJ, United States). Cell apoptosis was analyzed by Beckman Gallios™ Flow Cytometer (Beckman, Krefeld, Germany).
4
Cell Cycle Analysis of Gastric Cancer
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Gastric cancer cells were collected and centrifugated at 180g for 4 min, suspended in cold 70% ethanol overnight, washed with phosphate-buffered saline (PBS), and incubated for 1 h with light-free at room temperature after adding propidium iodide (Cat. P4170, Sigma). The cell cycles were tested by a Beckman Gallios™ Flow Cytometer (Beckman).
5
Apoptosis Assay in Cell Culture
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Reagents and instruments used in the study were as follows: Dulbecco's modified Eagle's medium nutrient mixture F-12, trypsin, paraformaldehyde (Sigma, Germany); fetal bovine serum (Hyclone, USA); TUNEL apoptosis assay kits (Roche, USA); IX7r0-142 inverted optical microscope (Olympus, Japan); micropipetter (Eppendorf, Germany); medical blue light tube (Yingze, Tianjin); TES1330A luminance meter (Taiguang, Taipei); high speed refrigerated table top centrifuge (5804R, Eppendorf, Germany); Beckman Gallios flow cytometer (Beckman, USA).
6
Annexin-V-FITC Apoptosis Assay
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Apoptosis assays were performed using the Annexin-V-FITC Apoptosis Detection Kit (Nanjing KeyGen Biotech Co., Ltd., Nanjing, China) according to the manufacturer's instructions. Early apoptotic cells were defined as annexin-V-positive or propidium iodide-negative cells. Analyses were performed using a Beckman Gallios Flow Cytometer (Beckman Coulter, Brea, CA, USA).
7
Monocyte Subsets Identification and Analysis
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The enriched monocytes were thawed and suspended in PBS supplemented with 10% fetal bovine serum (Gibco, Invitrogen), and then stained with phycoerythrin [PE]-conjugated anti-human CD14 and fluorescein isothiocyanate [FITC]-conjugated anti-human CD16 or their isotype controls which were PE and FITC-conjugated mouse IgG1, respectively (κ isotype control; BioLegend), for identifying monocytes before scRNA-seq. Fluorescence intensity was examined by a Beckman Gallios Flow Cytometer, and analyzed by flow cytometry software Kaluza analysis 2.0 (Beckman Coulter Life Sciences). For the validation of SELL+ CM in KD infants, the mononuclear cells were thawed, and stained with FITC conjugated anti-human CD14 (BD Pharmingen™), PE-Cy7 conjugated anti-human CD16 (BD Pharmingen™), PE-conjugated SELL (BioLegend), or their isotype controls (BioLegend). Fluorescence intensity was achieved on a Beckman Gallios Flow Cytometer, and analyzed by flow cytometry software FLOWJo (BD Pharmingen™). Student’s t test was used to compare the ratio of SELL+CM to CM between the healthy and KD infants.
8
Cell Cycle Analysis of AML Cells
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At 24 hours after adding different concentrations of GNE-987 to the AML cell line, the cell line was trypsinized, washed, and fixed in 70% ethanol at 4°C overnight. Then, the cells were washed with cold PBS, resuspended in 0.5 ml of PI/RNase staining fermentation broth (cat. No. 550825; BD Pharmingen™, San Diego, CA, USA), and then incubated at room temperature for 15 min. Flow cytometry was performed using the Beckman Gallios™ Flow Cytometer (Beckman, Krefeld, Germany), and the cell cycle was analyzed using Multicycle AV DNA analysis software (Verity Software House, Topsham, ME, USA).
9
Apoptosis Analysis of GNE-987 Treated Cells
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Different concentrations of GNE-987 were added to the cell line and collected after 24 h, centrifuged at 2000 rpm for 3 min. Then, the supernatant was removed, and the remains were washed once with cold PBS and centrifuged at 4000 rpm for 3 min. After taking the supernatant, they were suspended in a 1x binding buffer. The fluorescein isothiocyanate-Annexin V apoptosis kit and PI solution staining (cat. No. 556420; BD Biosciences, Franklin Lakes, NJ, USA) were used as per the manufacturer's instructions. The cell counting method was adopted to analyze cell apoptosis (Beckman Gallios™ Flow Cytometer; Beckman).
10
Cell Cycle Analysis by Flow Cytometry
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Cells (1×106 cells/well) were synchronized by culturing them in serum-free media for 24 h. Subsequently, cells were grown in regular medium for 24 h, washed, trypsinized and fixed with 70% ethanol overnight at 4°C. After fixation, cells were transparented with 0.5% Triton X-100 for 10 min. After that, cells were washed, stained with staining solution containing 1.5 µmol/l propidium iodide (PI) (P4170; Sigma-Aldrich, St. Louis, MO, USA) and 25 µg/ml RNase A. Then, samples were analyzed by flow cytometry on a Beckman Gallios™ Flow Cytometer (Beckman, Krefeld, Germany). The percentage of cell population in various phases of the cell cycle was calculated using MultiCycle AV DNA analysis software (Verity Software House, Topsham, ME, USA).
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