Short hairpin (sh)RNA lenti virus infected DU 145 and PC 3 cells at the logarithmic growth phase were trypsinized, resuspended in complete medium (RPMI 1640 + 10% FBS), counted and seeded in 96 well plates (2,000 cells/well). The next day, the plate was detected by Celigo Imaging Cytometry System (Nexcelom Bioscience) to count the cells, which was repeated once a day for 5 consecutive days. This experiment was performed independently three times.
Celigo imaging cytometry system
Manufactured by Revvity
Sourced in United States
The Celigo imaging cytometry system is a high-throughput cellular imaging platform designed for quantitative analysis of cells. It captures and analyzes images of individual cells, providing data on various cellular parameters such as cell count, size, and fluorescence intensity.
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Lab products found in correlation
Cell viability kit mtt, by Roche (1 mentions)
Cell viability kit, by Roche (1 mentions)
Cell proliferation kit 1 mtt, by Merck Group (1 mentions)
Smartpool sirna, by Horizon Discovery (1 mentions)
Antifade 4 6 diamidino 2 phenylindole dapi mounting solution, by Merck Group (1 mentions)
Alexa fluor 488 conjugated goat anti rabbit igg, by Thermo Fisher Scientific (1 mentions)
Rnaimax, by Thermo Fisher Scientific (1 mentions)
Methyl thiazolyl tetrazolium (mtt), by Roche (1 mentions)
7 protocols using celigo imaging cytometry system
1
Proliferation Kinetics of Prostate Cancer Cells
2
Cell Proliferation and Viability Assay
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Cell proliferation and viability were analyzed by the Celigo imaging cytometry system and MTT assay, respectively. The fluorescence intensity of entire wells, of transfected A549 cells was detected, and the number of cells was automatically calculated by the Celigo imaging cytometry system (Nexcelom Bioscience LLC). Cell viability was measured using a Cell Viability kit (MTT; Roche Diagnostics) solubilized in 150 µl dimethyl sulfoxide according to the manufacturer's instructions. The absorption of the solution was measured at 570 nm at various time points.
3
Quantifying Cell Proliferation Dynamics
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After infection with shRAD54B or shCtrl, the BEL-7404 and SMMC-7721 cells were seeded at a density of 1,000 cells/well in 96-well plates, and then cultured at 37°C in 5% CO2. The Celigo Imaging Cytometry System (Nexcelom Bioscience, Lawrence, MA, USA) was used to observe the fluorescence intensity and calculate the corresponding cell count.
4
Cell Growth Analysis and Colony Formation
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In our study, cell growth status was analyzed by two methods, the Celigo imaging cytometry system and MTT assays. The fluorescence intensity of cells was scanned and the number of cells was automatically calculated by Celigo imaging cytometry system (Nexcelom, Lawrence, MA, USA). Cell viability was measured with a Cell Viability Kit (MTT, Roche, Indianapolis, IN, USA) according to the manufacturer's instructions. The absorption of the solution was measured at 570 nm at various time points.
Colony formation assays were performed to assess the ability of a single cell to grow into a colony. Cells were plated at a low density (500-1000 cells/well) onto 6 well plates and observed for 2 weeks. Colonies were fixed in 4% paraformaldehyde, stained with 0.5% crystal violet staining solution and washed with PBS.
Colony formation assays were performed to assess the ability of a single cell to grow into a colony. Cells were plated at a low density (500-1000 cells/well) onto 6 well plates and observed for 2 weeks. Colonies were fixed in 4% paraformaldehyde, stained with 0.5% crystal violet staining solution and washed with PBS.
5
Transfected A549 Cells: Fluorescence and Viability
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Transfected A549 cells reached 80% confluency, the fluorescence of these cells was detected and the number of cells was automatically calculated using the Celigo Imaging Cytometry system version 2.0 (Nexcelom Bioscience LLC). The cell viability was monitored using a Cell Proliferation kit I (MTT) (Sigma-Aldrich; Merck KGaA) according to the manufacturer's instructions and 0.25% trypsin was used to dissolve the purple formazan. Cells were seeded at a density of 3×103 cells/well and cultured for 5 days. Subsequently, the optical density (OD) was measured by microplate reader under 450 nm. For each experimental condition, 5 parallel wells were assigned to each group. The experiments were performed in triplicate.
6
SARS-CoV-2 Infection Assay in Calu-3 Cells
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The siRNA transfection was conducted in 384-well plate format. 6,000 Calu-3 cells were plated and transfected with siRNA SMARTpools (Dharmacon) using RNAiMAX (Invitrogen). 48 h post-transfection, cells were infected with SARS-CoV-2 at MOI of 0.125 per well. 48 h post-infection, infected cells were fixed with 5% paraformaldehyde (PFA) for 4 h and permeabilized with 0.4% Triton X-100 for 15 min. After blocking with blocking buffer containing 3% bovine serum albumin (BSA) and 10% goat serum for 30 min, cells were incubated for overnight at 4°C with rabbit-anti-SARS-CoV-2 NP polyclonal antibodies. After three washes with phosphate-buffered saline (PBS), the cells were incubated with Alexa Fluor 488-conjugated goat-anti-rabbit IgG (Thermo Fisher Scientific, USA) for 1 h at room temperature. After three additional washes, antifade-4 6-diamidino-2-phenylindole (DAPI) mounting solution (Sigma) was added to the cells for 30 min before imaging. The fluorescence intensity of cells was scanned and the SARS-CoV-2 NP-positive cells were automatically calculated by Celigo imaging cytometry system (Nexcelom, Lawrence, MA, USA) or using the IC200 imaging system (Vala Sciences, CA, USA).
7
Cell Growth Analysis and Colony Formation
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Cell growth status was analyzed by two methods: the Celigo imaging cytometry system, and 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. The slides were seeded at a density of 104 cells to complete the cell slide for subsequent cytofluorimetric assay. The fluorescence intensity and cells were automatically calculated by the Celigo imaging cytometry system (Nexcelom, Lawrence, MA, USA). We defined cell viability by MTT (Roche, Indianapolis, IN, USA) according to the manufacturer’s instructions (absorbance was measured at 570 nm).
For colony formation assays, cells were plated at a low density (500–1,000 cells/well) onto 6-well plates and observed for 14 days. Colonies were then fixed in 4% paraformaldehyde, stained with 0.5% crystal violet staining solution, and counted.
For colony formation assays, cells were plated at a low density (500–1,000 cells/well) onto 6-well plates and observed for 14 days. Colonies were then fixed in 4% paraformaldehyde, stained with 0.5% crystal violet staining solution, and counted.
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