A real-time cell analyzer (RTCA, XceLLigence, Roche) was used to perform real-time monitoring of CD8+ T-cell cytotoxicity. The CD8+ T cells were purified through magnetic-activated cell sorting (Miltenyi Biotec) from the PBMC of patients with CHB. The target cells, PLC/PRF/5 cells and HepG2 cells, were purchased from the Institute of Cell Research in Shanghai, China, and were tested negative for mycoplasma contamination. PLC/PRF/5 cells or HepG2 cells were plated in 16-well E plates (ACEA Biosciences) at a density of 1 × 104 cells per well; CD8+ T cells were then added to the culture wells at a density of 1 × 105 cells per well after stable adherent growth. Impedance data were collected over 25 h on an RTCA multi plate system.
16 well e plate
Manufactured by Agilent Technologies
Sourced in United States
The 16-well E-plates are a lab equipment product designed for cell-based assays. The plates provide 16 individual wells for culturing and monitoring cells in real-time using the xCELLigence platform. The key function of the 16-well E-plates is to enable label-free, real-time monitoring of cellular processes.
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Lab products found in correlation
Xcelligence system, by Agilent Technologies (7 mentions)
Xcelligence rtca dp, by Agilent Technologies (3 mentions)
Rtca software 1, by Agilent Technologies (2 mentions)
Xcelligence rtca mp system, by Agilent Technologies (2 mentions)
Prism 7, by GraphPad (2 mentions)
Xcelligence real time cell analyzer, by Agilent Technologies (2 mentions)
Magnetic activated cell sorting (macs), by Miltenyi Biotec (1 mentions)
Rmil 2, by Thermo Fisher Scientific (1 mentions)
Rtca software, by Agilent Technologies (1 mentions)
Xcelligence real time cell analyzer dp, by Agilent Technologies (1 mentions)
Rtca dp analyzer, by Roche (1 mentions)
Trypan blue dye, by Bio-Rad (1 mentions)
Luna 2, by Logos Biosystems (1 mentions)
Rtca software 2, by Agilent Technologies (1 mentions)
Xcelligence real time cell analysis, by Agilent Technologies (1 mentions)
Xcelligence instrument, by Agilent Technologies (1 mentions)
Xcelligence rtca dp instrument, by Roche (1 mentions)
Xcelligence real time cell analyzer, by Roche (1 mentions)
Fc500 mpl flow cytometer, by Beckman Coulter (1 mentions)
Prism 5, by GraphPad (1 mentions)
22 protocols using 16 well e plate
1
Real-time Cytotoxicity Monitoring of CD8+ T cells
2
Liver Fluke Granulin Impacts H69 Cell Proliferation
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Proliferation of H69 and ΔhuPGRN-H69 cells in response to exposure to 100 nM recombinant liver fluke granulin [17] , [21] was quantified using the impedance-based xCELLigence real time cell analysis (RTCA) approach (ACEA Biosciences). The liver fluke granulin peptide (∼10 kDa) was concentrated using Centripep with cut-off 3 kDa (Eppendorf) and resuspended in low salt solution, Opti-MEM. The absorbance at 205 nm and concentration of liver fluke granulin was determined by using a Nanodrop 2000c spectrophotometer (ThermoFisher) [31] (link). Five thousand cells/well were seeded in 16-well E-plates (ACEA) in H69 complete media. E-plates was inserted in the xCELLigence DP platform at 37 °C, 5% CO2 and changes in impedance reflecting cell adhesion and proliferation record at intervals of 20 min for 24 h. On the following day, the medium was removed and replaced with H69 complete medium supplemented with liver fluke granulin at 100 nM. Cellular proliferation was monitored for 48 h for the wild type H69, liver fluke granulin-treated H69, and ΔhuPGRN-H69 cells with and without liver fluke granulin treatment, and displayed here as change of impedance (Cell Index) after normalizing to the signals for the wild type H69, assigned as the reference cell line (RTCA Software 1.2, ACEA) [32] (link).
3
Real-Time Monitoring of Cell Proliferation
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Cell proliferation was monitored by the xCELLigence RTCA MP System (ACEA Biosciences, San Diego, CA, USA) using 16-well E-Plates (ACEA Biosciences). The cells were seeded in triplicate at 5 × 103 cells/well in the plates. For the RTCA experiments, the cells were treated with puerarin after reaching steady growth (24 h). Impedance was measured every 15 min over 96 h and represented as the cell index by the RTCA-integrated software of the xCELLigence System. The cell index was normalized to 1 at the time point of drug administration. From this data, real-time cell growth curves were generated with GraphPad Prism 7 (GraphPad Software, La Jolla, CA, USA).
4
In Vivo Evaluation of Melanoma Immunotherapy
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Healthy male C57Bl/6 mice were divided into four groups and s/c administered in the interscapular region as follows: group 1—saline buffer (control); group 2—bone-marrow-derived DCs transfected with ML/RNA-B16 complexes (105 cells per mouse); group 3—ML/RNA-B16 lipoplexes (5 µg of RNA per mouse); group 4—CIMVs prepared using bone-marrow-derived DCs transfected with ML/RNA-B16 complexes (15 µg of vesicles (total protein counts) per mouse). Mice were sacrificed 7 days post injection, and splenocytes were isolated by density gradient centrifugation (Histopaque-1083). Splenocytes were seeded in complete IMDM medium (1 × 106 cells/mL) supplemented with 20 ng/mL of rmIL-2 (Invitrogen, USA) and melanoma B16 lysate (50 µg total protein counts per mL) and incubated under SC for 6 days to restimulate CTLs. Activated splenocytes (effector cells) were co-cultured with B16 melanoma cells (target cells) at effector-to-target ratio 20:1 in 16-well E-plates (ACEA Biosciences, USA) to estimate the cytotoxic response. Tumor cell viability was monitored for 70 h using the xCELLigence system (ACEA Biosciences, USA).
5
Real-time monitoring of cell viability
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The xCElligence system (ACEA Biosciences) provides noninvasive and label-free monitoring of cell viability and growth in real-time, based on measurement of the electrical impedance of cells adhered to an electrode on the well bottom. Increased impedance indicates that an increased number of cells is adhered to the bottom at this time [31 (link)]. HCT-116 cells were placed in 16 well E-plates (ACEA Biosciences, USA) at a concentrationof 40000 cells/ml. After 18 h, cells were treated with CL-43 in various concentrations for 20 h, and then etoposide (100 µM), doxorubicin (5 µM) or cisplatin (50 µM) were added to cells. Cell proliferation was then monitored for 48 h using the RTCA xCElligence system. Data analysis was performed using RTCA Analysis Software.
6
Real-time cell analysis of CoCl2-induced C6 cell proliferation and migration
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The xCELLigence system (ACEA Biosciences, San Diego, CA, USA) provides non-invasive and label-free monitoring of cell viability, growth, and migration in real-time based on measurement of the electrical impedance of cells adhered to an electrode on the well bottom. Elevated impedance indicates that an increased number of cells adhered to the bottom at that moment compared to the previous time point. C6 cells were placed in 16-well E-plates (4.0 × 104 cells/mL; ACEA Biosciences, San Diego, CA, USA). After 12 h, C6 cells were cultivated with CoCl2 in the presence of AEAC. Cell proliferation was then monitored for 48 h using the RTCA xCELLigence system. To estimate the migration capacity, we employed CIM plates according to the instructions of the manufacturer. Data analysis was performed using RTCA Analysis Software (RTCA Software Pro, xCELLigence Instruments, ACEA, San Diego, CA, USA).
7
Optimizing CRC Cell Proliferation
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The cell seeding density for CRC cell proliferation was optimized using colorectal adenocarcinoma cell line HT29. Briefly, HT29 cells (5 × 103 cells in 150 µl medium/well) were seeded in 16-well E-plates (ACEA Biosciences Inc, San Diego, CA, USA) according to the xCELLigence Real Time Cell Analyzer (RTCA) DP manufacturer’s instructions. The following day, HM (50–100 µg/ml) was added to the cells for 48 h of incubation. Baseline cell indices were calculated using RTCA software (ACEA Biosciences) for at least two measurements based on three replicate experiments.
8
Noninvasive Real-Time Cell Monitoring Assay
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The xCELLigence system (ACEA Biosciences) provides noninvasive and label-free monitoring of cell viability, growth, and migration in real-time, based on measurement of the electrical impedance of cells adhered to an electrode on the well bottom. Increased impedance indicates that an increased number of cells adhered to the bottom at this time [57 (link)]. Two pairs of tumor cells (scr and shHsp70), namely A549 and DLD1, were incubated with THP1 cells. After 24 h of co-cultivation, THP1 cells were removed and tumor cells were placed in 16-well E-plates (4.0 × 104 cells/mL; ACEA Biosciences, San Diego, CA, USA). Cell proliferation was then monitored for 48 h using the RTCA xCELLigence system. To estimate the migration capacity, we employed CIM plates according to the instructions of the manufacturer. Data analysis was performed using RTCA Analysis Software (RTCA Software Pro, xCELLigence Instruments, ACEA, San Diego, CA, USA).
9
Assessing Nanoparticle Impacts on HUVEC Viability
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The xCELLigence system (RTCA DP Analyzer, Roche Diagnostics, Mannheim, Germany) was used to monitor the effects of nanoparticles on HUVEC viability, essentially as described before in:41
Experiments were performed in 16-well E-plates (ACEA Bioscience, San Diego, USA), in which the impedance is measured with the help of microelectrodes localized at the bottom of the wells. For background measurements, 100 µL cell-free endothelial cell growth medium was added to each well. Afterwards, 50 µL of medium from each well was replaced with 50 µL of a cell suspension containing 1x103 HUVECs. Approximately 30 min after seeding the cells, impedance monitoring was initiated using the xCELLigence system. At 24 h after seeding, an additional 100 µL of media containing different concentrations of nanoparticles was added to the wells as follows: (a) for controls, 100 µL of pure medium without nanoparticles, and (b) for the treatment samples, 100 µL of medium containing nanoparticles at concentrations 2x higher than the required final nanoparticle concentration41 .
10
Cell Viability and Proliferation Assay
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The viability and the number of cells were evaluated on the automated cell counter LUNA-II (Logos Biosystems, South Korea) using Trypan Blue Dye (Bio-Rad Laboratories). Cell proliferation was assessed by real-time cell analysis using electrical impedance assay—xCELLigence System (ACEA Bioscience, San Diego, CA, USA). RTCA software was used to determine CI values through the measured impedance recordings. Briefly, cells were plated in 16-well E-plates (ACEA Bioscience) at a density of 20,000 cells per well in a total volume of 200 µl of complete medium and were monitored in real-time mode. The data were recorded every hour for 62 h; cell indexes were calculated using RTCA Software 1.2 (ACEA Bioscience). Cell index is a parameter reflecting the impedance of electron flow caused by adherent cells.
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