Cytoplasmic Ca2+ levels in DCs were assessed using three different approaches. (1) Time-lapse fluorescence microscopy was performed using a 1:1 ratio of Fluo-4-labeled DCs and CMTPX-labeled iNKT cells that were allowed to settle onto poly-L-lysine-coated glass slides. Microscopic analysis was performed in a 37°C and 5% CO2 chamber, with images taken by a Nikon Ti-Eclipse inverted wide-field microscope every 30 s. Data analysis was carried out using NIS Elements software (Nikon) version 4.13.04. (2) DCs were labeled with Fluo-4 and mixed with a 1:1 ratio of unlabeled iNKT cells, and 2 × 105 cells were added per well of a 96-well black-walled plate (Costar) and analyzed for 30–60 min at 37°C using a Synergy HT fluorescence plate reader (BioTek Instruments). (3) DCs were labeled with Fluo-4 and FuraRed (Invitrogen) and mixed at a 1:1 ratio with unlabeled iNKT cells. The cells were spun down for 2–3 min to initiate contact, incubated at room temperature for 2 min, and then vortexed and analyzed using a BD LSRII flow cytometer.
Synergy ht fluorescence plate reader
Manufactured by Agilent Technologies
Sourced in United States
The Synergy HT fluorescence plate reader is a multi-mode microplate reader that measures fluorescence intensity. It is designed to provide accurate and reliable results for a variety of fluorescence-based assays.
Automatically generated - may contain errors
Lab products found in correlation
Fura red, by Thermo Fisher Scientific (1 mentions)
Eclipse ti wide field inverted microscope, by Nikon (1 mentions)
Nis elements software, by Nikon (1 mentions)
Lsrii flow cytometer, by BD (1 mentions)
Prestoblue assay, by Thermo Fisher Scientific (1 mentions)
Calcein am fluorescent dye, by Thermo Fisher Scientific (1 mentions)
3 protocols using synergy ht fluorescence plate reader
1
Measuring Cytoplasmic Ca2+ in DC-iNKT Interactions
2
Evaluating Cardiomyocyte Metabolic Activity
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The metabolic activity of the cells was evaluated at days 1, 4, 7 post-seeding, using a PrestoBlue assay (Life Technologies) according to instructions from the manufacturer. Briefly, 2D and 3D cultures of primary CMs were incubated in 400 μL of growth medium with 10% PrestoBlue reagent for 2 h at 37 °C. The resulting fluorescence was measured (excitation 530 nm; emission 590 nm) using a Synergy HT fluorescence plate reader (BioTek). Control wells without cells were used to determine the background for all experiments.
3
THP-1 Cell Adhesion Assay on RA FLSs
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The adhesion of THP-1 cells to control siRNA-treated or ADAM-17 siRNA-treated RA FLSs grown to confluence in 96-well plates was examined. RA FLSs were serum-starved overnight. The next day, the cells were treated with TNF-α (10 ng/ml) for 24 h. THP-1 cells were collected and labeled with calcein AM fluorescent dye (Life Technologies, 5 μM) for 30 min. After being washed twice, 1 × 105 THP-1 cells were added to each well and incubated for 30 min at room temperature. Nonadherent cells were washed away, and the fluorescence was measured using a Synergy HT fluorescence plate reader (BioTek Instruments, Winooski, VT, USA).
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