Following isolation, 20,000 CD8+ cells were seeded in an anti-mouse CD3e-bound 96-well plate and treated with 10 µl of MDSC-derived exosomes or the same volume of PBS (control). After 48 h, 10 µl of WST-1 reagent (Alkali Scientific Inc.) was added to each well and incubated for 4 h. The absorbance of each well was measured at a wavelength of 450 nm by the Perkin Elmer Victor3 V 1420 multilabel plate reader.
Victor 3v 1420
Manufactured by PerkinElmer
Sourced in United States
The Victor 3V 1420 is a multimode microplate reader designed for various fluorescence, luminescence, and absorbance measurements. It is equipped with a xenon flash lamp and monochromator-based optics. The instrument can accommodate a wide range of microplate formats and is suitable for numerous applications in life science research and drug discovery.
Automatically generated - may contain errors
Lab products found in correlation
96 well plate, by Merck Group (2 mentions)
Alamar blue, by Thermo Fisher Scientific (2 mentions)
Trypan blue, by Thermo Fisher Scientific (2 mentions)
Alamarblue assay, by Thermo Fisher Scientific (2 mentions)
D1844, by Thermo Fisher Scientific (1 mentions)
Evom voltohmeter, by World Precision Instruments (1 mentions)
Dual glo luciferase assay system, by Promega (1 mentions)
96 well microplate, by Corning (1 mentions)
Alamarblue, by PerkinElmer (1 mentions)
Kaleidagraph software, by Synergy Software (1 mentions)
16 protocols using victor 3v 1420
1
Exosome-Mediated Modulation of CD8+ T Cells
2
Measuring Endothelial Cell Permeability
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TEER was measured using a EVOM voltohmeter connected to an Endohm 9 electrode chamber (World Precision Instruments, Sarasota, FL) as previously described [29 (link)]. Tissue culture inserts containing monolayers of HGEc cultured on Transwell-Collagen-coated membrane, were placed in the chamber and the TEER was recorded after 10 sec [29 (link)]. The permeability experiments were done as described before [31 (link)]. Briefly, 3 x 105 HGEC-1 were plated onto Transwell Collagen-coated membrane inserts (Corning Costar, Cat No 3415) in DMEM media with 10% FBS, and left for 3 days to form mature monolayer with a trans-endothelial electrical resistance (TEER) of ~ 60 ± 10 Ω /cm2. Subsequently, the cells were starved for 5 hours in serum free DMEM media without phenol red, treated with the corresponding reagents, and incubated with 1 mg/ml FITC-dextran (Molecular Probes, D-1844, Invitrogen) for 30 min at 37°C. Samples collected from the bottom chambers were read in triplicates on the Victor 3V1420 multi-counter (Perkin-Elmer, Wellesley, PA).
3
Rim1 Binding Affinity Determination
4
Assessing U373MG Cell Viability
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U373MG cells were seeded at a density of 1 × 104 per well into 96- well plates (Sigma-Aldrich, Arklow, Ireland) and allowed to adhere and grow overnight. Media was removed for the duration of CAP treatment and fresh media was replaced immediately after treatment and incubated at 37 °C as indicated. No deleterious effects were observed in the vehicle control samples. Cell viability was analysed using Alamar Blue (Fischer Scientific, Ballycoolin, Ireland). Cells were washed once with sterile PBS, incubated for 2.5 hours at 37 °C with a 10% Alamar Blue solution. Fluorescence was measured using an excitation wavelength of 530 nm and emission wavelength of 595 nm on a Victor 3 V 1420 (Perkin Elmer) multi-plate reader.
5
Gyrase-YacG Interaction Analysis
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Gyrase tetramers, individual subunits, and isolated domains were mixed at different concentrations with 50 nM N-terminally Alexa Fluor 488-labeled YacG in a buffer containing 120 mM potassium glutamate, 0.12 mg/mL bovine serum albumin (BSA), 6 μM ZnCl2, 12% (v/v) glycerol, and 30 mM Tris-HCl (pH 7.9). Measurements were collected using a Perkin Elmer Victor 3V 1420 multilabel plate reader at 535 nm. All points are normalized to wells that did not contain reconstituted gyrase, gyrase subunits, or gyrase domains. For competition experiments, 750 nM reconstituted gyrase was incubated with 50 nM labeled wild-type YacG, and various nonlabeled YacG constructs were titrated into the solution.
6
Evaluating Cell Viability with Trypan Blue and Alamar Blue
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Trypan Blue cell exclusion assay was performed as an initial evaluation of cell health following treatment with UA. Cell were trypsinized as described above. A Trypan Blue (Fischer Scientific, Ballycoolin, Ireland) cell suspension was counted using a haemocytometer as per manufacturer’s instructions. Cell viability was also measured biochemically using the Alamar Blue assay (Fischer Scientific, Ballycoolin, Ireland). Alamar blue is an oxidation-reduction (redox) fluorogenic indicator of cellular metabolic reduction. After each exposure time point, (24 h or 48 h) cells were washed once with sterile Phosphate-buffered saline (PBS). A 10% Alamar blue solution in the DMEM was added to each well and incubated at 37 °C for 2.5 h. Fluorescence was read at an excitation wavelength of 530 nm and an emission wavelength of 595 nm using the Victor 3V 1420 (Perkin Elmer) multi-plate reader.
7
UPF1 Silencing Impacts NMD Luciferase Assay
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HeLa cells were transfected with either control siRNA or UPF1si for two consecutive days, and on day 3 were co-transfected with NMD-firefly (FLuc-5xboxb) and Renilla luciferase (RLuc) and either λN or λN-ARS2n. Firefly and Renilla luminescence were analyzed 72 h after the first transfection using the Dual-Glo Luciferase Assay System (Promega) and a Perkin Elmer Victor3V 1420 multi-label plate reader. Firefly luminescence was normalized to Renilla luminescence.
8
E. coli Growth Kinetics Monitoring
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E. coli DP10 and PAS418 cells were grown overnight in LB medium. PAS418 cells were induced overnight with 2% (w/v) arabinose, and DP10/pCAM10 cells were induced for 4 h with 1 mM arabinose. Cells were washed three times in 1 × PBS, and then diluted 1:1,000 in fresh LB medium. 150 μl of diluted cells were transferred to 96-well microplates (Corning) with 100 μl of mineral oil overlaid on top to prevent evaporation. The plate was incubated in a Perkin Elmer Victor 3 V 1420 multilabel plate reader at 37 °C and medium shaking was performed for 8 h, and the optical density at 600 nm was recorded every 5 min.
9
Evaluating Cytotoxicity in U373MG and HeLa Cells
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U373MG cells were seeded at a density of 1 × 104, and HeLa cells were seeded at 0.8 × 104 into 96-well plates (Sigma-Aldrich) and allowed to adhere overnight. Media was removed for the duration of NTAP treatment, and fresh media was replaced immediately after treatment and incubated at 37 °C as indicated. No deleterious effects were observed in the vehicle control samples.
Cell viability was analysed using Alamar Blue, a redox fluorogenic indicator of metabolic reduction (Fischer Scientific, Ballycoolin, Ireland) (Page et al, 1993 (link)). Cells were washed once with sterile PBS, incubated for 2.5 h at 37 °C with a 10% Alamar Blue solution, and fluorescence was measured using an excitation wavelength of 530 nm and an emission wavelength of 595 nm on a Victor 3V 1420 (Perkin Elmer, Waltham, MA, USA) multi-plate reader. The Trypan Blue (Biosciences, Dun Laoghaire, Ireland) cell dye exclusion assay was also performed 48 h post treatment in HeLa cells and 96 h post treatment in U373MG cells. Both floating and trypsinised cells were collected, and a 1 × 106 cell per ml cell suspension was prepared. A 1 : 1 mixture of cell suspension and 0.4% trypan blue was loaded onto a haemocytometer for counting.
Cell viability was analysed using Alamar Blue, a redox fluorogenic indicator of metabolic reduction (Fischer Scientific, Ballycoolin, Ireland) (Page et al, 1993 (link)). Cells were washed once with sterile PBS, incubated for 2.5 h at 37 °C with a 10% Alamar Blue solution, and fluorescence was measured using an excitation wavelength of 530 nm and an emission wavelength of 595 nm on a Victor 3V 1420 (Perkin Elmer, Waltham, MA, USA) multi-plate reader. The Trypan Blue (Biosciences, Dun Laoghaire, Ireland) cell dye exclusion assay was also performed 48 h post treatment in HeLa cells and 96 h post treatment in U373MG cells. Both floating and trypsinised cells were collected, and a 1 × 106 cell per ml cell suspension was prepared. A 1 : 1 mixture of cell suspension and 0.4% trypan blue was loaded onto a haemocytometer for counting.
10
ABA-induced PP2C phosphatase activity
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Receptors were tested for activity as previously described [38 (link)]. Protein encoding receptors and PP2Cs were mixed at an approximate molar ratio of 10:1 receptor (0.024 μg/μL final concentration) to phosphatase (0.004 μg/μL final concentration), with ABA or ABA analog (0.1 μM unless otherwise noted) and buffer containing 100 mM Tris pH 7.9, 100 mM NaCl, 0.3 mM MnCl2 and 4 mM DTT in a total final volume of 50 μL. The mixture was incubated for 15 min at 30°C. Fifty μL of substrate (1 mM 4-Methylumbelliferyl phosphate) was then added to initiate the reaction and the assay mixture was incubated at 30°C. The intensity of the fluorescent product was the measured using a Perkin Elmer Victor 3 V 1420 fluorescent plate reader at 30 min after initiation of the assay. The excitation wavelength was 355 nm, the emission wavelength was 460 nm, and the measurement time was 0.1 s.
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