48 h following transfection 7 μl substrate of the Cell Titer-Blue® Cell Viability Assay (Promega) were added to each well. After a short centrifugation step (1,000 rpm for 10 sec) cells were incubated for further 3 h at 37°C / 5% CO2 before fluorescence reading (Victor X4, PerkinElmer). The activity of Caspase-3/7 was determined using the Caspase-Glo® 3/7 Assay (Promega). 20 μl substrate per well were applied and after 30 minutes shaking at room temperature in the dark, luminescence was detected (Victor X4, Perkin Elmer). The analysis of data was partially done online by Networkanalyst and GraphPad Prism.
Victor x4
Manufactured by PerkinElmer
Sourced in United States, Italy, Germany, Finland
The Victor X4 is a high-performance multimode microplate reader designed for a wide range of applications in life science research and drug discovery. It features advanced detection technologies, including absorbance, fluorescence, and luminescence, to enable reliable and sensitive measurements across multiple assay formats. The Victor X4 provides a flexible and configurable platform to meet the diverse needs of researchers and laboratories.
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Lab products found in correlation
Amplex red, by Thermo Fisher Scientific (5 mentions)
Caspase glo 3 7 assay, by Promega (5 mentions)
White 96 well plate, by Corning (4 mentions)
Celltiter glo, by Promega (4 mentions)
2 nbdg, by PerkinElmer (4 mentions)
Low glucose dmem, by Thermo Fisher Scientific (3 mentions)
Horseradish peroxidase, by Roche (3 mentions)
Celltiter glo luminescent cell viability assay, by Promega (3 mentions)
Ionomycin, by Merck Group (3 mentions)
Facscalibur, by BD (3 mentions)
Sytox green, by Thermo Fisher Scientific (3 mentions)
Slide 8 well, by Ibidi (2 mentions)
Glucose depleted dmem, by Thermo Fisher Scientific (2 mentions)
Lsm 700 confocal laser scanning microscope, by Zeiss (2 mentions)
Ex cytox, by DoGenBio (2 mentions)
Celltiter blue, by Promega (2 mentions)
Rnaimax, by Thermo Fisher Scientific (2 mentions)
Cs assay kit, by Merck Group (2 mentions)
Adipored, by Lonza (2 mentions)
Crystal violet, by Merck Group (2 mentions)
200 protocols using victor x4
1
Cell Viability and Apoptosis Assay
2
Cell Viability and Caspase Assay
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Forty-eight hours following transfection, 7 μl substrate of the Cell Titer-Blue Cell Viability Assay (Promega) was added to each well. After a short centrifugation step (1000 rpm for 10 seconds), cells were incubated for further 3 hours at 37°C/5% CO2 before fluorescence reading (Victor X4, PerkinElmer). The activity of Caspase-3/7 was determined using the Caspase-Glo 3/7 Assay (Promega). Twenty microliters of substrate per well was applied, and after 30-minute shaking at room temperature in the dark, luminescence was detected (Victor X4, Perkin Elmer). The analysis of data was done online by Networkanalyst and GraphPad Prism.
3
Intestinal Permeability Measurement
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On the day of euthanisation, intestinal integrity was determined by measuring the permeability of FITC-dextran, using a similar approach as previously described [9 (link)]. Animals were fasted for at least 9 hours before the assay. From each cage, one animal was orally dosed with 0.5 ml 120 mg/ml FITC-dextran (4 kDa, Sigma-aldrich FD-4) per 100 g (corresponding to 600 mg/kg animal) bodyweight while the other was dosed with 0.5 ml PBS per 100 g bodyweight. Two hours after dosage, animals were euthanized and blood was collected from the neck directly into 50 ml Falcon tubes with 100 μl EDTA (0.5 M, pH 8, Ambion). Blood samples were immediately centrifuged (3800 rpm, 5 min) to collect plasma. Plasma was centrifuged again, diluted 1:1 in PBS and stored at 5°C until analysis on the same day. Analysis of each sample was done in triplicate by transferring volumes of 60 μl plasma-PBS solution to a black 96-well microtiter plate (Proxiplate-96 F, Perkin Elmer) and measuring the florescence at excitation 485 nm/emission 535 nm (Victor TM X4, Perkin Elmer). Standard curves were prepared for each of the euthanisation days, by adding fixed concentrations of FITC-dextran to plasma-PBS prepared from animals dosed with PBS.
4
Assessing AuNP Cytotoxicity in Adipocytes
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EZ-Cytox assay kits (DOGEN Bio., Seoul, Korea) were used according to the manufacturer’s instructions for measuring cell viability. Differentiated 3T3-L1 adipocytes were incubated with various concentrations of AuNPs (0, 5, 10, 20, 30, and 60 μg/mL) for 48 h and were subsequently assessed at 450 nm using a VictorTM X4 (PerkinElmer, Waltham, MA, USA) microplate reader.
5
Cellular Viability Assay Protocol
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Cellular viability assays were performed as previously described [21 (link),22 (link)]. Briefly, cells were seeded into opaque black 96-well plates. To determine the cellular viability of siRNA-treated cells, reverse lipid-based transfections were performed (RNAiMAX, life technologies, Carlsbad, CA, USA). Following defined incubation periods, cellular viability was assessed using the CellTiter-Blue® reagent (Promega) in a plate reader (VICTORTM X4, Perkin Elmer, Waltham, MA, USA). All experiments were conducted as technical and biological triplicates.
6
Glucose Uptake Assay in 3T3-L1 Adipocytes
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The glucose uptake assay was performed using a previously published method, with slight modifications [57 (link)]. Fully differentiated 3T3-L1 adipocytes in µ-Slide 8 wells (Ibidi, München, Germany) were incubated overnight in low-glucose DMEM (Gibco BRL, Middlesex, UK). The cells were then treated with ethyl gallate in glucose-depleted DMEM (Gibco BRL, Middlesex, UK) for 6 h. Insulin treated independently for 30 min was used as the positive control. The fluorescent glucose analog, 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl)amino]-2-deoxyglucose (2-NBDG; 100 μM), was added, and the cells were incubated for 1 h. Cells were then quickly washed with pre-cooled DPBS, and the fluorescence signal in cells was detected using confocal microscopy (LSM700 laser scanning confocal microscope, Carl Zeiss, Oberkochen, Germany). For quantitative analysis, 3T3-L1 preadipocytes were differentiated in 96-well plates independently and handled as described above for confocal microscopy, but the fluorescence signal in the cells was measured using a microplate reader (VictorTM X4, PerkinElmer, Inc., Waltham, MA, USA).
7
Quantification of hCLEC14a-CTLD-Fc-HRP Binding
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Human umbilical vein endothelial cells (1 × 104) were grown in 0.1% (w/v) gelatin‐coated wells of a 96‐well plate overnight at 37 °C. Following fixation with 4% (w/v) PFA, the cells were incubated with 3 μg·mL−1 hCLEC14a‐CTLD‐Fc‐HRP in the presence or absence of increasing concentrations of 20 μg·mL−1 deglyco C1 IgG for 2 h at 37 °C. After three washes with ice‐cold PBS, 100 μL of TMB substrate solution (BD Biosciences) was added to each well. The reaction was stopped by the addition of an equal volume of 1 N H2SO4. The optical density was measured at 450 nm using a spectrophotometer (VICTORTM™ X4; PerkinElmer).
8
ROS-Glo Assay for Oxidative Stress
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Similar to GSH measurements, two 96-well plates were seeded at 1 × 104 cells/well density for each cell line. DMSO (vehicle) and 1E5 were used as treatments. Incubation times were same as in Section 2.9 . At 24 h of incubation, the media was replenished and supplemented with 20 μL of H2O2 substrate. The plates were then incubated for 6 h at 37 °C. After completion, 50 μL of (media + H2O2) substrate was pipetted to a 96-well white plate (Corning Inc., Corning, NY, USA, #4517). Freshly prepared 50 μL of ROS-GloTM (Promega, Madison, WI, USA, #G8820) solution was added and incubated for 20 min. After 20 min, luminescence was read using the PerkinElmer VictorTM X4.
9
Cell Viability Assay of PNS on H9c2 Cells
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The cell viability was tested by the methyl thiazolyl tetrazolium (MTT) assay (M2128, Sigma-Aldrich, USA). H9c2 cells were seeded in a 96-well plate with 5,000 cells per well for 48 h. Cells were then treated by various concentrations of PNS with or without 1 μM TG stimulation for 24 h. Twenty microliters of MTT (5 mg/ml) was added to each well for 4 h and then replaced by 150 µl dimethyl sulfoxide (DMSO). The optical density of the plate was measured at 570 nm using a multilabel microplate reader (VICTORTM X4, PerkinElmer, Inc., USA). The cell viability of the untreated control was considered as 100%.
10
IgG Detection in Chicken Sera via ELISA
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A laboratory-made indirect enzyme-linked immunosorbent assay (ELISA) using heat-killed whole S. enterica ser. Gallinarum bacteria as the coating antigen was applied to quantify IgG in the serum. Briefly, bacterial antigen (106 CFU/well) was coated in a 96-well plate and incubated overnight at 4°C. After blocking, the sera samples collected on days 14, 21, and 28 were diluted (1:400) with 2% bovine serum albumin (BSA) and added to the wells. The wells were incubated with a 1:8,000 dilution of affinity-purified, peroxidase-labeled goat anti-chicken IgG (H+L) (KPL Inc., MD, USA) for 1 h. OD450 was measured with a microplate reader VICTORTM X4 (PerkinElmer Inc., Waltham, MA, USA) after the reaction was stopped with 4.5 N H2SO4. The OD450 average value of negative serum samples plus the standard deviation (SD) of three times was used as the cut-off value (27 (link)).
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