Cell viability was evaluated using a CCK-8 kit (Bimake, Houston, TX, USA) to assess the cytotoxicity of piceatannol. In short, PK-15 cells in 96-well plates were treated with a serial twofold dilution of piceatannol and incubated at 37 °C and 5% CO2 for 48 h. At the same time, a group of cells without continuous s was maintained as a control. After 48 h, CCK-8 solution was added according to the instructions, and the cultures were incubated at 37 °C for 45 min. The OD value of each well at 450 nm was measured with a plate reader, and the cell survival rate was calculated to evaluate the cytotoxicity of piceatannol. To determine the inhibitory activity of piceatannol against PRV, different concentrations of piceatannol were mixed with an equal volume of 100 TCID50 of PRV solution at a nontoxic concentration and incubated at 37 °C for 1 h. The above mixture was added to 96-well cell culture plates, which were incubated for 1 h and then washed with PBS, and cell maintenance solution was then added. Forty-eight hours later, the OD values at 450 nm of each well were determined, and the virus inhibition rate was calculated to evaluate the inhibitory activity of piceatannol against PRV. The half inhibitory concentration (IC50) and half cytotoxic concentration (CC50) of piceatannol were calculated with GraphPad Prism 8.0 software.
Cck 8 kit
Manufactured by Selleck Chemicals
Sourced in United States, China
The CCK-8 kit is a colorimetric assay used to measure cell viability and cytotoxicity. It contains a water-soluble tetrazolium salt that is reduced by dehydrogenase enzymes in living cells, producing a colored formazan dye that can be measured spectrophotometrically.
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Lab products found in correlation
Cytation 5 cell imaging multi mode reader, by Agilent Technologies (2 mentions)
Hct 116 colon cancer cell line, by American Type Culture Collection (2 mentions)
Phosphate buffered saline (pbs), by PAN Biotech (2 mentions)
Dulbecco modified eagle medium (dmem), by PAN Biotech (2 mentions)
Prism 8, by GraphPad (1 mentions)
Microplate reader, by Thermo Fisher Scientific (1 mentions)
Spectramax m3, by Molecular Devices (1 mentions)
Cell light edu apollo567 in vitro kit, by RiboBio (1 mentions)
Crystal violet, by Beyotime (1 mentions)
Juli stage real time cell history recorder, by NanoEnTek (1 mentions)
Trypan blue, by Solarbio (1 mentions)
1 step pnpp substrate solution, by Thermo Fisher Scientific (1 mentions)
Synergy h1, by Agilent Technologies (1 mentions)
Erk1 2, by Cell Signaling Technology (1 mentions)
Phospho kit tyr719, by Cell Signaling Technology (1 mentions)
Ab15580, by Abcam (1 mentions)
Gapdh, by Cell Signaling Technology (1 mentions)
Imatinib, by Selleck Chemicals (1 mentions)
Sorafenib, by Selleck Chemicals (1 mentions)
Sunitinib, by Selleck Chemicals (1 mentions)
28 protocols using cck 8 kit
1
Cytotoxicity and Antiviral Effects of Piceatannol
2
CCK8 Viability Assay for ESCC
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The viabilities of treated ESCC cells were detected using CCK8 kit obtained from Bimake (Shanghai, China). The treated cells were grown in a 96-well plate at a density of 2 × 103 per well. After incubation for indicated time, the cells were harvested and added with 10 μl CCK8 solution and 90 μl medium in each well and cultured for another 4 h. Afterwards, the absorbance at 450 nm was evaluated in a microplate reader (Thermo Fisher Scientific, MA, USA).
3
Huh7 Cell Viability Assay
4
ETEC-Induced IPEC-J2 Cell Proliferation
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The impact of CP9 on the proliferation of IPEC-J2 cells during ETEC infection was assessed as previously described (124 (link)), with minor modifications. Briefly, IPEC-J2 cells were seeded in a 96-well plate at a density of 3 × 104 cells/well, followed by incubation at 37°C for 24 h. Spent medium was then replaced with CO2-independent medium without antibiotics. IPEC-J2 cells were stimulated with 108 CFU of ETEC/mL for 2 h at 37°C. The spent medium was then removed, and cells were incubated with fresh medium containing 108 CFU of CP9/mL at 37°C for an additional 6 h. The final volume in each well was 100 μL. Cells with or without FBS were used as controls. Cell proliferation was measured by using a CCK-8 kit (Bimake, Houston, TX) according to the manufacturer’s instructions. Briefly, 10 μL of the CCK-8 reagent was added to each well, followed by incubation at 37°C for 2 h. Metabolically active cells in proliferation were counted by measuring the absorbance at 450 nm using a Cytation 5 Cell Imaging Multi-Mode Reader (BioTek Instruments).
5
Cell Viability, Proliferation, and Colony Formation Assays
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Cell counting kit-8 (CCK-8) assay was performed using a CCK-8 kit (Bimake, TX, USA) according to the manufacturer’s instructions. In brief, cells were seeded at a density of 5 × 103 cells/well in 96-well plates in 100 μL complete culture medium and cultured successively for 24, 48, 72, and 96 h. Next, CCK-8 (10 μL) was added to each well and after 30 min, the optical density (OD) values were detected at 450 nm with a microplate reader (SpectraMax M3; Molecular Device, CA, USA).
5-Ethynyl-2′-deoxyuridine (EdU) incorporation assay was performed using a Cell-Light ™ EdU Apollo567 In Vitro Kit (RiboBio, Guangdong, China) according to the manufacturer’s instructions, as described previously.29 (link)
For colony formation assays, cells were plated into 6-well plates at a density of 1 × 103 cells/well. After 10 days, the supernatant was discarded and the cells were fixed with 4% paraformaldehyde and stained with 0.1% crystal violet (Beyotime, Shanghai, China). Images were captured using an EPSON V330 scanner (EPSON, Nagano, Japan).
5-Ethynyl-2′-deoxyuridine (EdU) incorporation assay was performed using a Cell-Light ™ EdU Apollo567 In Vitro Kit (RiboBio, Guangdong, China) according to the manufacturer’s instructions, as described previously.29 (link)
For colony formation assays, cells were plated into 6-well plates at a density of 1 × 103 cells/well. After 10 days, the supernatant was discarded and the cells were fixed with 4% paraformaldehyde and stained with 0.1% crystal violet (Beyotime, Shanghai, China). Images were captured using an EPSON V330 scanner (EPSON, Nagano, Japan).
6
Cell Cycle, Growth, and Mobility Analysis
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Cell cycle analysis was conducted as described previously 30 . Cell growth and cell mobility were monitored by JULI Stage Real-time Cell History Recorder (NanoEntek, Seoul, South Korea) as described previously 30 . Images were taken, and the growth rate and cell mobility ability were quantified. Cell viability was analyzed by trypan blue (Solarbio Life Sciences, Beijing, China) staining. Cell numbers were counted by CellDrop FL Fluorescence Cell Counter (Devovix, USA). Cell growth was also checked by CCK-8 Kit (Bimake, China) 21 (link), 28 (link). Wound-healing assays were conducted as described previously 28 (link).
7
Cell Viability Assay Using CCK-8 Kit
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CCK‐8 kit (Bimake, Houston, TX, USA) was used to evaluate cell viability. Cell suspension was inoculated in 96‐well plates (3 × 103 cells per well). CCK‐8 solution (10 μL) was added to the wells every 24 h. After incubation for 1.5 h, the absorbance was measured at 450 nm using a microplate reader.
8
Osteogenic Differentiation ALP Activity
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After 6 days of osteogenic differentiation, ALP activity was determined using 1-Step PNPP Substrate Solution (Thermo Fisher Scientific; Cat N°: 37621) following manufacturer’s instructions. For ALP data normalization, cell number was determined by CCK-8 kit (Bimake, Cat N°: B34304) in a parallel 96-well plate.
9
Cellular Proliferation Evaluation with CP Extract
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The cells were plated at the density of 104 cells in 96 wells until confluent and changed to CP extract containing serum-free medium for 24 hours before testing with a WST based proliferation/cytotoxicity assay using the CCK8 kit (cat no B34304, Bimake, Houston, USA). Four groups were tested, which include (1) the one treated directly with the CP extract (dir-CP), (2) cells treated with conditioned medium collected from the 24 hour CP extract treated cells (CP-sup), (3) cells treated with conditioned medium collected from the 24-hour incubation without CP extract (sup-Control), and (4) cells without any treatment (control).
10
Cell Viability Assay of Immunocompromised hPDLC
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The immunocompromised hPDLC cells after indicated treatment were seeded into 96-well plates at the density of 1 × 103 cells/well. Cell viability was detected with CCK-8 kit (Bimake, Houston, USA). Briefly, cells were plated into 96-well plates at about 104 cells/well and treated with CCK-8 solution for 2 h. The absorbance was detected with a microplate reader at 450 nm wavelength.
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