CCK-8 (Beyotime Institute of Biotechnology) was carried out to determine the cell viability of HCT116 cells following the manufacturer's protocol. Cells were transfected for 24 h, re-seeded into 96-well plates (6×103 cells/well) and incubated for 0, 24, 48 and 72 h at 37°C. Subsequently, CCK-8 reagent (10 µl) was added to the cells and incubated for 4 h at 37°C. The absorbance was then analyzed at 450 nm using a microplate reader (FilterMax F3/F5; Molecular Devices, LLC).
Filtermax f3 f5
Manufactured by Molecular Devices
Sourced in United States
The FilterMax F3/F5 is a microplate reader that measures absorbance, fluorescence, and luminescence in microplates. It is designed to perform various assays, including cell-based, biochemical, and ELISA assays. The instrument provides accurate and reliable results with its advanced optical system and user-friendly software.
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5 protocols using filtermax f3 f5
1
Cell Viability Assay of HCT116 Cells
2
Cell Viability Assay with Zey
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Cells were inoculated in 96-well plates (2.5×103 cells/well) and cultured in an incubator for 24 h. Subsequent to being cultured, cells were exposed to different concentrations of Zey at 0, 2.5, 5, 10, 20 and 40 µmol/l for 12, 24 and 48 h. CCK-8 solution (Beyotime Institute of Biotechnology, Haimen, China) was added to the cells and the plate was transferred to the incubator for 4 h. Finally, absorbance was measured at 450 nm with a FilterMax F3/F5 microplate reader (Molecular Devices, LLC, Sunnyvale, CA, USA).
3
Quantifying Protein in Bronchoalveolar Lavage
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The protein concentration in the bronchoalveolar lavage fluids (BALF) was measured using the BCA assay. After the rats were sacrificed, PBS was injected into the lungs via the trachea, the liquid was aspirated slowly and the procedure was repeated 3 times. The supernatant of the aspirated liquid was separated by centrifugation at 12,000 × g for 10 min at 4°C and the protein concentration in the supernatant was measured. BCA kit was purchased from Biomiga, Inc., and the reagents were added according to the manufacturer's protocol. Absorbance at 570 nm was measured by a microplate reader (FilterMax F3/F5; Molecular devices, LLC) and the protein concentrations were calculated from the standard curve concentration.
4
Cell Viability Assay of LM3 Cells
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CCK-8 (MSK, Wuhan, China) was used to detect the viability of LM3 cells according to the manufacturer’s instructions. Specifically, cells were inoculated in 96-well plates at the density of 1.5×103 cell/well and were cultured in an incubator for 24 h. After being cultured, cells were treated with PBS, unspecific scrambled siRNA vector, and human STOML2-target siRNA vector. The experiments were divided into control, negative control (NC), and si-STOML groups. After grouping, CCK-8 reagent was added to cells, and cells were seeded into an incubator for 4 h. Absorbance was analyzed at 450 nm by a microplate reader (FilterMax F3/F5; Molecular devices, California, USA).
5
Interferon Stimulatory DNA Enhances IFN-β Expression
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Interferon stimulatory DNA (ISD ), a 45-bp non-CpG oligomer, can strongly enhance the expression of IFN-β in various cell types (Stetson and Medzhitov, 2006 (link); Ishikawa et al., 2009 (link)). ISD (InvivoGen, San Diego, CA) was applied as a simulator of IFN-β production for determining the effect of VP1 on IFN-β. Briefly, 5 × 104 cells were seeded onto 24-well plates and transiently transfected with the firefly luciferase reporter plasmid (100 ng), pRL-TK-Renilla luciferase reporter (50 ng), and indicated plasmids or pCMV-myc empty control vector (500 ng) using Lipofectamine 3000 from Thermo Fisher Scientific (Shanghai, China), with the STING plasmid or 2.5 μg of ISD transfection as a simulator.
After 24 h, cells were lysed, and samples were assayed with the dual luciferase reporter assay kit (Vazyme, Nanjing, China) and measured by a multitube chemiluminescence detector (FilterMax F3/F5, Molecular Devices). The reporter assays were repeated at least 3 times.
After 24 h, cells were lysed, and samples were assayed with the dual luciferase reporter assay kit (Vazyme, Nanjing, China) and measured by a multitube chemiluminescence detector (FilterMax F3/F5, Molecular Devices). The reporter assays were repeated at least 3 times.
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