Cells were seeded at 2.0 x 104 cells per well in a 96-well tissue culture plate and allowed to adhere overnight. Media was changed the next day 1 hour prior to stimulation. Cells were stimulated with 1 ng/ml LPS +/- IL10 (0.01–30 ng/ml) for 1 hour. Triplicates wells were used for each stimulation condition. Supernatant was collected and secreted TNFα protein levels were measured using a BD OptEIA Mouse TNFα Enzyme-Linked Immunosorbent Assay (ELISA) kit (BD Biosciences, Mississauga, ON). The inhibitory concentration (IC50) of IL10 needed to inhibit LPS-stimulated TNFα production was calculated using GraphPad (non-linear regression curve fit). The mean IC50 was determined from three independent IC50 determination experiments.
Opteia mouse tnfα enzyme linked immunosorbent assay elisa kit
Manufactured by BD
The BD OptEIA Mouse TNFα Enzyme-Linked Immunosorbent Assay (ELISA) kit is a laboratory tool designed to quantitatively measure the levels of tumor necrosis factor alpha (TNFα) in mouse biological samples.
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2 protocols using opteia mouse tnfα enzyme linked immunosorbent assay elisa kit
1
Inhibition of LPS-stimulated TNFα by IL10
2
Measuring TNFα Secretion in LPS-Stimulated Macrophages
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Cells were seeded at 1.5 x 104 cells per well in a 96-well tissue culture plate and allowed to adhere overnight. Media was changed the next day 1 hour prior to stimulation. Cells were stimulated with 1 ng/ml LPS +/- IL10 (0.5 or 10 ng/ml) for 1 hour. Triplicate wells were used for each stimulation condition. Supernatant was collected and secreted TNFα protein levels were measured using a BD OptEIA Mouse TNFα Enzyme-Linked Immunosorbent Assay (ELISA) kit (BD Biosciences, Mississauga, ON).
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