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The Cignal Reporter Assays (luc) include pre-formulated, transfection-ready reporter, negative control, and positive control. The transcription factor reporter and negative control are transfected and subjected to experimental treatments, in parallel. Dual-luciferase results are calculated for each transfectant. The change in the activity of each signaling pathway is determined by comparing the normalized luciferase activities of the reporter in treated versus untreated transfectants. The identically treated negative control transfectants serve as a specificity control. The positive control serves as a control for transfection efficiency, by monitoring GFP expression, as well as a positive control for both the firefly and Renilla luciferase assays.
Excellent signal to noise ratio
Cignal ISRE reporter assay showed specific response
to IFN-α: Hela cells were transfected with ISRE reporter, negative
control and positive control (for transfection protocol refer our user
manual). After 16 hours of transfection, medium was changed to assay
medium (Opti-MEM + 10% heat inactivated FBS + 0.1mM NEAA + 1mM Sodium
pyruvate + 100 U/ml penicillin + 100 µg/ml streptomycin). After 24 hours
of transfection, cells were treated with 1000 U/ml of IFN-α for 18 hours.
Dual Luciferase assay was performed, and promoter activity values are
expressed as arbitrary units using a Renilla reporter for internal
normalization. Experiments were done in triplicates, and the standard
deviation is indicated.
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