Science Catalog 2012 Life Cell Signaling Chroma-Glo™ Luciferase Assay System Product Chroma-Glo™ Luciferase Assay System For Research Use Only. Not for Use in Diagnostic Procedures. Description: The Chroma-Glo™ Luciferase Assay System and the Chroma-Luc™ Vectors provide a method to generate red and green (dualcolor) luminescence from a single sample upon a single-reagent addition. Filtered measurement of the dual-color luminescence produced by the Chroma-Luc™ luciferases permits each reporter to be measured independently and virtually simultaneously. The Chroma-Glo™ Assay is in a homogeneous format that generates luminescence with >30-minute signal half-lives for each of the Chroma-Luc™ luciferases, thereby enabling the processing of many plates without prior sample handling. Use the high-homology Chroma-Luc™ luciferases to establish an ideal internal control for normalizing cytotoxicity in downregulation applications and for decreasing inter- and intrasample variability. You can also use the reporters to multiplex experimental reporters to increase the data content from cell-based assays. Features: • Measure Dual Reporters Using a Single Substrate Addition: Increase your accuracy and precision through normalization, or use both reporters to multiplex experimental measurements. Use filters to spectrally separate the luminescent signals. • Establish the Ideal Control or Multiplexed System: Use the highhomology red and green luciferases to minimize potential RNA and protein effects on reporter expression. • Increase Your Throughput: Use the stable luminescence for batch or continuous processing of multiple plates. • Perform Fewer Steps: Add Chroma-Luc™ Reagent directly to cells in medium, then measure. • Automate This Assay: Validated automated methods available at: www.promega.com/automethods/ • Choose Your Configuration: Learn more about our custom options for this product at: www.promega.com/myway/ Storage Conditions: Store the Chroma-Glo™ Substrate at –20°C. Store the Chroma-Glo™ Assay Buffer below 25°C. Protocol Chroma-Glo™ Luciferase Assay System Technical Manual Part# TM062 Size Cat.# Price (Fr) 10 ml E4910 338.00 100 ml E4920 2366.00 10 15 20 25 0 5 CRE-CBG99luc NFκB-CBRluc ISO TNFα Treatment Using the Chroma-Luc™ Technology to monitor two independent experimental signals from the same sample. DNA segments containing either CRE or the NFκB consensus sequence were cloned into pCBG99-Basic (Cat.# E1431) or pCBR-Basic (Cat.# E1411). The resulting constructs, pCRECBG99-luc and pNFκB-CBRluc, were cotransfected into 293 cells. At 24 hours post transfection, the cells received one of three treatments: ISO (1μM)/ RO(100μM), TNFα (0.1μg/ml)/RO(100μM), or ISO(1μM)/RO(100μM) plus TNFα (0.1μg/ml). Only RO(100μM) was added to the Control wells. At six hours post treatment, cells were harvested and assayed with the Chroma-Glo™ Reagent. Relative light units were measured using the Mithras LB940 (Berthold Technologies) configured with a red filter (610 long pass) and a green filter (510/60). The red and green signals were deciphered by using the ChromaLuc™ Calculator (available as a downloadable file at: www.promega.com/chromacalc/). Fold inductions were calculated by dividing the three treatments by the RO Control. ISO +TNFα 262 For complete and up-to-date product information visit: www.promega.com/catalog Fold Induction 4222MA06_3A Pagina 265
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