HEK293T cells were transfected with a firefly luciferase reporter plasmid (pGL2 Sv40 min or pGL3 basic, as indicated), a Renilla reporter plasmid pTK-RLUC (Promega), and as indicated with pCMX-LXRα and pCMV-RXRα, or empty pCMX. 48h after transfection cells were treated with vehicle (DMSO) or 1 μM GW3965 for 24 hours. Subsequently, the cells were washed twice in PBS and lysed in passive lysis buffer following the manufacturers instructions. The samples were then measured using the Dual-Luciferase Reporter Assay System (Promega) on a Glowmax Multi detection system (Promega) according to the manufacturer’s protocol. Each experiment was repeated at least three times in triplicate.
Glowmax multi detection system
Manufactured by Promega
Sourced in United States
The Glowmax multi detection system is a versatile lab equipment designed for sensitive and accurate detection of various luminescent and fluorescent assays. It provides reliable measurement capabilities for a wide range of applications, including cell-based assays, reporter gene studies, and biochemical analyses. The Glowmax multi detection system employs advanced optical technology to deliver consistent and reproducible results, enabling researchers to obtain high-quality data for their experimental needs.
Automatically generated - may contain errors
Lab products found in correlation
Dual luciferase reporter assay system, by Promega (2 mentions)
Mouse elisa kit, by R&D Systems (2 mentions)
White flat bottom 96 well plate, by Corning (1 mentions)
Celltiter glo luminescent reagent, by Promega (1 mentions)
Caspase glo 3 7 assay reagent, by Promega (1 mentions)
Celltiter glo, by Promega (1 mentions)
Prl tk plasmid, by Promega (1 mentions)
Prl cmv, by Promega (1 mentions)
Gene pulser 2, by Bio-Rad (1 mentions)
9 protocols using glowmax multi detection system
1
Dual-Luciferase Assay for LXR Activity
2
Evaluating Synergistic Cell Viability
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Cell proliferation was measured with an impedance-based instrument system (xCELLigence, Roche/ACEA Biosciences, Basel, Switzerland) enabling label-free real time cell analysis. Briefly, 2-4 × 104 cells were seeded into 96-wells with 200 μl culture media containing FBS and allowed to grow up to 150 h. Cellular impedance was measured periodically every 4 h (relative cell index) and HDACi, EEDi, chemotherapeutics or DMSO was added as indicated in the Results section.
For synergy evaluation 0.5-1 × 104 cells were seeded into white, flat-bottom, 96-well plates (Corning). Twenty-four hours later, doxorubicin and/or HDACi (FK228 or MS-275) were added, and viability was assessed after 72 h. Cell viability was measured using CellTiter-Glo Luminescent Reagent according to the manufacturer’s protocol (Promega). Luminescence was measured on a Promega GlowMax-Multi Detection System. Synergistic interactions were evaluated using the package Synergyfinder [30 (link)] (Package version 2.2.4, R version 4.0.3, RStudio version 1.3.1093) according to the Bliss algorithm [31 ].
For synergy evaluation 0.5-1 × 104 cells were seeded into white, flat-bottom, 96-well plates (Corning). Twenty-four hours later, doxorubicin and/or HDACi (FK228 or MS-275) were added, and viability was assessed after 72 h. Cell viability was measured using CellTiter-Glo Luminescent Reagent according to the manufacturer’s protocol (Promega). Luminescence was measured on a Promega GlowMax-Multi Detection System. Synergistic interactions were evaluated using the package Synergyfinder [30 (link)] (Package version 2.2.4, R version 4.0.3, RStudio version 1.3.1093) according to the Bliss algorithm [31 ].
3
Fluorometric ATP Assay Protocol
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Cellular ATP concentration was determined using a fluorometric ATP assay kit (BioRad, Milpitas, CA) following the manufacturer’s instructions, and fluorescence was read at 525 nm on a Glowmax Multi Detection System (Promega Corporation, Madison, WI).
4
Caspase-3/7 Activity Assay for BL31 Cells
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BL31 and BL31 wtBAC2 cells (100 µl) were seeded into 96 well plates at a density of 20,000 cells/well and cultured for 8 or 18 hours in the presence or absence of UNC1999. An equal volume of Caspase-Glo 3/7 Assay reagent (Promega, UK) was added, and cells were incubated for 30 min at room temperature. Luminescence was measured using a Glowmax multi detection system (Promega).
5
Quantifying CXCL13 Levels in TB Treatment
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The Quantikine ELISA assay for human CXCL/13/BLC/BCA-1 was used to measure the amount of CXCL13 in the plasma of 19 participants before and after 6 months of successful TB treatment. Standards, controls and samples were run in triplicate. Results were measured at 450nm using the GlowMax- Multi detection system (Promega). Concentrations were determined based on the standard curve generated on GraphPad prism version 6.0 (GraphPad Software, Inc.). Protein levels for mouse cytokines (IL-17, IL-22 and IL-23) in culture supernatants were measured using mouse ELISA kits or multiplex according to manufacturer’s instruction (R&D Systems, MBL International Corporation).
6
Cell Viability Assay Using CellTiter-Glo
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Cell viability was assessed using CellTiter-Glo (Promega). Briefly, for CellTiter-Glo measurement, 1,000 cells were seeded in white, flat-bottom, 96-well plates. After 24 hours, drugs were added to the medium and cells were incubated for 72 hours. CellTiter-Glo luminescent reagent was added according to the manufacturer's protocol, and the luminescence signal measured on a Glowmax-Multi Detection System (Promega).
7
PTEN Promoter Regulation in NB4 Cells
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The PTEN regulatory promoter region was subcloned in PGL4.1 FLuc reporter plasmid (Promega, Madison, USA), using the following primers: forward: 5′-CCCTCTTTAGACTTTGCTAGGC-3′ and reverse: 5′-GGTAGGAGGGGGCAGAGC-3′. NB4 cells were electroporated using the Amaxa Nucleofector system according to the manufacturer's protocol. Briefly, 2 × 106 cells were co-transfected with 2 ug of PGL-4 PTEN promoter and 200 ng of pRLTK plasmid (Promega, Madison, USA) encoding for renilla luciferase gene as internal control. Electroporation was performed by re-suspending the cells in 100 μl Cell Line Nucleofector® Solution V, Program X-001. Immediately after electroporation, cells were treated with 1uM ATRA for 48hrs. Luciferase was measured using the Dual-Luciferase Reporter Assay System (Promega, Madison, USA), according to the manufacturer's instructions. Luminescence was measured with the “Dual Glow” protocol of the Glowmax MULTI + Detection System (Promega, Madison, USA).
8
Quantifying CXCL13 Levels in TB Treatment
9
Promoter Activity Assay for RUNX1 and RUNX3
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For RUNX1 and RUNX3 promoter reporter assays, DG75 cells were electroporated with plasmid DNA at 230 V and 950 μF (BioRad Gene Pulser II) and luciferase assays carried out as described previously (45 (link)) using sequential injection on a Glowmax multi detection system (Promega). Cells were transfected with 2 μg of the pGL3 luciferase reporter plasmids and 0.5 μg pRL-CMV (Promega) as a transfection control, in the absence or presence of 10 or 20 μg of the EBNA2-expressing plasmid pSG5 EBNA2A. One tenth of each transfection was processed for western blotting to analyse protein expression levels.
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