A549 cells (1×105) were plated in a 24-well plate before transfection. Next day, cells were co-transfected with the control pcDNA3.1 empty vector (EV, 225 ng) or the wild-type TTF-1 plasmid (225ng)-based firefly luciferase reporter plus the renilla luciferase control vector pGL4.73 (50 ng, Promega) using the K-2 transfection reagent (Biontex, #T060). After transfection (48 hr later), cell lysates were prepared and firefly/renilla luciferase values were quantified using Firely & Renilla Dual Luciferase assay (Biotium, #30005-2) on a GloMax-96 plate reader (Promega). Firefly luciferase values were normalized to Renilla luciferase values and expressed as relative values. See Supplementary Information for additional experimental methods on RNA profiling, cholesterol efflux, intracellular cholesterol quantitation and animal studies.
Glomax 96 plate reader
Manufactured by Promega
Sourced in United States
The GloMax-96 plate reader is a microplate luminometer designed for detecting luminescent signals. It is capable of reading 96-well plates and can be used for various luminescence-based assays, such as cell viability, gene reporter, and enzyme activity measurements.
Automatically generated - may contain errors
Lab products found in correlation
Firely renilla dual luciferase assay, by Biotium (2 mentions)
K2 transfection reagent, by Biontex (2 mentions)
Glo lysis buffer, by Promega (2 mentions)
Nanoglo reagent, by Promega (2 mentions)
Flexi rabbit reticulocyte lysate system, by Promega (2 mentions)
Mouse anti flag antibody, by Merck Group (2 mentions)
Adcc reporter bioassay, by Promega (1 mentions)
Dual glo luciferase assay, by Promega (1 mentions)
Firefly and renilla dual luciferase assay kit, by Biotium (1 mentions)
Lipofectamine 3000, by Thermo Fisher Scientific (1 mentions)
Dual luciferase reporter assay system, by Promega (1 mentions)
8 protocols using glomax 96 plate reader
1
Luciferase Assay for TTF-1 Transactivation
2
Luciferase Reporter Assay for p75NTR Promoter Analysis
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Dual‐Luciferase Reporter Assay System was applied for luciferase reporter assay. First, we cloned the 2 kb sequences of p75NTR promoter from human genomic DNA into plasmid pGL4.10 (Promega, #E6651) by PCR assay. Truncated promoter fragments were generated by FL promoter via PCR assay. Second, in luciferase reporter assay, 1 × 105 HKE293T cells were plated onto 24‐well plates before transfection. The next day, cells were co‐transfected with a firefly luciferase reporter vector based on control pcDNA3.1 empty vector (EV, 225 ng) or wild‐type CASZ1 plasmid (225 ng), and a henifella luciferase control vector pGL4.73 (50 ng, Promega). After transfection, cell lysates were prepared and the values of firefly/Henilla luciferase were quantified using the Firely and Renilla dual luciferase assay (Biotium, #30005‐2) on a Glomax 96‐plate reader (Promega).
3
In vitro Translation of nLuc Reporter
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In vitro translation reactions using the Flexi Rabbit Reticulocyte Lysate (RRL) System (Promega), were performed with nLuc reporter mRNAs and analyzed as in Kearse et al., 2016. Briefly, luciferase assays were performed using a mixture of prepared NanoGlo reagent (Promega) and in vitro reactions diluted in Glo Lysis Buffer at 25μL:25μL (Promega), and quantified using a GloMax-96 plate reader (Promega). For immunoblot analysis, 10μL RRL reactions were performed with saturating mRNA levels, mixed with 40μL of SDS sample buffer and heated at 70° for 15 minutes, and then resolved using SDS-PAGE. nLuc-3xFLAG reporter proteins were detected by immunoblot using the anti-FLAG antibody (mouse, Sigma, F1804).
4
ADCC Bioassay for HIV-1 gp140 Evaluation
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The ADCC Reporter Bioassay was performed according to the manufacturer's protocol (Promega), using soluble HIV‐1 UG37 gp140 (CFAR, UK) instead of target cells. 0.08, 0.4, 2 and 10 μg/mL antibody was pre‐incubated with free HIV‐1 gp140 (5 μg/mL) for 1 h. ADCC Bioassay Effector cells (Jurkat cells stably expressing the high‐affinity (V158) FcγRIIIa receptor and an NFAT response element driving expression of firefly luciferase) were added at 1.5 × 105 cells/well, and the assay plates were incubated at 37 °C and 5% CO2 for 24 h. Bio‐Glo™ luciferase assay reagent was added and luminescence measured in a GloMax 96 plate reader (Promega). In this assay, luciferase activity in the effector cells is directly proportional to FcγRIIIa activation.
5
Promoter Reporter Assay in A549 Cells
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Promoter reporter assays were carried out in 24-well plates using A549 parental cells as previously described11 (link)15 (link). Briefly, cells were co-transfected with Firefly luciferase reporter construct and Renilla luciferase control vector pGL4.10 (Promega). Firefly and Renilla luciferase values were measured 48 hours after transfection using Firefly and Renilla Dual Luciferase Assay Kit (Biotium #30005-1) or Dual-Glo luciferase assay (Promega #PAE2920) on a GloMax 96 plate reader (Promega). Renilla luciferase signals were normalized to Firefly luciferase signals.
6
In vitro Translation of nLuc Reporter
7
Luciferase Assay for TTF-1 Transactivation
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A549 cells (1×105) were plated in a 24-well plate before transfection. Next day, cells were co-transfected with the control pcDNA3.1 empty vector (EV, 225 ng) or the wild-type TTF-1 plasmid (225ng)-based firefly luciferase reporter plus the renilla luciferase control vector pGL4.73 (50 ng, Promega) using the K-2 transfection reagent (Biontex, #T060). After transfection (48 hr later), cell lysates were prepared and firefly/renilla luciferase values were quantified using Firely & Renilla Dual Luciferase assay (Biotium, #30005-2) on a GloMax-96 plate reader (Promega). Firefly luciferase values were normalized to Renilla luciferase values and expressed as relative values. See Supplementary Information for additional experimental methods on RNA profiling, cholesterol efflux, intracellular cholesterol quantitation and animal studies.
8
Foxq1 Regulation of EGFR Expression
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Before transfection, 293T cells (1 × 105 cells) were plated in a 24-well plate for 24 h and co-transfected with the control pcDNA3.1 plasmid (2.0 µg/ml) or the Foxq1 plasmid (2.0 µg/ml) together with the control vector pGL3 (2.0 µg/ml) or the EGFR plasmid (2.0 µg/ml) using Lipofectamine 3000 (L3000008 ,Thermo Fisher Scientific, USA). After 36 h incubation, cell lysates were prepared and firefly/renilla luciferase values were quantified using the Dual-Luciferase Reporter Assay System (E1910, Promega, USA) on a GloMax-96 plate reader (Promega).
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