For luciferase assay, as described previously (7 (link)), cells were maintained in RPMI with 10% FBS and seeded at 1 × 105 cells/well in 12-well plates 1 day prior to the assay. The reporter constructs, pGL3 basic, pGL3-NANOG, or pGL3-NANOG E2F1 Mut together with pCMV-β-Gal, an internal control for transfection efficiency, were cotransfected into CaSki cells using Lipofectamine 2000 (Invitrogen). After 24 hours, cells were washed with PBS and lysed with Cell Culture Lysis Reagent (Promega). Luciferase activity was measured with a Turner Biosystems TD-20/20 luminometer after addition of 40 µL of luciferase assay reagent (Promega). β-Galactosidase activity was measured with a uQuant microplate reader (BioTek) at 570 nm wavelength after addition of β-galactosidase assay reagent containing 1 mmol/L chlorophenol red β-d-galactopyranoside substrate (Roche). Relative luciferase activity was normalized to the β-galactosidase activity in the cell lysate and was calculated as an average of three independent experiments.
Turner biosystems td 20 20 luminometer
Manufactured by Promega
Sourced in United States
The Turner Biosystems TD-20/20 luminometer is a laboratory instrument designed to measure luminescence. It provides quantitative analysis of light-emitting chemical reactions, which can be used to detect and quantify various biomolecules or cellular processes.
Automatically generated - may contain errors
Lab products found in correlation
Lipofectamine 2000, by Thermo Fisher Scientific (2 mentions)
Chlorophenol red β d galactopyranoside substrate, by Roche (1 mentions)
Luciferase assay reagent, by Promega (1 mentions)
Cell culture lysis reagent, by Promega (1 mentions)
Uquant microplate reader, by Agilent Technologies (1 mentions)
Pierce renilla luciferase flash assay kit, by Thermo Fisher Scientific (1 mentions)
Luciferase assay system kit, by Promega (1 mentions)
Deae dextran, by Merck Group (1 mentions)
Polybrene, by Merck Group (1 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (1 mentions)
Cyclosporine, by Merck Group (1 mentions)
Enliten atp assay, by Promega (1 mentions)
Hmgb1 elisa kit 2, by Shino-Test (1 mentions)
4 protocols using turner biosystems td 20 20 luminometer
1
Luciferase Assay for NANOG Transcription
2
Measuring TCF-4 Promoter Activity in Breast Cancer Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
MCF-7 and T47-D cells were seeded in 24-well plates with 1×105 cells/well, and only cells growing in the log phase at a passage number ≤15 were used. Cells were transfected with Let-7c mimic and Renilla luciferase plasmid containing a TCF-4-responsive promoter when at 50–70% confluence ratio using Lipofectamine® 2000 (Invitrogen; Thermo Fisher Scientific, Inc.), non-transfected and non-treated cells were set as control, as described previously (9 (link),11 (link)). Each group included three replicates, and independent triplicate experiments were performed. Specifically, the cells were collected with cell lysis buffer after transfection for 24 h, and Renilla luciferase activity was detected using the Pierce®Renilla Luciferase Flash Assay kit (no. 16164; Thermo Fisher Scientific, Inc.), following the manufacturer's instructions. Cell lysate (20 μl) and 50 μl working solution were added to one column, and a Turner Biosystems TD-20/20 luminometer (Turner Biosystems; Thermo Fisher Scientific, Inc.) was used to detect the light output (prime the pump prior to injections with working solution). Firefly luciferase activity was normalized to Renilla luciferase activity.
3
Transduction Efficiency of Macrophages
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The supernatant from transfected 293T cells was mixed with complete mBMM culture media at an approximate ratio of 2:1 and was further supplemented with 6 μg/ml polybrene (Sigma-Aldrich), 10 μg/ml sterile filtered DEAE-dextran (Sigma-Aldrich) or 10 μM cyclosporine (Sigma-Aldrich). To achieve a MOI of 10, 6 ml of this infection media was used for macrophages in 10 cm dishes and 15 ml for the cells in 175 cm2 culture flasks. After 24 hours infection media was removed and complete mBMM media added. The cells were allowed to recover for 24 hours after which the infection was repeated in selected groups (single or double infection). Three days after the second infection the GFP expression was determined by fluorescence microscopy and flow cytometry. The FLUC activity was determined using a Luciferase Assay System kit (Promega, Madison, WI) and a Turner BioSystems Luminometer TD-20/20 (Promega). The FLUC activity was normalized to the amount of total protein measured using a Pierce BCA Protein Assay Kit (Thermo Scientific). The viability of the reporter macrophages was assessed three days after the second infection by flow cytometry and propidium iodide staining
4
Extracellular ATP and HMGB1 Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The supernatants of RH2-infected SCCVII cells and uninfected control cells were collected; and cell debris was removed by centrifugation at 6200 r. p.m. for 5 min. Secreted extracellular ATP in the supernatants was measured with the ENLITEN ATP assay (Promega, Madison, WI, USA) according to the manufacturer's protocol, using a Turner Biosystems luminometer (TD-20/20; Promega). Supernatants were also used to detect HMGB1 with HMGB1 ELISA Kit II (Shino-Test, Kanagawa, Japan). Enzyme-linked immunosorbent assay was performed according to the manufacturer's protocol outlined for the normal sensitivity format of the assay; and the plates were read on a microplate spectrometer at a wavelength of 450 nm.
About PubCompare
Our mission is to provide scientists with the largest repository of trustworthy protocols and intelligent analytical tools, thereby offering them extensive information to design robust protocols aimed at minimizing the risk of failures.
We believe that the most crucial aspect is to grant scientists access to a wide range of reliable sources and new useful tools that surpass human capabilities.
However, we trust in allowing scientists to determine how to construct their own protocols based on this information, as they are the experts in their field.
Ready to get started?
Sign up for free.
Registration takes 20 seconds.
Available from any computer
No download required
Revolutionizing how scientists
search and build protocols!