A total of 10,000 cells were plated into each well of a white-bottom 96-well plate (Corning, New York, NY, USA). The cells were treated with varying concentrations (1, 5, 15, or 30 μM) of REV1 inhibitor drugs (JH-RE-06, JH-RE-06.NaOH, drug 4, drug 5, drug 6) with either 0, 1, or 4 Gy of ionizing radiation. After 24, 48, or 72 h of incubation, the proportion of viable cells was evaluated by adding 100 µL of the CellTiter-Glo Luminescence stain (Promega, Madison, WI, USA) was added into each well. The CellTiter Glo stain was prepared according to the manufacturer’s recommendation. The endpoint luminescence was measured on the Synergy H1 Microplate Reader plate reader. Relative cell viability was determined by dividing treated sample luminescence measurements by their respective control samples without drugs and/or ionizing radiation.
White bottom 96 well plate
Manufactured by Corning
Sourced in United States
The White-bottom 96-well plates are a laboratory equipment product designed for a variety of cell-based assays. The white-colored bottom of the plates provides enhanced signal-to-noise ratio and improved sensitivity for luminescent and fluorescent applications.
Automatically generated - may contain errors
Lab products found in correlation
Synergy h1 microplate reader, by Synergy Software (1 mentions)
Dual glo luciferase assay, by Promega (1 mentions)
Bio glo luciferase reagent, by Promega (1 mentions)
Adcc reporter bioassay kit, by Promega (1 mentions)
Firefly luciferase substrate, by Promega (1 mentions)
Tristar lb 941, by Berthold Technologies (1 mentions)
Celltiter glo, by Promega (1 mentions)
6 protocols using white bottom 96 well plate
1
Cell Viability Evaluation of REV1 Inhibitors and Radiation
2
Rapid Dinucleotide Extraction and Quantification
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
All dinucleotides (NAD+, NADH, NADP+, NADPH) were extracted using a custom protocol from Promega. In brief, 300 µl high pH Bicarbonate Buffer + 1% DTAB was added to cell pellets to lyse the cells. The lysate was split into two 100 uL aliquots, where one was treated with 100 µl 0.4N HCl to one tube for acid treatment. Both aliquots were heated at 60°C for 15 min and then cooled at RT for 10 min. The acid treated aliquot was then neutralized with 100 µl 0.5 M Trizma Base to get oxidized forms of NAD and NADP. The base-only treated sample was neutralized with 200 µl 0.4 NHCl/0.5 M Trizma Base to get reduced forms of NADH and NADPH.
After dinucleotide extraction, extracts were used in the corresponding GloTM Assay (Promega) using the standard protocol where 50 uL of extract was added with 50 uL of GloTM Detection Reagent (Promega) in a white bottom 96 well plate (Corning). After 30 minutes of incubation the plates were measured for luminescence using a Glomax Navigator with dual injection pumps, model GM2010 luminometer.
After dinucleotide extraction, extracts were used in the corresponding GloTM Assay (Promega) using the standard protocol where 50 uL of extract was added with 50 uL of GloTM Detection Reagent (Promega) in a white bottom 96 well plate (Corning). After 30 minutes of incubation the plates were measured for luminescence using a Glomax Navigator with dual injection pumps, model GM2010 luminometer.
3
Dual-Reporter Luciferase Assay Protocol
4
ADCC Assay for Monoclonal Antibodies
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
ADCC capacity of each MAb was measured using an ADCC Reporter Bioassay kit (Promega) largely in line with the manufacturer’s instructions. In brief, 3 × 104 Vero.E6 cells per well were seeded in white-bottom 96-well plates (Corning) and infected with VSV-ANDV in MEM at an MOI of 1 16 h later. After 2 days of incubation with virus at 37°C, 3-fold serial dilutions of each MAb were prepared in MEM, starting with 90 μg/ml. A negative-control antibody (KL-2G12) was used as well for determining the background. These MAbs were then added to the infected plate with effector cells from the Promega kit at a ratio of 3:2 (effector cells to infected cells) and additional medium such that the effective starting concentration of each MAb was 30 μg/ml. The plates were then incubated at 37°C for 6 h. Bio-Glo Luciferase reagent (Promega) was then added, and luminescence was measured immediately.
5
TZM-bl Infectivity Assay for Viral Supernatant
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
TZM-bl cells were plated at 1 x 105 cells / 100 μL RPMI + 10% fetal calf serum / well into 48 well plates and infected with 100 μL viral supernatant with 16 μg / ml DEAE-Dextran and 10 nM of the protease inhibitor saquinavir. Plates were spinoculated for 1200 x g for 2 hours at room temperature and infection was allowed to proceed for 48 hours at 37°C for 48 hours. Culture media was then removed, and cells were resuspended in 200 μL firefly luciferase passive lysis buffer (Promega). Lysate was then placed into white-bottom 96 well plates (Corning) at 20 μL per well. Luciferase readings were performed by dispensing 25 μL of firefly luciferase substrate (Promega) and measuring the subsequent bioluminescent output via a TriStar LB 941 multimode reader (Berthold, Bad Wildbad, Germany). Relative light units were then normalized to an efavirenz uninfected control and then to reverse transcriptase units / ml viral supernatant.
6
Vero Cell Cytotoxicity Assay for Ricin
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Vero cell cytotoxicity assays were performed as described [29] (link). In brief, Vero cells were trypsinized, adjusted to ∼5 ×104 cells per mL and seeded (100 µl/well) into white bottom 96-well plates (Corning Life Sciences, Corning, NY), and allowed to adhere overnight. Vero cells were then treated with ricin (0.01 µg/mL; 154 pM), ricin:Ab mixtures, or medium alone (negative control) for 2 h at 37°C. Cells were washed to remove non-internalized toxin or ricin:Ab mixtures, and 100 µl of fresh medium was added to the wells. Fresh medium was allowed to incubate for 48 h and cell viability was measured using CellTiter-Glo (Promega, Madison, WI). All samples were performed in quadruplicate and 100% viability was defined as the average value obtained from wells in which cells were treated with medium only.
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!