After cells were seeds onto a 24-well plate at a density of 105 cells/well, mitochondrial membrane potential was measured as described previously [34 (link)35 (link)]. Briefly, at the indicated time points, cells were incubated with 200 nM tetramethylrhodamine ethyl ester perchlorate (TMRE) in 0.1% DMSO for 20 minutes at 37℃. After washing six times with PBS, 1% Triton X-100 was added to each well. TMRE intensity for 200 µl cell lysate on a Nunc 96-well black plate (Thermo Fisher Scientific) was then quantified with the SpectraMax i3 plate reader (Molecular Devices) using 549 nm for excitation and 575 nm for emission.
Nunc 96 well black plate
Manufactured by Thermo Fisher Scientific
Sourced in United Kingdom
The Nunc 96-well black plate is a laboratory equipment used for various microplate-based assays. It is a standard 96-well microplate with a black-colored polystyrene surface. The black color minimizes background fluorescence, making it suitable for fluorescence-based experiments.
Automatically generated - may contain errors
5 protocols using nunc 96 well black plate
1
Quantifying Mitochondrial Membrane Potential
2
Quantifying ROS Production using DCFDA
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
An oxidative sensitive dye 2′,7′-dichlorodihydrofluorescein diacetate (DCFDA) was used to measure ROS production as described previously [33 (link)]. Briefly, HK-2 cells were seeded onto a 24-well plate at a density of 105 cells/well and treated with or without H2O2 plus/minus CsA as indicated. After that, they were incubated with 20 µM DCFDA for 45 minutes at 37℃. After washing twice with PBS, 1% Triton X-100 was added to each well. Then 2′,7′-dichlorofluorescein intensity of 200 µl of cell lysate on a Nunc 96-well black plate (Thermo Fisher Scientific) was quantified with a SpectraMax i3 plate reader (Molecular Devices) using 485 nm for excitation and 535 nm for emission.
3
Antioxidant Capacity Measurement by ORAC
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The ORAC assay was performed as described by Huang et al. (2002b) (link). Twenty-five microliters of sample extract or standard solution were placed in a Nunc 96-well black plate (Thermo Fisher Scientific, Leicestershire, UK), 150 µL of fluorescein solution (1.2 × 10-8 mM in 75 mM phosphate buffer, pH 7.42) was added, and the plates were incubated for 30 min at 37 °C. The assay was initiated by adding 25 µL AAPH solution (15 mM in 75 mM phosphate buffer, pH 7.42). Fluorescence was recorded every minute for one hour at an excitation wavelength of 485 nm and an emission wavelength of 515 nm using the microplate reader previously described. The net area under the curve (net-AUC) in relation to the blank control was calculated (Prior et al., 2005 (link)). Trolox was diluted in pure ethanol and results were expressed as Trolox equivalents per gram of FW (μM TEq/g FW).
4
MALAT1 RNA Binding Assay
5
Monitoring Amyloid Fibril Formation
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Synthetic peptides that correspond to the fourth and fifth repeats of CsgA, CsgAR4-5, and CsgAR5-4N122A described previously42 (link) were synthesized by Biosynthesis Inc. Polymerization of the synthetic peptides was previously described43 (link). To monitor the fibrillization of the synthetic CsgA peptide, 100 μM CsgAR4-5, or 100 μM CsgAR5-4N122A was mixed with an equal volume 10 μM ThT in the presence or absence of 0.5 mg/ml mAb 3H3 or control antibody 6A in a black Nunc 96-well plate (Fisher). The plate was sealed with an optical plate cover (Eppendorf). Fluorescence of ThT (excitation 440 nm/emission 490 nm) was monitored at 37 °C using a BMG Labtech POLARstar Omega plater reader. Readings were taken at 8-min intervals for 36 h. Lag time (t0) was calculated using the following formula: t0 = t1/2 − 2te, where t1/2 is the time required to reach half the maximum fluorescence intensity and te is the elongation time43 (link).
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!