Protocol: Seed a dark clear-bottomed 96-well microplate with 50,000 cells per well then culture overnight under standard conditions using standard media. Freshly prepare a working solution of H2DCFDA by diluting the 20mM stock in 1x assay buffer to a 20µM working concentration (10µL sto...
看了一些protocol,但是都没有具体说到背景值需要去掉哪些,如何去计算。 另一个问题是,单位如何设置更合理呢?文献上有的用relative fluorescence unit,没有说跟什么相对,我想大概是酶标仪给出的读书吧;有的用的是H2O2的系列浓度作为标准去衡量ROS的产生。 两个问题,困惑了很久,希望大家提供宝贵的意见。欢迎探讨!谢谢...
FunctionalStudies-DCFDA-Cellular ReactiveOxygenSpeciesDetection AssayKit(ab113851) JurkatcellswerelabeledwithDCFDA(20 µM)orunlabeled(none)andthenculturedan additional3hourswithorwithout50µMtert- butylhydrogenperoxide(TBHP)accordingto theprotocol.Cellswerethenanalyzedona fluorescentplatereader.Mean+/-standard...
The DCFDA assay protocol is based on the diffusion of DCFDA / H2DCFDA / DCFH-DA / DCFH into the cell. It is then deacetylated by cellular esterases to a non-fluorescent compound, which is later oxidized by ROS into 2’, 7’ –dichlorofluorescein (DCF). DCF is highly fluorescent and ...
DCFDA Assay Kit ab113851 includes DCFDA reagent, buffer, TBHP positive control, and a full protocol, for a fast way to set up a DCFDA assay on live cells. DCFDA taken up by cells is oxidized by reactive oxygen species with fluorescent readout (Ex/Em 485/535 nm) by microscopy, flow cyt...
This article introduces the detailed operation protocol and attentions for ROS detection using 2′,7′-dichlorodi-hydrofluorescein diacetate (H2DCFDA) fluorescent probe based on flow cytometry and confocal microscope. These methods can be used to detect ROS levels and distribution in model plant ...
Cells constantly generate reactive oxygen species (ROS) during aerobic metabolism. The ROS generation plays an important protective and functional role in the immune system. The cell is armed with a powerful antioxidant defense system to combat excessive production of ROS. Oxidative stress occurs in ...
Protocol Cell experiment [1, 2]: Cell lines hCECs Preparation Method Telomerase-immortalized human corneal epithelial cells (hCECs) were cultured at 37℃ under 5% CO2 atmosphere in bronchial epithelium growth medium supplemented with 5 mg/mL insulin, 0.5 mg/mL hydrocortisone, a mixture of 50 mg/...
The protocol I have used for ROS detection by this dye is straightforward: 1) Stable cells derived from MDA-MB-231 mammary carcinoma cells were plated in 6 well plates at 3 x 105in full serum media 2) Approx. 24 hours later the cells were treated with 10 uM Hydrogen Peroxide (...
In Vitro: Guidelines (Following is our recommended protocol. This protocol only provides a guideline, and should be modified according to your specific needs). ?1. H2DCFDA is dissolved in DMSO to obtain a 10 mM stock solutions and further diluted before use. ?2. Cells are incubated with 5...