The IFRS1 cells were reseeded into 8-well Ibidi μ-Slides (Ibidi, Grafelfing, Germany) (5 × 104 cells/well) and cultured in a humidified atmosphere containing 5% CO2 at 37 °C for 12 h before a treatment to identify the prophylactic effect of DHA on tBHP-induced ROS production. The IFRS1 cells were pretreated with 10 μM DHA for 12 h. Intracellular ROS were measured using a ROS assay kit (Dojindo, Kumamoto, Japan). The DHA pretreated cells were washed with a Hanks Balanced Salt Solution (HBSS; FUJIFILM Wako Pure Chemical, Tokyo, Japan). A DCFH-DA working solution was added to the cells and incubated for 30 min, followed by a treatment with 50 μM tBHP or HBSS alone for 30 min. After washing with HBSS, the fluorescence intensity was measured by a fluorescence microscope (IX70; Olympus, Tokyo, Japan) and quantified using ImageJ software version 1.53m (NIH, Bethesda, MD, USA).
Ros assay kit
Manufactured by Dojindo Laboratories
Sourced in Japan
The ROS assay kit is a laboratory tool designed to measure the levels of reactive oxygen species (ROS) in biological samples. It provides a quantitative assessment of ROS, which are highly reactive molecules that can cause oxidative stress and cellular damage.
Automatically generated - may contain errors
Lab products found in correlation
Bz x700, by Keyence (2 mentions)
μ slide, by Ibidi (1 mentions)
Elispot analyzer, by Cellular Technology (1 mentions)
Concanavalin a (cona), by Cayman Chemical (1 mentions)
Lab tek chamber slide, by Thermo Fisher Scientific (1 mentions)
Hoechst 33342, by Dojindo Laboratories (1 mentions)
Hanks balanced salt solution (hbss), by Fujifilm (1 mentions)
Hanks balanced salt solution (hbss), by Keyence (1 mentions)
Porphyromonas gingivalis lps, by Merck Group (1 mentions)
Dcfh da dye working solution, by Dojindo Laboratories (1 mentions)
Rpmi 1640, by Merck Group (1 mentions)
Glomax multi detection system, by Promega (1 mentions)
Anhydrous ethanol, by Merck Group (1 mentions)
In vitro toxicology assay kit, by Merck Group (1 mentions)
Penicillin streptomycin solution, by Thermo Fisher Scientific (1 mentions)
Wst 8, by Dojindo Laboratories (1 mentions)
Antibiotic antimycotic solution, by Thermo Fisher Scientific (1 mentions)
Hydrochloric acid (hcl), by Merck Group (1 mentions)
H2o2 aqueous solution, by Merck Group (1 mentions)
Graphite gt powder, by Merck Group (1 mentions)
8 protocols using ros assay kit
1
DHA Attenuates tBHP-induced ROS
2
Characterizing Tumor-Infiltrating CD8+ T Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Tumors were minced and dissociated using the Tumor dissociation kit. Tumor‐infiltrating CD8+ T cells were purified with antibody‐coated magnetic beads (CD8 [TIL] MicroBeads mouse, Miltenyi Biotec). ELISpot assays were performed as previously described.19 Briefly, tumor‐infiltrating CD8+ T cells were sorted and plated at 6 × 104 cells/well in 96‐well plates coated with anti‐interferon (IFN)‐γ antibodies (AN18; Mabtech). Tumor‐infiltrating CD8+ T cells were stimulated with BNL‐T (2 × 104 cells), medium alone (negative control), or concanavalin A (positive control; Cayman Chemical). Each condition was tested in duplicate wells. Spots were counted using computer‐assisted image analysis (ELISpot analyzer; Cellular Technology). The total production of ROS was assessed using a ROS assay kit (Dojindo). Briefly, tumor cells were sorted to isolate tumor‐infiltrating CD8+ T cells. ROS production was determined using a fluorescence microscope (BZ‐X700; Keyence).
3
ROS Measurement in Cultured Cells
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
ROS levels were evaluated using a ROS Assay Kit (R252; Dojindo, Kumamoto, Japan). The cells were plated in an 8-well 0.8 cm2 Lab-Tek chamber slide (177445PK; Thermo Fisher Scientific). After 24 h, the cells were washed with phenol red-free Hanks’ balanced salt solution (HBSS, 084-08965; Wako) and incubated in DCFH-DA solution and 0.5 μg/mL Hoechst 33342 (H342; Dojindo, Tokyo, Japan) for 30 min at 37 °C with 5% CO2. Thereafter, the cells were washed with HBSS and treated with H2O2 and IL/TNF for 30 min. After washing with HBSS, the fluorescence signal was detected using a fluorescence microscope (BZ-X700, KEYENCE).
4
Measuring ROS Induced by P. gingivalis LPS
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The production of ROS induced by Porphyromonas gingivalis LPS was evaluated following the ROS assay kit (Dojindo, Kumamoto, Japan) manual. Briefly, 1 × 105 BPH-1 cells/well were seeded into 6-well plates with RPMI-1640 (Hyclone, USA), containing 10% fetal bovine serum (Hyclone, USA), and cultured at 37°C for 24 h. The medium was replaced with RPMI-1640 containing 1 μg/mL Porphyromonas gingivalis LPS (Sigma, SMB00610) and cultured at 37°C for a further 24 h. The supernatant was removed, and highly sensitive DCFH-DA dye working solution (Dojindo, Kumamoto, Japan) was added then incubated at 37°C for 30 min. The changes in the levels of ROS were detected using flow cytometry.
5
Progranulin Regulates Myoblast ROS
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Primary myoblasts were cultured at 1 3 10 3 cells/well on a 96-well plate in DMEM-LG supplemented with 20% fetal calf serum, basic FGF. After 12 h, half of the medium was changed and myoblasts were cultured in the presence or absence of recombinant mouse Progranulin. After 24-72 h, the medium was discarded, the cells were washed twice with PBS, and 100 mL of the solution in the ROS assay kit (Dojindo, Kumamoto, Japan) was added to the cells. Thirty minutes after incubation in the dark, the cells were washed twice with PBS, and fluorescence was measured using a GloMax multi+ Detection System (Promega, Tokyo, Japan).
6
Quantifying Intracellular ROS Levels
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The intracellular ROS levels were measured using a ROS assay kit (Dojindo, Japan). The kit contains highly sensitive 2′,7′-dichlorofluorescein diacetate (DCFH-DA), which is more susceptible to oxidation to fluorescent dichlorofluorescein (DCF) by intracellular ROS than normal DCFH-DA. Thus, the ROS level can be quantified by the fluorescence of DCF. At 24 h after light exposure, the cells were incubated with highly sensitive DCFH-DA for 20 min at 37°C and observed by fluorescence microscopy or measured at 488 nm excitation and 525 nm emission using a fluorescence microplate reader.
7
Cytotoxicity Evaluation of Nanomaterials
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Penicillin-streptomycin solution, trypsin-EDTA solution, RPMI 1640 medium, and 1% antibiotic-anti-mycotic solution were obtained from Life Technologies/Gibco (Grand Island, NY, USA). Vanillin, fetal bovine serum, and the in vitro toxicology assay kit were purchased from Sigma-Aldrich (St. Louis, MO, USA). Graphite (Gt) powder, NaOH, KMnO4, NaNO3, anhydrous ethanol, 98% H2SO4, 36% HCl, 30% H2O2 aqueous solution, and all other chemicals were purchased from Sigma-Aldrich unless otherwise stated. The cell viability assay WST-8, ROS assay kit, 2′,7′-dichlorofluorescin diacetate, and lactate dehydrogenase (LDH) cytotoxicity detection kit were purchased from Dojindo (Kumamoto, Japan), Sigma, and Takara (Shiga, Japan), respectively.
8
Evaluating Hepatocyte Apoptosis and Oxidative Stress
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Paraformaldehyde-fixed cell cultures were stained for DNA fragmentation using the DeadEnd Fluorometric TUNEL System (Promega, Madison, WI, USA) and an anti-albumin antibody (Abcam). Hepatocytes were distinguished from ASCs/ASCL in the co-culture group as albumin-positive stained cells. ROS was detected in cell cultures using an ROS assay kit (Dojindo, Tokyo, Japan) according to the manufacturer’s protocol.
The liver tissue of each group was fixed using 4% paraformaldehyde phosphate buffer solution at 4°C. The sections were cut to a thickness of 4 μm and stained with hematoxylin and eosin for histological examination. The TUNEL assay was performed using the DeadEnd Fluorometric TUNEL System (Promega).
The liver tissue of each group was fixed using 4% paraformaldehyde phosphate buffer solution at 4°C. The sections were cut to a thickness of 4 μm and stained with hematoxylin and eosin for histological examination. The TUNEL assay was performed using the DeadEnd Fluorometric TUNEL System (Promega).
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!