Lipid peroxide was detected using the Image-iT™ Lipid Peroxidation Kit, which is based on the lipophilic BODIPY®581⁄591 C11 probe (Thermo Fisher Scientific) (Kim et al., 2020 (link)). After treatment as indicated, 1 µM probe was added and incubated for a further 30 min at 37°C. Cells were collected by trypsinization and the fluorescence with excitation/emission at 488/525 nm was measured from 5,000 cells using a Guava® easyCyte flow cytometer and analyzed with InCyte2.6 software (Luminex, TX, USA). Mean fluorescence intensity of C11-BODIPY were calculated from independent experiments.
Bodipy 581 591 c11 probe
Manufactured by Thermo Fisher Scientific
Sourced in United States
The BODIPY 581/591 C11 probe is a lipid peroxidation indicator. It is a fluorescent dye that can be used to detect and quantify lipid peroxidation in cells and tissues.
Automatically generated - may contain errors
Lab products found in correlation
H2dcfda, by Thermo Fisher Scientific (4 mentions)
Sytox green, by Thermo Fisher Scientific (3 mentions)
Image it lipid peroxidation kit, by Thermo Fisher Scientific (2 mentions)
Facscalibur flow cytometer, by BD (2 mentions)
Pierce bca protein assay kit, by Thermo Fisher Scientific (2 mentions)
Accuri c6 flow cytometer, by BD (2 mentions)
Varioskan lux, by Thermo Fisher Scientific (1 mentions)
Calcein am, by Yeasen (1 mentions)
Cm h2dcfda probe, by Thermo Fisher Scientific (1 mentions)
Annexin 5 pi, by Keygen Biotech (1 mentions)
Cell counting kit 8 (cck8), by Dojindo Laboratories (1 mentions)
Vectashield hardset with dapi, by Vector Laboratories (1 mentions)
Tcs sp8 confocal microscope, by Leica (1 mentions)
Fluorescence microplate reader, by Agilent Technologies (1 mentions)
Fer 1, by Targetmol (1 mentions)
Mouse spleen lymphocyte separation kit, by Solarbio (1 mentions)
Dnase 1, by Merck Group (1 mentions)
Liberase, by Merck Group (1 mentions)
Collagenase type 4, by Merck Group (1 mentions)
Lovastatin, by MedChemExpress (1 mentions)
38 protocols using bodipy 581 591 c11 probe
1
Lipid Peroxidation Detection in Cells
2
Quantitative Assessment of Cell Viability and Oxidative Stress
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Cell viability was detected by Cell Counting Kit-8 (CCK-8) kits. HepG2 cells were seeded onto cultured in 96-well plates at 1 × 104 cells per well. After treatment with or without HG, CCK-8 reagents were added to the plates and then incubated at 37°C for 2 h. A multifunctional microplate reader measured the absorbance at 450 nm (Varioskan LUX, Thermo Fisher Scientific).
HepG2 cells were seeded onto 6 cm cell culture dishes at 2 × 105 cells. Calcein-AM (40719ES50, YEASEN Biotechnology) was applied to measure the intracellular labile iron pool (LIP) [28 (link)]. After treatment, cells were digested and incubated with 0.25 μM Calcein-AM at 37°C for 15 min. Washing with PBS, the fluorescence intensity (Ex/Em = 490 nm/515 nm) was quantified by a multifunctional microplate reader microplate. The intracellular lipid ROS levels were detected by BODIPY 581/591 C11 probe (D3861, Thermo Fisher Scientific) [29 (link)]. Cells were incubated with a 2 μM probe for 37°C for 15 min in the dark. After washed with PBS, the fluorescence spectrum was measured by a multifunctional microplate reader microplate.
HepG2 cells were seeded onto 6 cm cell culture dishes at 2 × 105 cells. Calcein-AM (40719ES50, YEASEN Biotechnology) was applied to measure the intracellular labile iron pool (LIP) [28 (link)]. After treatment, cells were digested and incubated with 0.25 μM Calcein-AM at 37°C for 15 min. Washing with PBS, the fluorescence intensity (Ex/Em = 490 nm/515 nm) was quantified by a multifunctional microplate reader microplate. The intracellular lipid ROS levels were detected by BODIPY 581/591 C11 probe (D3861, Thermo Fisher Scientific) [29 (link)]. Cells were incubated with a 2 μM probe for 37°C for 15 min in the dark. After washed with PBS, the fluorescence spectrum was measured by a multifunctional microplate reader microplate.
3
Apoptosis and Oxidative Stress Assays
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Annexin V/PI (KeyGEN, Nanjing, China) was used for the detection of cell apoptosis induced by oxaliplatin. The CM-H2DCF-DA probe (C6827, Thermo Fisher Scientific) and BODIPY™ 581/591 C11 probe (D3861, Thermo Fisher Scientific) were added, and the mixture was incubated in the dark at 37 °C for 30 min to measure the cellular ROS and cellular lipid ROS levels. All these measurements were conducted with a flow cytometer (Beckman Coulter, CA, USA) according to the manufacturer’s instructions. The gating strategy for ROS/lipid ROS analysis is provided in Supplementary Fig. 3f .
4
Hydrophobic Bis-Pyrene Unit Synthesis and Cell Studies
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The hydrophobic bis‐pyrene unit (BP‐COOH) was synthesized by Apeptide Biotech Co., Ltd (shanghai, China) according to a previous report.[29 ] Murine mammary carcinoma (4T1), HUVECs, and L929 cell lines were purchased from the MEIXUAN Biological science and technology Co., Ltd. Nile red (NR) was purchased from Shanghai Macklin Biochemical Co., Ltd. (China). Recombinant human caspase‐3 protein was purchased from Sino Biological, Inc. (China). BODIPY‐581/591C11 probe was purchased from Thermo Fisher Scientific Co., Ltd (Carlsbad, California, U.K.). Propidium iodide(PI), calcein AM, and Cell Counting Kit‐8 were purchased from Dojindo (Tokey, Japan). FITC‐CD11c+, PE‐CD80+, and APC‐CD86+ antibodies were purchased from Biolegend, (California, MA). TNF‐α, IFN‐γ, and IL‐6 ELISA kits were purchased from Uscn Life Science (China).
5
Intracellular and Lipid ROS Measurement
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Intracellular ROS and lipid ROS levels were measured using previously described methods (Dixon et al., 2012). Briefly, after the treatment with or without PQ, intracellular levels of ROS and lipid ROS were examined using 2′,7′-dichlorofluorescein diacetate (H2DCF-DA, Thermo Fisher Scientific) and the BODIPY 581/591 C11 probe (Thermo Fisher Scientific), respectively. Cells were incubated with DCFH-DA (20 μM) or BODIPY 581/591 C11 (10 μM) for 30 min at room temperature in the dark, washed three times with PBS, and evaluated using an inverted microscope (IX83, Olympus, Japan) or a FACS Calibur flow cytometer (BD Biosciences, CA, USA) to measure ROS production.
6
Visualizing Lipid Oxidation in Tissue
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
10 μm tissue cryosections were fixed in 4% paraformaldehyde, blocked with 10% normal serum, and permeabilized with 0.2% Tween 20. Tissue sections were stained with 10 μM BODIPY 581/591 C11 probe (Thermo Fisher, D3861). After 30 min in the dark, the slides were mounted with Vectashield Hardset with DAPI (Vector Laboratories) and viewed under a Leica TCS SP8 confocal microscope (Leica Microsystems). Each wound section was imaged from edge to edge with consecutive frames.
7
Lipid Peroxidation Measurement by BODIPY 581/591 C11
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
BODIPY 581/591 C11 probe (D3861; Thermo Fisher Scientific) was used to detect lipid peroxidation according to the manufacturer’s instructions. Briefly, cells were incubated with BODIPY 581/591 C11 at a final concentration of 5 μmol/L for 30 minutes at 37°C and washed 3 times with phosphate-buffered saline. Oxidation of the polyunsaturated butadienyl portion of the dye resulted in a shift of the fluorescence emission peak from approximately 590 nm to approximately 510 nm, which was measured in a Bio-Tek fluorescence microplate reader (Winooski, VA).
8
Lipid Peroxidation Quantification by Flow Cytometry
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Lipid peroxide was detected using the Image-iT™ Lipid Peroxidation Kit, which is based on the lipophilic BODIPY®581⁄591 C11 probe (Thermo Fisher Scientific, Waltham, MA, USA). After treatment as indicated, 1 µM of the probe was added and the solution was incubated for 30 min at 37 °C. Cells were collected by trypsinization and the fluorescence from 5000 cells was measured with the excitation/emission set at 488/525 nm using a Guava® easyCyte flow cytometer. Data were analyzed using InCyte2.6 software (Luminex, Austin, TX, USA).
9
Immune Profiling and Ferroptosis Assay
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
Lovastatin (catalog HY-N0504) was purchased from MedChemExpress. Recombined active IFN-γ protein (catalog KGH2016) was purchased from KeyGEN (Nanjing, China). Mouse spleen lymphocyte separation kit (catalog P8860) and mouse tumor–infiltrating tissue lymphocyte separation kit (catalog P9000) were purchased from Solarbio. Liberase (catalog 5401020001), DNase I (catalog DN25), and type IV Collagenase (catalog C5138) were purchased from MilliporeSigma. The PD-1 in vivo mAb (catalog BE0273) was purchased from BioXCell. The inhibitor for ferroptosis, fer-1 (catalog T6500), was purchased from TargetMol. The BODIPY 581/591 C11 probe (catalog D3861) was purchased from Thermo Fisher Scientific. Antibodies used in the study and their source were exhibited in Supplemental Table 2 .
10
Quantifying Lipid Peroxidation via BODIPY C11
Check if the same lab product or an alternative is used in the 5 most similar protocols
+ Expand
The RSL3-induced lipid peroxidation was measured by using the BODIPY 581/591 C11 probe (Thermo Fisher Scientific). Briefly, cells plated in 6-well plates were loaded with 10 μM BODIPY C11 at 37°C for 30 minutes and then treated with RSL3 and ferrostatin-1 for indicated times. After treatment, cells were harvested by trypsinization and resuspended in PBS. The cell suspension was loaded into 96-well black-walled plates and measured the fluorescence signal using a microplate reader. Lipid peroxidation was determined by calculating the ratio of oxidized (Ex/Em = 485:535 nm) to reduced (Ex/Em = 560/591 nm) C11 fluorescence signal.
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!