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C11 BODIPY 581/591

Catalog No.: GC40165

C11-BODIPY581/591 is a fluorescent ratio-probe of lipid oxidation.

C11 BODIPY 581/591 Chemical Structure

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1mg
$145.00
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Sample solution is provided at 25 µL, 10mM.

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Protocol

Cell experiment [1]:

Cell lines

microalgal cells (unicellular green alga Chlamydomonas reinhardtii); mammalian cells

Preparation method

The stock solution of C11-BODIPY581/591 was prepared by dissolving 1 mg of the product in DMSO to a concentration of 1mM. Stock solution can be stored in freezer (-5 to -30°C) and protect from light.

Reaction Conditions

2.5 µM C11-BODIPY581/591 for 30 min. The excitation and emission band of oxidized type is pass of 460–495 and 510–550, respectively. But the excitation and emission band of reduced type is pass of 565–581 and 585–591, respectively.

Applications

C11-BODIPY591/581 staining was combined with flow cytometry measurements to detect reactive oxygen species (ROS) and oxidative stress damage in living cells and membrane systems. Oxidation of the polyunsaturated butadienyl portion of the dye results in a shift of the fluorescence emission peak from ~590 nm to ~510 nm.

Cell experiment [2][3]:

Cell lines

Rat-1 fibroblasts

Preparation Method

Rat-1 fibroblasts were cultured at 37℃ in DMEM supplemented with 7.5% fetal calf serum in a 7.5% CO2 humidified atmosphere. Small unilamellar vesicles of egg PC were made through ethanolic injection. The C11-BODIPY581/591 concentration of the stock solutions was determined by measuring the absorption at 581 nm using a molar extinction coefficient of 139 444 l mol-1 cm-1.

Reaction Conditions

Cells were incubated for 30 min at 37℃ with C11-BODIPY581/591 (1μM) in growth medium. Prior to use, fibroblasts were rinsed with enriched phosphate buffered saline (PBS+, 137 mM NaCl, 2.7 mM KCl, 0.5 mM MgCl2, 0.9 mM CaCl2, 1.5 mM KH2PO4, 8.1 mM Na2HPO4 and 5 mM glucose, pH 7.4). Oxidation of C11-BODIPY581/591 was induced by incubating cells with 50μM CumOOH at 37℃ with or without hemin in PBS+.

Applications

C11-BODIPY581/591 is oxidized only when CumOOH and hemin are present. The oxidation rate correlates directly with CumOOH concentration, up to 0.2 mM. Administration of C11-BODIPY581/591 dissolved in fetal calf serum to the growth medium of rat-1 fibroblasts leads to a rapid incorporation into cellular membranes. When the ratio of C11-BODIPY581/591 to cellular phospholipid is 1:13 047±734, it causes the minimal perturbation of the membranes and the absence of spectroscopic artifacts after 30 min of labeling.

References:

[1]. Giulia Cheloni, Vera I. Slaveykova. Optimization of the C11-BODIPY581/591 Dye for the Determination of Lipid Oxidation in Chlamydomonas reinhardtii by Flow Cytometry. Cytometry A,2013 Oct;83(10):952-61..

[2]. De Wit, R. et al. Large Scale Screening Assay for the Phosphorylation of Mitogen-Activated Protein Kinase in Cells. Journal of Biomolecular Screening, 1998,3(4), 277–284.

[3]. Pap EH, et al. Ratio-fluorescence microscopy of lipid oxidation in living cells using C11-BODIPY581/591. FEBS Lett. 1999 Jun 25;453(3):278-82.

Background

C11-BODIPY581/591 is an oxidation-sensitive fluorescent fatty acid analogue with fluorescent properties in the red range of the visible spectrum (emission maximum 595 nm), allowing its application in fluorescence microscopy. C11-BODIPY581/591 is easily incorporating into membranes and fluoresces red in the intact state but shifts to green upon free radical-induced oxidation. This characteristic is highly advantageous, it makes the ratio-imaging of oxidant activities at the (sub)cellular level feasible. In addition, the fluorescent properties of C11-BODIPY581/591 allow the use of this probe in fast- and medium- throughput screening of antioxidants in living cells and model membranes in a multiwell/fluorescence reader approach[1][2].

The wavelengths of maximal excitation and emission of fluorophore C11-BODIPY581/591 corresponded to 581 and 591 nm, respectively. Addition of CumOOH/hemin, as an initiator of lipid oxidation, shifted the excitation and emission spectra to shorter wavelengths corresponding to green fluorescence (peak excitation 500 nm, emission 510 nm). C11-BODIPY581/591 is also easily oxidized by other hydroxy-, peroxy- and oxy-radical generating systems such as hydrogen peroxide/Fe2+ and 2,2’-azobis. However, this probe is relatively insensitive to SIN-1, which generates nitric oxide and superoxide[3]

References:
[1]. Drummen GP, et al. C11-BODIPY581/591, an oxidation-sensitive fluorescent lipid peroxidation probe: (micro)spectroscopic characterization and validation of methodology. Free Radic Biol Med. 2002 Aug 15;33(4):473-90.
[2]. Partyka A, et al. Detection of lipid peroxidation in frozen-thawed avian spermatozoa using C11-BODIPY581/591. Theriogenology. 2011 Jun;75(9):1623-9.
[3]. Pap EH, et al. Ratio-fluorescence microscopy of lipid oxidation in living cells using C11-BODIPY581/591. FEBS Lett. 1999 Jun 25;453(3):278-82.

Chemical Properties

Cas No. 217075-36-0 SDF
Synonyms N/A
Chemical Name (T-4)-difluoro[5-[[5-[(1E,3E)-4-phenyl-1,3-butadien-1-yl]-2H-pyrrol-2-ylidene-κN]methyl]-1H-pyrrole-2-undecanoato(2-)-κN1]-borate(1-), monohydrogen
Canonical SMILES [F-][B+3]1([N]2=C(/C=C/C=C/C3=CC=CC=C3)C=CC2=CC4=CC=C(CCCCCCCCCCC([O-])=O)[N-]14)[F-].[H+]
Formula C30H34BF2N2O2 • H M.Wt 504.4
Solubility 30mg/ml in DMSO,Slightly soluble in Methanol Storage Store at -20°C
General tips For obtaining a higher solubility , please warm the tube at 37 ℃ and shake it in the ultrasonic bath for a while.Stock solution can be stored below -20℃ for several months.
Shipping Condition Evaluation sample solution : ship with blue ice
All other available size: ship with RT , or blue ice upon request

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Research Update

Optimization of the C11-BODIPY(581/591) dye for the determination of lipid oxidation in Chlamydomonas reinhardtii by flow cytometry

Cytometry A2013 Oct;83(10):952-61.PMID: 23943236DOI: 10.1002/cyto.a.22338

Lipid oxidation is a recognized end point for the study of oxidative stress and is an important parameter to describe the mode of micropollutant action on aquatic microorganisms. Therefore, the development of quick and reliable methodologies probing the oxidative stress and damage in living cells is highly sought. In the present proof-of-concept work, we examined the potential of the fluorescent dye C11-BODIPY(591/581) to probe lipid oxidation in the green microalga Chlamydomonas reinhardtii. C11-BODIPY(591/581) staining was combined with flow cytometry measurements to obtain multiparameter information on cellular features and oxidative stress damage within single cells. First, staining conditions were optimized by exploring the capability of the dye to stain algal cells under increasing cell and dye concentrations and different staining procedures. Then lipid oxidation in algae induced by short- and long-term exposures to the three metallic micropollutants, copper, mercury, and nanoparticulate copper oxide, and the two organic contaminants, diethyldithiocarbamate (DDC) and diuron was determined. In this work we pointed out C11-BODIPY(591/581) applicability in a wide range of exposure conditions, including studies of oxidation as a function of time and that it is suitable for in vivo measurements of lipid oxidation due to its high permeation and stability in cells and its low interference with algal autofluorescence. © 2013 International Society for Advancement of Cytometry.

Prolonged lipid oxidation after photodynamic treatment. Study with oxidation-sensitive probe C11-BODIPY581/591

FEBS Lett2005 Feb 14;579(5):1255-60.PMID: 15710422DOI: 10.1016/j.febslet.2005.01.024

Photodynamic treatment (PDT) is an emerging procedure for the therapy of cancer, based on photosensitizers, compounds that generate highly reactive oxygen species on illumination with visible light. Photodynamic peroxidation of cellular lipids is a consequence of PDT associated with cytolethality. We used chloromethyl dichlorodihydrofluorescein diacetate and a novel fluorescent ratiometric oxidation-sensitive probe, C11-BODIPY581/591 (C11-BO), which reports on lipid peroxidation, for visualizing oxidative stress in cells subjected to PDT with a phthalocyanine photosensitizer Pc4. With C11-BO loaded into the cells before or immediately after PDT, we observed a prolonged oxidation, which continued up to 30 min after illumination. In contrast, H2O2 caused oxidation of C11-BO only when the cells were in direct contact with H2O2. PDT-induced oxidative stress was most pronounced in vesicular perinuclear organelles, most likely photodamaged lysosomes. We hypothesize that the lysosomal localization of the prolonged oxidative stress is a consequence of the presence of redox-active iron in lysosomes. In conclusion, we have found that oxidative stress induced in cells by PDT differs from one induced by H2O2 in respect of induction of prolonged oxidation of lipids.

Supra-physiological doses of testosterone affect membrane oxidation of human neutrophils monitored by the fluorescent probe C₁₁-BODIPY⁵⁸¹/⁵⁹¹

Eur J Appl Physiol2013 May;113(5):1241-8.PMID: 23160653DOI: 10.1007/s00421-012-2538-y

The purpose of this study was to determine the effects of supra-physiological doses of testosterone (TES) on membrane oxidation of activated human neutrophils in vitro using an innovative and sensitive technique: the real-time detection with the fluorescence probe C11-BODIPY(581/591). Methodological controls were performed with the lipid-soluble and powerful antioxidant astaxanthin at different neutrophil density cultures. Neutrophils from nine healthy young men (23.4 ± 2.5 years, 174.4 ± 7.0 cm height, and 78.3 ± 7.0 kg weight) were isolated and treated with 0.1 or 10 μM TES for 24 h and subsequently labeled with the free radical-sensitive probe C11-BODIPY(581/591) for monitoring membrane oxidation after neutrophil activation with phorbol-12-myristate-13-acetate (PMA). First-order exponential decay kinetic indicated that both 0.1 and 10 μM TES severely increased baseline membrane oxidation in non-activated human neutrophils (compared to control). However, similar kinetics of membrane oxidation were observed in control and 0.1 μM TES-treated neutrophils after PMA activation, whereas chemical activation did not alter the baseline higher rates of membrane oxidation in 10 μM TES-treated neutrophils. The data presented here support the hypothesis that TES exerts distinct effects on the membrane oxidation of human neutrophils, depending on its dose (here, 10(2) to 10(4)-fold higher than physiological levels in men) and on PMA activation of the oxidative burst. Furthermore, this paper also presents an innovative application of the free radical-sensitive probe C11-BODIPY(581/591) for monitoring (auto-induced) membrane oxidation as an important parameter of viability and, thus, responsiveness of immune cells in inflammatory processes.

Detection of lipid peroxidation in equine spermatozoa based upon the lipophilic fluorescent dye C1l-BODIPY581/591

J Androl2002 Mar-Apr;23(2):259-69.PMID: 11868820DOI: 10.1139/apnm-2019-0966

The lipophilic fluorescent probe, 4,4-difluoro-5-(4-phenyl-1 ,3-butadienyl)-4-bora-3a,4a-diaza-s-indacene-3-undecanoic acid (C11-BODIPY581/591) was used to evaluate changes in lipid peroxidation in equine spermatozoa during both short-term exposure to ferrous sulfate and sodium ascorbate in the presence of cumene hydroperoxide as well as during storage of spermatozoa at 5 degrees C for 48 hours. Peroxidation of C11-BODIPY581/591 was accompanied by a shift in fluorescence from red to green, and the relative amount of nonoxidized probe was determined as the ratio of red:(red + green) fluorescence as detected by either fluorescence microplate reader or by flow cytometry. The addition of Fe2SO4 (0 to 0.5 mM), low concentrations of sodium ascorbate, and the addition of cumene hydroperoxide increased peroxidation of C11-BODIPY581/591. The addition of high concentrations (10 or 20 mM) of sodium ascorbate or alpha-tocopherol reduced peroxidation of C11-BODIPY581/591 during short-term incubations. During storage at 5 degrees C in a skim milk-based extender, equine spermatozoa demonstrated a progressive decline in motility and a small but significant increase in lipid peroxidation based upon ratiometric analysis of C11-BODIPY581/591. The addition of Fe2SO4 increased lipid peroxidation in cooled spermatozoa in a dose-dependent fashion and decreased sperm motility. The addition of alpha-tocopherol, however, did not reduce lipid peroxidation during cooled semen storage. These data demonstrate that the lipophilic fluorescent probe C11-BODIPY581/591 is a useful measurement of lipid peroxidation in equine spermatozoa and that there is an increase in lipid peroxidation during cooled storage of equine spermatozoa that is increased in the presence of ferrous promoters.

Lipid droplets are both highly oxidized and Plin2-covered in hepatocytes of diet-induced obese mice

Appl Physiol Nutr Metab2020 Dec;45(12):1368-1376.PMID: 32585124DOI: 10.1139/apnm-2019-0966

Chronic high-fat diet feeding is associated with obesity and accumulation of fat in the liver, leading to the development of insulin resistance and nonalcoholic fatty liver disease. This condition is characterized by the presence of a high number of intrahepatic lipid droplets (LDs), with changes in the perilipin pattern covering them. This work aimed to describe the distribution of perilipin (Plin) 2, an LD-associated protein involved in neutral lipid storage, and Plin5, which favors lipid oxidation in LD, and to evaluate lipid peroxidation through live-cell visualization using the lipophilic fluorescent probe C11-BODIPY581/591 in fresh hepatocytes isolated from mice fed a high-fat diet (HFD). Male C57BL/6J adult mice were divided into control and HFD groups and fed with a control diet (10% fat, 20% protein, and 70% carbohydrates) or an HFD (60% fat, 20% protein, and 20% carbohydrates) for 8 weeks. The animals fed the HFD showed a significant increase of Plin2 in LD of hepatocytes. LD from HFD-fed mice have a stronger lipid peroxidation level than control hepatocytes. These data provide evidence that obesity status is accompanied by a higher degree of lipid peroxidation in hepatocytes, both in the cytoplasm and in the fats stored inside the LD. Novelty Our study shows that lipid droplets from isolated hepatocytes in HFD-fed mice have a stronger lipid peroxidation level than control hepatocytes. C11-BODIPY581/591 is a useful tool to measure the initial level of intracellular lipid peroxidation in single isolated hepatocytes. Perilipins pattern changes with HFD feeding, showing an increase of Plin2 covering lipid droplets.

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