Cell Meter™ Fluorimetric Cellular Lipid Peroxidation Assay Kit

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Telephone	1-800-990-8053
Fax	1-800-609-2943
Email	sales@aatbio.com
International	See distributors
Bulk request	Inquire
Custom size	Inquire
Shipping	Standard overnight for United States, inquire for international

Storage, safety and handling

H-phrase	H303, H313, H333
Hazard symbol	XN
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R22
UNSPSC	12352200

Overview SDS Protocol

Platform

Flow cytometer

Excitation	488 nm laser
Emission	530/30 nm, 575/26 nm filter
Instrument specification(s)	FITC channel

Fluorescence microscope

Excitation	490 nm (FITC) and 545 nm (TRITC)
Emission	530 nm (FITC) and 600 nm (TRITC)
Recommended plate	Black wall/clear bottom
Instrument specification(s)	FITC and TRITC channels

Components

Example protocol

AT A GLANCE

Protocol summary

Plate cells at desired confluency.
Treat cells with compound of interest and incubate.
Add Lipoxite™ R590/G525.
Remove media, and wash with PBS.
Analyze sample by fluorescence microscope or flow cytometer through FITC/TRITC or FITC/PE channels.

Important

Thaw one of each kit component at room temperature before starting the experiment.

PREPARATION OF WORKING SOLUTION

Prepare 10X working solution of Lipoxite™ R590/G525 by making 1:50 dilution of 500X Lipoxite™ R590/G525 (Component A) into HHBS (Component B)

For guidelines on cell sample preparation, please visit
https://www.aatbio.com/resources/guides/cell-sample-preparation.html

SAMPLE EXPERIMENTAL PROTOCOL

Grow cells at desired density and incubate overnight in a humidified chamber at 37°C with 5% CO₂.
Treat cells with test compounds as desired. Note: For a positive control, add hydrogen peroxide (Component C) to the cells at a final concentration of ~250 µM (1X) for 30 minutes.
Add 10X Lipoxite™ R590/G525 to the cells at a final concentration of 1X (for example add 10 uL to 90 uL of the cells).
Incubate the cells for 30 min at 37°C with 5% CO₂cell incubator.
Remove media and wash cells with HHBS (Component B) or DPBS for three times.
Monitor fluorescence of cells with a fluorescence microscope or flow cytometer through FITC/TRITC or FITC/PE channels within 2 hours of staining.

Images

Figure 1. HeLa cells were stained with 1X Lipoxite™ R590/G525 for 30 mins in complete growth medium at 37°C. For H₂O₂ treatment, approximately 250 µM of H₂O₂ were added to the cells and incubated for 30 mins. The cells were then incubated with 1X Lipoxite™ R590/G525, and stained with Hoechst 33342 during the last 10 mins of incubation. The cells were washed 3 times with HHBS and imaged with a Keyence fluorescent microscope. With H₂O₂ treatment, a clear shift of fluorescence signal of red to green was observed.

Figure 2. Jurkat cells were stained with 1X Lipoxite™ R590/G525 for 30 mins in complete growth medium at 37°C. For H₂O₂ treatment, approximately 250 µM of H₂O₂ were added to the cells and incubated for 30 mins. The cells were then incubated with 1X Lipoxite™ R590/G525, and analyzed with a flow cytometer through FITC (488/530 nm) and PE (488/572 nm) channels. The data are represented as the ratios of red (PE)/green (FITC) fluorescence intensities. The ratio of red/green decreases in H₂O₂ treated cells indicating the presence of H₂O₂ induced lipid peroxidation.

Citations

View all 19 citations: Citation Explorer

PDT-Enhanced Ferroptosis by a Polymer Nanoparticle with pH-Activated Singlet Oxygen Generation and Superb Biocompatibility for Cancer Therapy
Authors: Li, Jing and Li, Junhua and Pu, Yuji and Li, Sai and Gao, Wenxia and He, Bin
Journal: Biomacromolecules (2021): 1167--1176

Using fluorescence-activated flow cytometry to determine reactive oxygen species formation and membrane lipid peroxidation in viable boar spermatozoa
Authors: Guthrie, H. D., Welch, G. R.
Journal: Methods Mol Biol (2010): 163-71

Use of fluorescence-activated flow cytometry to determine membrane lipid peroxidation during hypothermic liquid storage and freeze-thawing of viable boar sperm loaded with 4, 4-difluoro-5-(4-phenyl-1,3-butadienyl)-4-bora-3a,4a-diaza-s-indacene-3-undecanoic
Authors: Guthrie, H. D., Welch, G. R.
Journal: J Anim Sci (2007): 1402-11

Fluorescence microplate reader measurement of tissue susceptibility to lipid peroxidation
Authors: Zhang, J. G., Fariss, M. W.
Journal: Curr Protoc Toxicol (2004): Unit17 3

Response to ozone in two lettuce varieties on chlorophyll a fluorescence, photosynthetic pigments and lipid peroxidation
Authors: Calatayud, A., Barreno, E.
Journal: Plant Physiol Biochem (2004): 549-55

Fluorescence imaging of lipid peroxidation in isolated rat lungs during nonhypoxic lung ischemia
Authors: Matot, I., Manevich, Y., Al-Mehdi, A. B., Song, C., Fisher, A. B.
Journal: Free Radic Biol Med (2003): 785-90

Effect of radiation induced lipid peroxidation on diphenylhexatriene fluorescence in egg phospholipid liposomal membrane
Authors: P, undefined and ey, B. N., Mishra, K. P.
Journal: J Biochem Mol Biol Biophys (2002): 267-72

Micellar electrokinetic chromatography separation and laser-induced fluorescence detection of the lipid peroxidation product 4-hydroxynonenal
Authors: Claeson, K., Thorsen, G., Karlberg, B.
Journal: J Chromatogr B Biomed Sci Appl (2001): 133-8

Chlorophyll a fluorescence, antioxidant enzymes and lipid peroxidation in tomato in response to ozone and benomyl
Authors: Calatayud, A., Barreno, E.
Journal: Environ Pollut (2001): 283-9

A fluorescence method for the determination of plasma susceptibility to lipid peroxidation
Authors: Cervato, G., Viani, P., Cazzola, R., Cestaro, B.
Journal: Clin Biochem (1999): 171-7