The AldeRed ALDH Detection Kit provides cancer and stem cell scientists with new capabilities for live cell isolation and characterization. The AldeRed reagent is a red-shifted fluorescent substrate for aldehyde dehydrogenase (ALDH), allowing cells to be identified and isolated by flow cytometry with concurrent use of green fluorescent cell lines, antibodies, transgenic animals and reporter assays.
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High ALDH activity serves as a universal marker of stem cells, both normal and malignant. Cells can be identified and isolated based upon the enzymatic activity of ALDH, a detoxifying enzyme responsible for oxidation of hazardous aldehyde byproducts. The marker ALDH has been used to isolate cancer stem cells from various human malignancies including bladder, breast, cervical, colon, head and neck, liver, lung, pancreas, prostate and ovary. The AldeRed assay employs a fluorescent and non-toxic ALDH substrate (AldeRed 588-A) that diffuses freely into intact and viable cells, but remains trapped inside the cells once converted by ALDH into the corresponding acid. The amount of fluorescence produced is proportional to the ALDH activity in the cells and is measured by flow cytometry, allowing fluorescence-activated cell sorting (FACS). This kit supplies the ALDH inhibitor diethylaminobenzaldehyde (DEAB), which is used in negative control testing necessary for background fluorescence assessement.
• For optimal excitation and emission of AldeRed 588-A, use a combination of blue laser (488 nm) or green laser (532 nm) with PE-Texas Red detector (615 nm). Detection in proximal measurement channels (e.g. PE) is also possible, but may affect delta MFI (mean fluorescence intensity) between Test sample and DEAB-treated control sample.
• All live intact cells (even with low ALDH expression) become fluorescent upon exposure to activated AldeRed 588-A. The generation of a DEAB-treated control is necessary for each cell sample to establish baseline fluorescence.
• Handle all reagents using aseptic techniques.
Stem Cell Research
Apoptosis & Cancer
1. AldeRed Reagent: (Part No. CS216596). One (1) vial containing 50 µg of AldeRed reagent powder.
2. DEAB Reagent: (Part No. CS216595). One (1) vial containing 1 mL of DEAB (Diethylaminobenzaldehyde) reagent.
3. Hydrochloric Acid (2N): (Part No. CS216594). One (1) vial containing 1.5 mL of 2 Normal hydrochloric acid.
4. Dimethylsulphoxide: (Part No. CS216593). One (1) vial containing 1.5 mL of Dimethylsulphoxide (DMSO).
5. AldeRed Assay Buffer: (Part No. CS216592). Four (4) bottles containing 25 mL of AldeRed Assay Buffer each.
6. Verapamil: (Part No. CS220012). Four (4) vials containing 615 µg Verapamil powder each.
• Each lot is tested by measuring ALDH activity in SK-BR-3 cells by flow cytometry.
• AldeRed® Reagent is 95% pure (PMR)
Storage and Stability
• Store kit at 2-8°C.
• Reconstituted and activated AldeRed Reagent may be stored in aliquots at -20°C protected from light for up to 1 year. Repeated freezing and thawing is not recommended.
• AldeRed Assay Buffer supplemented with Verapamil may be stored at 2-8°C in aliquots for up to 3 months.
ALDERED is a registered trademark of Merck KGaA, Darmstadt, Germany
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