AO/EB Staining Kit Cat.No.: E607308 Package: 100 Tests/200 Tests Description Acridine orange (AO) is a nucleic acid selective fluorescent cationic dye. It is cell-permeable, and will stain both live and dead cells. AO is commonly used for fluorescence microscopy and flow cytometry analysis of cellular physiology and cell cycle status. This cell-permeant cellular stain can be utilized in conjunction with a number of other staining solutions. Under the fluorescent microscope: Live cells will appear uniformly green; Apoptotic cells will stain green and contain bright green dots in the nuclei as a consequence of chromatin condensation and nuclear fragmentation; Necrotic cells will stain orange, but the fluorescent is weak or even disappear. EB can stain only cells that have lost membrane integrity. Combined with EB, necrotic cells stain orange, but have a nuclear morphology resembling that of viable cells, with no condensed chromatin. Then normal cells, apoptotic cells and necrotic cells can be distinguished by using this AO/EB staining kit. Kit Components Component 100 Tests 200 Tests Component A: AO Staining Solution 0.5 ml 1 ml Component B: EB Staining Solution 0.5 ml 1 ml Component C: 10X Buffer 10 ml 20 ml Protocol 1 1 Storage Store at 2~8°C and protect from light. Reagent is stable for at least one year. Procedure Note: The follow procedure is adapted for most cell types. 1. Dilute 10X Buffer (Component C) with distilled water to 1X Buffer. 2. Wash cells twice with phosphate buffer saline (PBS) and re-suspend cells in desired volume of 1X Buffer. Note: Adherent cells in culture may be stained in situ on cover slips or in the cell culture wells. 3. Add 5 μl AO Staining Solution (Component A) and 5 μl EB Staining Solution (Component B) to 90 μl cell suspension, and mix gently. 4. Incubate 1~5 mSin at rooangonm temperature, protected from light. Biotech Note: Each cell line should be evaluated on an individual basis to determine the optimal concentration and incubation time for staining. 5. Analyze staining cells by florescent microscope or flow cytometry. WARNING! Before use, please read this protocol carefully. For your safety, please use gloves, goggles or other protective measures. This kit is for R&D use only, not for drug, household or other uses. .