A viability assay was carried out using PI/FDA staining. First 20 μl PI (propidium iodine solution, 1 mg/ml, Sigma) and 10 μl FDA (fluorescein diacetate solution 1 mg/ml, Sigma) were added to ELS and incubated at room temperature for 90 s. Next the ELS were washed once in PBS (Invitrogen) and then florescence at 617 nm (excitation) and 520 nm (emission) measured, with 1 s and 150 ms exposure for PI and FDA staining respectively. The total FDA intensity was compared to the total PI plus FDA intensity using Nikon imaging software, giving both a cell membrane integrity and metabolic viability read-out.
Fluorescein diacetate solution
Manufactured by Merck Group
Fluorescein diacetate solution is a chemical reagent used in various laboratory applications. It is a colorless, non-fluorescent compound that can be used as a substrate for the detection of enzymatic activity, particularly esterase enzymes. When the diacetate compound is hydrolyzed by esterases, it releases the fluorescent compound fluorescein, which can be detected using fluorescence-based techniques.
Automatically generated - may contain errors
Lab products found in correlation
3 protocols using fluorescein diacetate solution
1
Cell Viability Assay: PI/FDA Staining
2
Cell Viability Assay for Cryopreserved Cells
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A viability assay was carried out using PI/FDA staining. 20 μl PI (propidium iodine solution, 1 mg/ml, Sigma) and 10 μl FDA (fluorescein diacetate solution 1 mg/ml, Sigma) were added to ELS and incubated at room temperature for 90 s. The ELS were washed once in PBS (Invitrogen) and then florescence at 617 nm (excitation) and 520 nm (emission) measured, with 1 s and 150 ms exposure respectively. The total FDA intensity was compared to the total PI plus FDA intensity using Nikon imaging software, giving both a cell membrane integrity and metabolic viability read-out. This was carried out at 6, 24, 48, and 72 h post-thaw. The 6 h timepoint was chosen as this was the minimum time required to fully remove residual (pre-freeze) FDA-sensitive enzymes from non-viable cells.
3
Fluorescent Viability Assay for Cells
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A viability assay was carried out using PI/FDA staining. 20μl PI (propidium iodine solution, 1mg/ml, Sigma) and 10μl FDA (fluorescein diacetate solution 1mg/ml, Sigma) were added to ELS and incubated at room temperature for 90 seconds. The ELS were washed once in PBS (Invitrogen) and then florescence at 617 nm (excitation) and 520 nm (emission) measured, with 1 s and 150 ms exposure for PI and FDA staining respectively. The total FDA intensity was compared under a phase-contrast microscope to the total PI plus FDA intensity using Nikon imaging software giving a cell membrane integrity and metabolic viability read-out [4 (link), 6 (link), 31 (link)].
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