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CELLESTIAL® Assays
Mito-ID cell staining is too strong
Mitochondrial membrane potential dye is too concentrated for the cell type used. Optimize for different cell types by diluting the…
Tue, Jul 18, 2023
Fluorescence does not increase after incubation with inhibitor.
Cells do not express any MDR1, MRP1/2 or BCRP. Use a positive control cell type expressing corresponding ABC transporter.
Tue, Jul 18, 2023
Healthy cells show very high fluorescence intensity
PMT setting is too high. Lower PMT setting on flow cytometer. Incubation temperature. Incubation with the dye should be at…
Tue, Jul 18, 2023
Cells do not exhibit fluorescence after incubation with the detection reagent.
Cell viability is low. Cells should be in log growth phase. Cell samples cannot be kept longer than 6 hours…
Tue, Jul 18, 2023
There are differences in MFI (mean fluorescence intensity) values between cell lines.
Detection reagent accumulation may be influenced by cell size, endogenous esterase activity, etc. Using the inhibitors and calculating the MAF…
Tue, Jul 18, 2023
Inconsistent fluorescence shift using the same cell (irreproducible results).
Inadequate incubation condition. Always use a water bath (not incubator) and ensure temperature of water bath is 37°C.
Tue, Jul 18, 2023
High baseline fluorescence
Growth medium and organic anion transport contribute to high baseline fluorescence. Remove the medium before adding indicator dye to the wells.…
Tue, Jul 18, 2023
Untreated cells have calcium response
Inconsistent DMSO concentration. Ensure that buffer used for the negative control wells has the same final concentration of DMSO as…
Tue, Jul 18, 2023
Well-to-well variation observed
Incorrect dispensing and/or experimental setup parameter used. A fluorescence microplate reader with automatic dispensing should be used for assays requiring precise…
Tue, Jul 18, 2023
Fluorescence drop upon compound addition
Cells may become dislodged during addition. Decrease the rate of addition or seed fewer cells per well.
Tue, Jul 18, 2023