Hey everyone! I need to inhibit transport in ovarian cell culture line for the following live cell microscopy. Does anyone know sufficient time needed to use for Brefeldin before ABs staining? and is it necessary to wash it off before ABs?
Would be so much grateful for your advices!
Dear Daria Valeryevna Savenkova ,
If you are stimulating your ovarian cells with certain stimulants than you must add BFA at the same time you add stimulant.
Otherwise 3-4 hour is sufficient for transport blocking by BFA.
If you are using any stimulant then yes, you need to wash cells before antibody staining. But if you are not using any stimulant then you can directly add antibody.
Hope this is helpful.
Depending on type and duration of stimulation, it can be different. Prolonged Brefeldin A incubation may reduce viability, however it again depends on the cell type. Here's some thoughts: Article The Duration of Antigen-Stimulation Significantly Alters the...
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