What is the purpose of adding Brefeldin A to cells being tested in flow cytometry experiments? Please explain using an example.

Brefeldin A (BFA) is a lactone antibiotic that is often used in cell biology experiments to study protein transport. In the context of flow cytometry, BFA is commonly used to inhibit protein transport from the endoplasmic reticulum to the Golgi apparatus. This is particularly useful when studying the intracellular accumulation of proteins.

For example, in an experiment studying the production of a specific cytokine (a type of protein) by immune cells, BFA can be added to the cell culture. The BFA will prevent the cytokine from being transported out of the cell, causing it to accumulate within the cell. The cells can then be stained with a fluorescent antibody that binds to the cytokine, and analyzed using flow cytometry. The amount of fluorescence detected in each cell is proportional to the amount of cytokine that the cell has produced. This allows researchers to quantify cytokine production by different types of immune cells, or under different experimental conditions.