Is probenecid required for calcium assays?

In CHO (Chinese Hamster Ovary) cell calcium flux assays, the presence of high organic anion transporter activity can result in poor retention of fluorescent calcium dyes, such as Fluo-3 and Fluo-4 within the cells. To enhance the retention of these dyes and achieve optimal signal sensitivity, it is necessary to include probenecid in the assay protocol. Probenecid is a compound which inhibits organic anion transporters and prevents the expulsion of fluorescent dyes from the cells. In a standard calcium flux assay for CHO cells using Fluo-3, Fluo-4, or similar commercial kits, a concentration of 2.5 mM probenecid is recommended. It is important to note that probenecid is not required when using certain types of assay kits (e.g. BD™ PBX Calcium Assay Kit).