A novel Ca2+ indicator for long-term tracking of intracellular calcium flux.

Abstract

The major drawback of using Fluo-4AM is that it requires an organic anion transporter inhibitor, such as probenecid, to prevent leakage. This can hinder the studies that require extended monitoring time and longer cellular retention. To address the issue, a novel Ca2+indicator, Calbryte 520AM, was developed. We compared the performance of Fluo-4AM and Calbryte 520AM following prolonged incubation periods aftercell loading. Cells loaded with Calbryte 520AM retained the dye for up to 24hwhile exhibiting significant fluorescence brightness and superior Fmax/F0 ratios (Fmax: fluorescence intensity upon stimulation; F0: intensity before stimulation). It demonstrated that the longer retention of Calbryte 520AM can be exploited to accommodate for the extended time required when monitoring calcium dynamics.