Deleterious effects of calcium indicators within cells; an inconvenient truth

[Display omitted] •Ca2+ signalling is widely studied using fluorescent indicators that can be readily loaded into living cells.•Chemical Ca2+ indicators can measure cellular Ca2+ changes over long periods of time with sub-second resolution and microscopic precision.•There are known problems with the use of chemical Ca2+ indicators, which are largely overlooked.•Genetically-encoded Ca2+ indicators overcome some of the issues of using chemical Ca2+ indicators, but may introduce problems of their own.•Careful planning and control experiments may overcome the deleterious, off-target effects of chemical Ca2+ indicators. The study of cellular Ca2+ signalling is indebted to Roger Tsien for the invention of fluorescent indicators that can be readily loaded into living cells and provide the means to measure cellular Ca2+ changes over long periods of time with sub-second resolution and microscopic precision. However, a recent study [1] reminds us that as useful as these tools are they need to be employed with caution as there can be off-target effects. This article summarises these recent findings within the wider context of confounding issues that can be encountered when using chemical and genetically-encoded Ca2+ indicators, and briefly discusses some approaches that may mitigate against misleading outcomes.