Calcium chelators

Chelators are compounds that bind to metal ions forming a complex and some of them are also fluorescent. If the complex has fluorescence properties different from those of the free chelator, it can be used as an ion indicator.

Calcium chelators are able to bind calcium (usually in a relationship one to one) in a selective way (they have higher affinity for calcium than for any other metal ions). Binding to calcium is performed through carboxylic groups, so it can be affected by pH, other ions or co-ordination to proteins, lipids, etc.

This process is a reversible equilibrium:

{Chelator-Ca}complex <===> Chelator + Ca2+

 

Calcium, chelator and complex concentration are related by an equilibrium constant called dissociation constant, Kd, that is defined as:

Kd = ([Ca2+] [Chelator] ) / [{Chelator-Ca}complex]

 

When Kd is very low, it is a high-affinity chelator (it has a high tendency to bind calcium). If Kd is high (µM or higher), we talk about low-affinity chelators.

Optimum calcium concentration range for an indicator is between 0,1 · Kd < [Ca2+] < 10 · Kd. Nearly all chelator is forming the fluorescent complex at high calcium concentration, so no variations are observed in fluorescence intensity. At lower [Ca2+] , there is a very low indicator response.

High-affinity chelators trap calcium very efficiently, so high intracellular chelator concentration can buffer calcium response. This attribute is used in the case of the non-fluorescent BAPTA (Kd = 190 nm if there is no Mg2+) to make intracellular calcium concentration zero and obtain Fmin. A high concentration of BAPTA is loaded along with the desired calcium indicator. In these conditions, all intracellular calcium is sequestered by BAPTA, so fluorescence in absence of calcium can be obtained.

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