The effects of pH and temperature on fluorescent calcium indicators as determined with Chelex-100 and EDTA buffer systems.
A novel method of determining the apparent dissociation constants of fluorescent calcium indicators is described which utilizes Chelex-100 ion exchange resin and 45Ca. The affinity for calcium of indicators fluo-3, fura-2 and indo-1 measured at either 22 degrees or 37 degrees C decreases as pH is decreased from 7.4 to 5.5. These measurements agree with determinations made using EDTA-calcium buffers. The 1:1 calcium:indicator complex is maintained under all conditions. The necessity to correct dissociation constants during intracellular acidification to properly interpret fluorescence measurements is illustrated by indo-1 measurements in the ischemic rat heart.
Lattanzio
F A
FA
Department of Pharmacology, Eastern Virginia Medical School, Norfolk 23501.
D000814:Aniline Compounds; D001572:Benzofurans; D002118:Calcium / Q000032:analysis*; D002614:Chelating Agents; D004492:Edetic Acid; D016257:Fura-2; D006863:Hydrogen-Ion Concentration; D007211:Indoles; D012117:Resins, Synthetic; D013696:Temperature; D014966:Xanthenes
D000814:Aniline Compounds; D001572:Benzofurans; D002614:Chelating Agents; D007211:Indoles; D012117:Resins, Synthetic; D014966:Xanthenes; C024997:Chelex 100; C059715:Fluo-3; D004492:Edetic Acid; C048960:indo-1; D002118:Calcium; D016257:Fura-2