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AAT Bioquest

Which calcium dyes work with flow cytometry lasers?

Posted February 2, 2024


Answer

There are various types of calcium dyes which work with flow cytometry lasers. 

  • Fura-2 dye is a UV light-excitable calcium indicator commonly used for ratio-imaging microscopy. It exhibits a shift in absorption upon binding calcium, allowing observation through scanning the excitation spectrum between 300 and 400 nm with an emission wavelength of approximately 510 nm.
  • Fluo-3 is a fluorescence indicator for intracellular calcium commonly used in flow cytometry and confocal laser scanning microscopy. Its fluorescence sharply increases upon binding calcium, with an emission maximum at 525 nm, suitable for detection using a fluorescein isothiocyanate (FITC) channel.
  • Indo-1, another UV light-excitable calcium indicator, is preferred for flow cytometry using a single laser for excitation. Its emission maximum shifts from 475 nm in calcium-free conditions to 400 nm when saturated with calcium. 
  • Quin-2 is a calcium indicator known for its high selectivity, being unaffected by sodium gradients, intracellular pH or membrane potential. It displays high affinity for calcium ions and is sensitive enough to monitor low calcium levels, with excitation and emission maxima at 339/492 nm.
  • BioTracker NIR Ca2+ dyes (SCT021, SCT022, SCT023), which are far-red fluorescent calcium indicators. These dyes exhibit a substantial change in fluorescence intensity (up to 1000 fold) upon binding to calcium in viable cells. 
  • Cal-520 is a novel calcium indicator that offers several improvements and advantages over older generations of calcium indicators like Fluo-3 and Fluo-4. One key improvement in Cal-520 is an alteration in its ideal absorption and emission wavelengths, approximately 12 nm shorter than those of Fluo-3. This shift results in a maximum absorption wavelength of 494 nm. 
Additional resources

Ratiometric Analysis of Fura Red by Flow Cytometry: A Technique for Monitoring Intracellular Calcium Flux in Primary Cell Subsets

Calcium Indicators

Cal-520®, AM