Actively helping customers, employees and the global community during the coronavirus SARS-CoV-2 outbreak.  Learn more >>

Cal-520™ Calcium Indicators

Cal-520™ provides the most robust homogeneous fluorescence-based assay tool for detecting intracellular calcium mobilization. Cal-520™ AM is a new fluorogenic calcium-sensitive dye with a significantly improved signal to background ratio and intracellular retention compared to the existing green calcium indicators (such as Fluo-3 AM and Fluo-4 AM). The higher signal/background ratio and better intracellular retention make the Cal-520™ calcium assay a robust tool for evaluating GPCR and calcium channel targets as well as for screening their agonists and antagonists. Our preliminary in-house research indicated that Cal-520™ AM can be readily loaded to a guinea pig heart and stays there for a few hours in the absence of probenecid. The calcium signal can be readily monitored with Cal-520™ AM while it is difficult to observe the calcium signal under the same conditions with other green calcium dyes, such as Fluo-3 AM and Fluo-4 AM.

Key Features of Cal-520™ AM:

  • Better Intracellular Retention, Cal-520™ AM is better retained in live cells than Fluo-3 AM and Fluo-4 AM.
  • Higher Sensitivity, Cal-520™ AM has much higher signal/background ratio than Fluo-3 AM and Fluo-4 AM in cells.
  • Convenient, Cal-520™ AM has almost identical spectra to those of Fluo-4 AM.


Left: ATP-stimulated calcium responses of endogenous P2Y receptor in CHO-K1 cells incubated with Cal-520™ AM (red curve, Cat# 21131), or Fluo-4 AM (blue curve) respectively with probenecid under the same conditions. CHO-K1 cells were seeded overnight at 50,000 cells/100 µL/well in a Costar 96-well black wall/clear bottom plate. 100 µL of 5 µM Fluo-4 AM or Cal-520™ AM in HHBS with 2.5 mm probenecid was added into the cells, and the cells were incubated at 37 °C for 2 hours. ATP (50 µL/well) was added using FlexSation® to achieve the final indicated concentrations. Right: ATP-stimulated calcium responses of endogenous P2Y receptors in CHO-K1 cells incubated with Cal-520™ AM (red curve, Cat# 21131), or Fluo-4 AM (blue curve) respectively, without probenecid under the same conditions. CHO-K1 cells were seeded overnight at 50,000 cells/100 µL/well in a Costar 96-well black wall/clear bottom plate. 100 µL of 5 µM Fluo-4 AM or Cal-520™ AM in HHBS was added into the cells, and the cells were incubated at 37 °C for 2 hours. ATP (50 µL/well) was added using FlexSation® to achieve the final indicated concentrations.

 

Table 1. Spectral Comparison of Fluo-3, Fluo-4, Fluo-8® and Cal-520

Dye
Ex (nm)
Em (nm)
QY*
Kd (nM)
Cal-520™ 4925140.75320
Fluo-3 5065250.15390
Fluo-4 4935150.16345
Fluo-8®4905140.16389
  1. *QY = Fluorescence Quantum Yield in the presence of 5 mM calcium citrate.

Table 2. Fluorescent Cal Calcium Indicators

Cat No.
Product Name
Ex (nm)
Em (nm)
Kd
Unit Size
21131Cal-520®, AM492514320 nM1 mg
21141Cal-520®, potassium salt492514320 nM1 mg
21136Cal-520®, sodium salt492514320 nM1 mg
21142Cal-520FF™, AM4925149.8 µM 1 mg
21144Cal-520FF™, potassium salt49251410x50 ug 9.8 µM
20501Cal Green™ 1, AM [Equivalent to Calcium Green-1, AM]50653110x50 ug 190 nM
20500Cal Green™ 1, hexapotassium salt50653110x50 ug 190 nM