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The Brightest and Most Sensitive Red Fluorescent Ca2+ Indicator


Calcium acts as a universal second messenger in a variety of cells.
Since the 1920s, scientists have attempted to measure Ca2+, but few were successful due to limited availability of Ca2+ probes. The first reliable measurements of Ca2+ were performed by Ridgway and Ashley by injecting the photoprotein aequorin into the giant muscle fiber of the barnacle. Subsequently, in the 1980s, Tsien and colleagues produced a variety of fluorescent indicators. Among them the rhodamine-based Ca2+ reagents (such as Rhod-2) have been the most valuable longer wavelength dye for measuring Ca2+ with red fluorescence.

Although Rhod-2 has been the most popular red fluorescent Ca2+ indicator, its mitochondrial localization and high basal Ca2+ signal in cells have severely limited its cellular applications. In addition, the less optimal excitation of Rhod-2 at 488 nm makes it less robust to use with some instruments (such as FLIPR™) that have only 488 nm excitation light source. Quest Rhod-4™ serial calcium detection reagents have been developed to address these limitations of Rhod-2. The absorption and emission peaks of Quest Rhod-4™ reagents are 530 nm and 555 nm, respectively. Although Rhod-4 has maximum absorption at 530 nm, its absorption at 488 nm is quite strong. It is quite unique that Quest Rhod-4™ can be well excited with an argon ion laser at 488 nm besides the longer wavelength 514 nm, 532 nm and 546 nm excitations. Quest Rhod-4™ emits fluorescence (at wavelengths 555 nm) that increases with increasing Ca2+. Quest Rhod-4™ is determined to undergo a >200-fold increase in fluorescence upon binding Ca2+. Because the range of increase in Ca2+ in many cells after stimulation is generally 5- to10-fold, Quest Rhod-4™ is an excellent probe to use with high sensitivity in this region. The Kd of Quest Rhod-4™ is estimated to be 525 nM (22°C, pH 7.0 –7.5), but this value may be significantly influenced by pH, viscosity, and binding proteins in vivo conditions.
 Cat #  Product Name  Unit Size
ABD-21121  Quest Rhod-4™, AM  5x50 µg
ABD-21123  Quest Rhod-4™, AM  20x50 µg
ABD-21128  Quest Rhod-4™, sodium salt  5x50 µg
ABD-36331  Screen Quest™ Rhod-4 NW Calcium Assay Kit *Medium Removal*  10 Plates
ABD-36334  Screen Quest™ Rhod-4 NW Calcium Assay Kit *1% FBS Growth Medium *  10 Plates

Calcium Detection Reagents

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