Description
Key facts
Detection method:Fluorescent,
Sample types:Suspension cells, Adherent cells,
Assay type:Cell-based,
Assay time:1h 20m,
Assay Platform:Microplate reader
Product details:
Calcium Flux Assay Kit (Fura-2, No Wash, Ratiometric) ab176766 is a no-wash, ratiometric calcium assay that allows homogeneous measurement of intracellular calcium mobilization caused by activation of G-protein-coupled receptors (GPCR) or calcium channels. The assay is suitable for use with 96-well and 384-well microplates.
In the Fura-2 calcium flux assay protocol, cells are pre-loaded with Fura-2 AM (which can cross the cell membrane). Once inside the cell, the lipophilic blocking groups of Fura-2 AM are cleaved by esterases, resulting in a negatively charged fluorescent dye that stays inside the cell.
The excitation maxima of the dye shifts from 363 nm to 335 nm upon binding of the dye to calcium. The emission spectra is relatively unaffected. The change is most easily measured by comparing the emission at 510 nm on excitation separately at 380 nm and 340 nm.
Measuring the ratio reduces the effects of uneven dye loading and cell numbers, dye leakage and photo bleaching, compared to measuring cellular calcium concentrations using a single wavelength dye like Fluo-8.
Fura-2 calcium flux assay protocol summary:
- add Fura-2 solution to cells
- incubate for 1 hr at 37°C, then for 20 min at room temp
- analyze with a microplate reader at both Ex/Em 340/510 nm and Ex/Em 380/510nm
This product is intended to be used for monitoring calcium fluctuations in vivo in live cells using the following HTS imaging plate readers: FLIPR™, FDSS, BMG NOVOstar™, FLexStation, ViewLux, IN Cell Analyzer or Arrayscan.
Properties and Storage Information:
Shipped at conditions-Blue Ice, Appropriate short-term storage conditions--20°C, Appropriate long-term storage conditions--20°C, Storage information--20°C
Supplementary Information:
This supplementary information is collated from multiple sources and compiled automatically.
Calcium ions (Ca²⁺) serve as essential signaling molecules in various cellular processes. These ions have an atomic mass number of 40.08. Calcium ions are highly concentrated in bone tissues teeth and within cells mostly stored in the endoplasmic reticulum and mitochondria. Common indicators of calcium concentration in research include the use of dyes such as Fura-2 and Calbryte 520 AM. These indicators help in visualizing the transient increases in intracellular calcium levels which are important in cellular signaling.
Biological function summary
Calcium ions mediate various physiological functions including muscle contraction neurotransmitter release and hormone secretion. They typically do not act as lone agents; instead they often function as part of protein complexes particularly with calcium-binding proteins like calmodulin and troponin C. Calcium ions also interact with enzymes such as protein kinases and phosphatases regulating metabolic pathways and genetic expression.
Pathways
Calcium ions play pivotal roles in the calcium signaling pathway and the cardiac excitation-contraction coupling pathway. In the calcium signaling pathway calcium ions once released from stores like the endoplasmic reticulum activate proteins such as calmodulin which then modulates various cellular activities. In the cardiac excitation-contraction coupling pathway calcium bows its link to proteins like sarcoplasmic reticulum calcium ATPase essential for heart muscle function.
Calcium ion dysregulation can lead to conditions such as osteoporosis and cardiac arrhythmias. In osteoporosis low calcium levels contribute to decreased bone density affecting bone health and making them prone to fractures. In cardiac arrhythmias abnormal calcium handling can result in irregular heartbeats. Proteins such as calbindin-D28k are closely connected to calcium metabolism disorders highlighting the significance of calcium in maintaining physiological balance.
