Calcium Flux Assay Kit (Fluo-8, No Wash)

Product details

Fluo-8 Calcium Flux Assay Kit ab112129 is a no-wash, fluorescence-based assay for detecting intracellular calcium mobilization in cells.

This product is optimized 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.

Readers with an in built pipetter are essential for measuring calcium response accurately due to transient property of the calcium signal. Depending on dose of the agonist, the peak response is few seconds to 20 seconds, so one will need to add the compound by instrument and read the fluorescent signal simultaneously every second for ~1-2 min.

Calcium flux assay principle
In the Fluo-8 Calcium Flux Assay Kit protocol, cells are pre-loaded with Fluo-8 which can cross the cell membrane. Once inside the cell, the lipophilic blocking groups of Fluo-8 are cleaved by an esterase, resulting in a negatively charged fluorescent dye that stays inside the cell. Its fluorescence is greatly enhanced upon binding to calcium.If intracellular calcium levels increase, the fluorescence of Fluo-8 is significantly increased.

The characteristics of its long wavelength, high sensitivity, and >100 times fluorescence enhancement make Fluo-8 the brightest green calcium indicator available, and it is an ideal tool for the measurement of cellular calcium through HTS screening.

Fluo-8 Calcium Flux assay protocol summary
- Prepare cells in growth medium with 1-5% FBS
- Add Fluo-8 NW dye-loading solution
- Incubate at room temperature for 1 hour
- Monitor fluorescence intensity at Ex/Em = 490/525 nm

Related Fluo-8 assay kits
Fluo-8 Calcium Flux Assay Kits are available in No Wash and Medium Removal variants:
- No wash kit Fluo-8 Calcium Flux Assay Kit ab112129 is generally preferred as the workflow is simpler. It is also recommended for cell lines that are only weakly adherent and risk losing adherence when washing.
- Medium removal kit Fluo-8 Calcium Flux Assay Kit ab112128 is recommended for the testing of compounds, receptors and calcium channels that are sensitive to the presence of cell culture media or serum. Alternatively, for cells that can tolerate it, consider using a low serum % in culture media and use No Wash Fluo-8 kit ab112129.

Fluo-8 is also available as a standalone biochemical ab142773.

If you would like to quantify calcium concentration in vitro using cell extracts, we recommend using Calcium Detection Kit (Colorimetric) (ab102505) or Calcium Quantification Assay Kit (ab112115).

How other researchers are using ab112129
ab112129 has been used in a variety of sample types including:
- Human OA chondrocytes and C28I2 cells 1
-Neuronal cells 2
- Human U2OS, HeLa, BJ, BJ-5ta (BJ-T), T98G, U87 MG and MRC5 cell lines and mouse embryonic fibroblasts (MEF) 3
References: 1- Fu W et al. 2024; 2- Lui L et al. 2023; 3- Munk S et al. 2023