Fura-2 Calcium Flux Assay Kit
The Fura-2 calcium kit is a simple mixed reading format that uses the industry standard Fura-2 ratio calcium indicator to accurately measure calcium mobilization.
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Description
View the largest Fura-2 signal window
Eliminate washing artifacts and increase throughput through homogeneous measurement
Minimize the impact of uneven dye loading and leakage on the results
Use a no-wash protocol to interrogate low-density, weak or non-adherent cells
Analysis on FlexStation®3 and FLIPR®Tetra systems
Cite thousands of citation forms, among which Fura-2 has been used in cell lines and targets
The Fura-2 Calcium Flux Assay kit uses Fura-2, AM, a calcium-sensitive dye, which is absorbed into the cytoplasm of the cell during the incubation process, and the lipophilic blocking group is cleaved, causing the negatively charged fluorescent dye to remain in the cell Inside. After the target is activated, it releases intracellular calcium and binds to the dye, thereby increasing the fluorescence intensity. The kit uses extracellular masking technology to block background fluorescence and increase the signal window of the measurement, and reduce the measurement steps.
Fig.2 Calcium flux signal changes
The calcium-bound Fura-2 is excited at 340/510 nm, and the unbound form is excited at 380/510 nm. After calcium has flowed through, the cytoplasmic calcium concentration increases, resulting in a proportional increase in the fluorescence intensity at 340/510 nm. On the contrary, the intensity of 380/510 nm decreases in tandem with the decrease in the concentration of unbound dye. By matching the fluorescence intensity generated by excitation at two wavelengths, artifacts related to uneven dye distribution and cell number changes will be minimized because they will affect both measurements.
Fig.3 Fura-2 Calcium Flux Assay dye when bound to Ca2+
The Fura-2 calcium dye is excited at 340nm and 380nm, and emits at 510nm. Through the calcium flux, the changes in the intracellular calcium concentration can be measured.
On the FlexStation®3 multi-mode microplate reader, carbachol was used to stimulate the muscarinic M1 receptor on CHO M1 cells to generate a concentration response curve (CRC). The Fura-2 dye was combined with BD Kit and traditional Fura-2 washing solution was compared with protocol. The EC50 values of all methods are similar and within the range of published values.
Fig.2 Antagonistic effect of muscarinic M1 receptor on CHO M1 cells
On the FlexStation 3 multi-mode microplate reader, 50 nM carbachol was used as an agonist to attack compared with the atropine antagonist CRC. The experiment Fura-2 Calcium Flux Assay kit provides the largest signal window and the most stable Z factor at EC80.
Fig.3 Agonistic effects of endogenous H1 receptors on HeLa cells
Histamine is used to stimulate endogenous histamine receptors on HeLa cells. The FLIPRTetra® high-throughput cell screening system with UV LED was used for the assay. The generated concentration response curve (CRC) compares the Fura-2 dye with the BD Ratiometric calcium kit and the traditional Fura-2 washing protocol. Using the Fura-2 Calcium Flux Assay kit, the EC50 value is in the semi-log range, and the Z factor at EC80 is the largest.
Fig.4 The antagonistic effect of endogenous H1 receptor on HeLa cells
The FLIPR Tetra high-throughput cell screening system with UV LED was used for the assay, and similar experimental settings and system settings were used in Figure 3. Compared to pyrrolamide's antagonist CRC, 40 nM histamine was used as an agonist attack.
Fig.1 Antagonistic effect of muscarinic M1 receptor on CHO M1 cells
On the FlexStation®3 multi-mode microplate reader, carbachol was used to stimulate the muscarinic M1 receptor on CHO M1 cells to generate a concentration response curve (CRC). The Fura-2 dye was combined with BD Kit and traditional Fura-2 washing solution was compared with protocol. The EC50 values of all methods are similar and within the range of published values.
Fig.2 Antagonistic effect of muscarinic M1 receptor on CHO M1 cells
On the FlexStation 3 multi-mode microplate reader, 50 nM carbachol was used as an agonist to attack compared with the atropine antagonist CRC. The experiment Fura-2 Calcium Flux Assay kit provides the largest signal window and the most stable Z factor at EC80.
Fig.3 Agonistic effects of endogenous H1 receptors on HeLa cells
Histamine is used to stimulate endogenous histamine receptors on HeLa cells. The FLIPRTetra® high-throughput cell screening system with UV LED was used for the assay. The generated concentration response curve (CRC) compares the Fura-2 dye with the BD Ratiometric calcium kit and the traditional Fura-2 washing protocol. Using the Fura-2 Calcium Flux Assay kit, the EC50 value is in the semi-log range, and the Z factor at EC80 is the largest.
Fig.4 The antagonistic effect of endogenous H1 receptor on HeLa cells
The FLIPR Tetra high-throughput cell screening system with UV LED was used for the assay, and similar experimental settings and system settings were used in Figure 3. Compared to pyrrolamide's antagonist CRC, 40 nM histamine was used as an agonist attack.
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