Monitor calcium mobilization in GFP-expressing cell lines
The brightest and most sensitive red-shifted fluorescent Ca2+ indicator
Provides comparable EC50 and IC50 values as obtained using Rhod-4 AM and Fluo-4 AM
Larger assay window and higher Z' factor values, allowing measurements of challenging cell lines and receptors
Enzo Life Sciences’ GFP-CERTIFIED® FLUOFORTE® Calcium Assay Kit (high-throughput) provides a homogeneous fluorescence-based assay for detecting intracellular calcium mobilization across a broad spectrum of biological targets. Relative to other commercially available dyes, the GFP-CERTIFIED® FLUOFORTE® dye is the brightest and most sensitive fluorescent calcium indicator. The kit provides a homogeneous mix-and-read, no-wash calcium mobilization assay. The homogenous cell-based assay for calcium offers fewer steps, lower variability and an easier protocol for adherent and non-adherent cell lines. In addition, it requires neither a washing step, nor exogenous addition of a quencher dye, which could adversely effect receptor-ligand interaction kinetics.
Histamine H1 receptors: HeLa cells were seeded overnight at 40,000 cells per 100 µL per well in a 96-well black wall/clear bottom microplate. The cells were incubated with 100 µL of GFP-CERTIFIED® FLUOFORTE® reagent or Rhod-2 AM dye for 1 hour at 37°C. Histamine (20µL/well) was added using a BioTek two-syringe pump dispenser to achieve the final indicated concentrations. No significant difference in EC50 of Histamine for GFP-CERTIFIED® FLUOFORTE® and Rhod2-AM dye was observed. GFP-CERTIFIED® FLUOFORTE® reagent generated higher intensity signal and a larger assay window.
U-2 OS cells were incubated with (A) GFP-CERTIFIED® FLUOFORTE® AM dye or (B) Rhod-2 AM dye. Cells were washed and treated with a final concentration of 100nM ATP and imaged immediately. GFP-CERTIFIED® FLUOFORTE® AM dye exhibits brighter fluorescence intensity than Rhod-2 AM.
The GFP-CERTIFIED® FLUOFORTE® Calcium Assay kit provides a homogeneous fluorescence-based assay for detecting intracellular calcium mobilization across a broad spectrum of biological targets.
Quality Control:
% purity of Reagent A (GFP-CERTIFIED® FLUOFORTE® dye) by HPLC is ≥95%.
Quantity:
ENZ-51020-KP010: for 10 x 96-well plates ENZ-51020-KP100: for 100 x 96-well plates
Handling:
Protect from light. Avoid freeze/thaw cycles.
Shipping:
Dry Ice
Short Term Storage:
-20°C
Long Term Storage:
-80°C
Contents:
Starter pack (Prod. No. ENZ-51020-KP010): Reagent A (lyophilized GFP-CERTIFIED® FLUOFORTE® dye), 1 vial Reagent B (dye efflux inhibitor), 10ml Reagent C (Hanks’ buffer with 20mM HEPES, 100ml)
Sufficient for 10 x 96-well plates
High-throughput (Prod. No. ENZ-51020-KP100): Reagent A (lyophilized GFP-CERTIFIED® FLUOFORTE® dye), 10 vials Reagent B (dye efflux inhibitor), 10 x 10ml
Sufficient for 100 x 96-well plates
Technical Info/Product Notes:
The GFP-CERTIFIED® FLUOFORTE® Calcium assay kit is a member of the CELLESTIAL® product line, reagents and assay kits comprising fluorescent molecular probes that have been extensively benchmarked for live cell analysis applications. CELLESTIAL® reagents and kits are optimal for use in demanding cell analysis applications involving confocal microscopy, flow cytometry, microplate readers and HCS/HTS, where consistency and reproducibility are required.
Regulatory Status:
RUO - Research Use Only
Product Literature References
Perfluorooctane sulfonate and bisphenol A induce a similar level of mast cell activation via a common signaling pathway, Fyn-Lyn-Syk activation: S.J. Park, et al.; Food Chem. Toxicol. 156, 112478 (2021), Abstract;
Phorbol myristate acetate induces differentiation of THP-1 cells in a nitric oxide-dependent manner: Y.Y. Chang, et al.; Nitric Oxide 109, 33 (2021), Abstract;
Absolute proteomic quantification reveals design principles of sperm flagellar chemosensation: C. Trötschel, et al.; EMBO J. 39, e102723 (2020), Abstract; Full Text
Nanobody-directed targeting of optogenetic tools to study signaling in the primary cilium: J.N. Hansen, et al.; Elife 9, e57907 (2020), Abstract; Full Text
Interplay between sub-cellular alterations of calcium release and T-tubular defects in cardiac diseases: M. Scardigli, et al.; Front. Physiol. 9, 1474 (2018), Abstract; Full Text
Characterization of cholesterol homeostasis in sphingosine-1-phosphate lyase-deficient fibroblasts reveals a Niemann-Pick disease type C-like phenotype: H. Vienken, et al.; Sci. Rep. 7, 43575 (2017), Abstract; Full Text
Novel insights on the relationship between T-tubular defects and contractile dysfunction in a mouse model of hypertrophic cardiomyopathy: C. Crocini, et al.; J. Mol. Cell. Cardiol. 91, 42 (2016), Abstract; Full Text
Sulfur dioxide protects against collagen accumulation in pulmonary artery in association with downregulation of the transforming growth factor β1/smad pathway in pulmonary hypertensive rats: W. Yu, et al.; J. Am. Heart Assoc. 5, e003910 (2016), Abstract; Full Text
T-tubular electrical defects contribute to blunted β-adrenergic response in heart failure: C. Crocini, et al.; Int. J. Mol. Sci. 17, 1471 (2016), Abstract; Full Text
Defects in T-tubular electrical activity underlie local alterations of calcium release in heart failure: C. Crocini, et al.; PNAS 111, 15196 (2014), Abstract; Full Text
Gα13/PDZ-RhoGEF/RhoA signaling is essential for gastrin-releasing peptide receptor-mediated colon cancer cell migration: M. Patel, et al.; Mol. Pharmacol. 86, 252 (2014), Application(s): Microplate reader using Caco-2 cells, Abstract; Full Text
Paxillin is an intrinsic negative regulator of platelet activation in mice: A. Sakuta, et al.; Thromb. J. 12, 1 (2014), Abstract; Full Text
GABAergic signaling in the pulmonary neuroepithelial body microenvironment: functional imaging in GAD67-GFP mice: K. Schnorbusch, et al.; Histochem. Cell Biol. 140, 549 (2013), Abstract;
The N-terminally truncated µ3 and µ3-like opioid receptors are transcribed from a novel promoter upstream of exon 2 in the human OPRM1 gene: S. Andersen, et al.; PLoS One. 8, e71024 (2013), Application(s): Microplate reader using HEK293 cells, Abstract; Full Text
Leukotriene C4 induces migration of human monocyte-derived dendritic cells without loss of immunostimulatory function: J. Dannull, et al.; Blood 119, 3113 (2012), Application(s): Intracellular calcium release in human monocyte derived dendritic cells with Tecan Infinite plate reader, Abstract; Full Text
FLIM FRET technology for drug discovery: Automated multiwell‐plate high‐content analysis, multiplexed readouts and application in situ: S. Kumar, et al.; Chemphyschem. 12, 609 (2011), Abstract; Full Text
Fluorescence imaging of Influenza virus H1N1 mRNA in living infected cells using single chromophore FIT-PNA: S. Kummer, et al.; Angew. Chem. Int. Ed. Engl. 50, 1931 (2011), Abstract;
N-way FRET microscopy of multiple protein-protein interactions in live cells: A.D. Hoppe, et al.; PLoS One 8, e64760 (2011), Abstract; Full Text
General Literature References
Novel Fluo-4 analogs for fluorescent calcium measurements: V.V. Martin, et al. ; Cell Calcium 36, 509 (2004), Abstract;
Calcium measurements in perfused mouse heart: quantitating fluorescence and absorbance of Rhod-2 by application of photon migration theory: C. Du, et al.; Biophys. J. 80, 549 (2001), Full Text
Rhod-2 based measurements of intracellular calcium in the perfused mouse heart: cellular andsubcellular localization and response to positive inotropy: G.A. MacGowan, et al.; J. Biomed. Opt. 6, 23 (2001), Abstract;
Flow cytometric kinetic assay of calcium mobilization in whole blood platelets using Fluo-3 and CD41: M. do Céu Monteiro, et al.; Cytometry 35, 302 (1999), Abstract;
Nucleoplasmic and cytoplasmic differences in the fluorescence properties of the calcium indicator Fluo-3: C. Perez-Terzic, et al.; Cell Calcium 21, 275 (1997), Abstract;
Improved four-color flow cytometry method using fluo-3 and triple immunofluorescence for analysis of intracellular calcium ion ([Ca2+]i) fluxes among mouse lymph node B- and T-lymphocyte subsets: R. Greimers, et al.; Cytometry 23, 205 (1996), Abstract;
