Screen Quest™ Calbryte-520 Probenecid-Free and Wash-Free Calcium Assay Kit

Calcium flux assays are the preferred methods in drug discovery for screening G protein coupled receptors (GPCR). Screen Quest™ Calbryte-520 Probenecid-Free and Wash-Free Calcium Assay Kit provides the most robust homogeneous fluorescence-based assay for detecting the intracellular calcium mobilization. Cells expressing a GPCR of interest that signals through calcium are pre-loaded with our proprietary Calbryte™-520 AM which can cross cell membrane. Calbryte™-520 AM is the brightest calcium indicator available for HTS screening. Once inside the cell, the lipophilic blocking groups of Calbryte™-520 AM are cleaved by non-specific cell esterase, resulting in a negatively charged fluorescent dye that stays inside cells, and its fluorescence is greatly enhanced upon binding to calcium. When cells stimulated with screening compounds, the receptor signals release of intracellular calcium, which greatly increase the fluorescence of Calbryte™-520 AM. The characteristics of its excellent cell retention, high sensitivity, and 100-250 times fluorescence increases (when it forms complexes with calcium) make Calbryte™-520 AM an ideal indicator for measurement of cellular calcium. Calbryte™-520 AM is the only calcium dye that does not require probenecid for better cellular retention. This Screen Quest™ Calbryte-520 Probenecid-Free and Wash-Free Calcium Assay Kit provides the most optimized assay method for monitoring G-protein-coupled receptors (GPCRs) and calcium channels with fragile or difficult cell lines. The assay can be performed in a convenient 96-well or 384-well microtiter-plate format and easily adapted to automation.

Comparison of fluorescent signal response of endogenous P2Y receptor to ATP in CHO-K1 cells. CHO-K1 cells were seeded overnight at 50,000 cells/100 µL/well in a 96-well black wall/clear bottom costar plate. Calcium flux response was measured with Screen Quest™ Calbryte™ 520 Probenecid-Free and Wash-Free Calcium Assay Kit, FLIPR Calcium 4 Assay Kit , and Fluo-4 Direct Calcium Assay kit. ATP (50 µL/well) was added by FlexStation 3 to achieve the final indicated concentrations.

Protocol

COA

Catalog	Size	Price
36317	1 Plate	Price
36318	10 Plates	Price
36319	100 Plates	Price

Components

Catalog 36317 (1 Plate)

Component A: Calbryte™ 520 AM	1 vial
Component B: 10X Pluronic® F127 Plus	1 vial (1 mL)
Component C: HHBS (Hanks’ buffer with 20 mM Hepes)	1 vial (9 mL)

Catalog 36318 (10 Plates)

Component A: Calbryte™ 520 AM	1 vial
Component B: 10X Pluronic® F127 Plus	1 bottle (10 mL)
Component C: HHBS (Hanks’ buffer with 20 mM Hepes)	1 bottle (100 mL)

Catalog 36319 (100 Plates)

Component A: Calbryte™ 520 AM	10 vials
Component B: 10X Pluronic® F127 Plus	10 bottles (10 mL/bottle)
Component C: HHBS (Hanks’ buffer with 20 mM Hepes)	Not provided

Citations

View all 16 citations: Citation Explorer

OPEN ACCESS EDITED BY

Authors:

Taggart, Clifford and Kesturu, Girish S and Brown, Ryan and Wu, Xiaojun and Wang, Zhengtao and Chen, Z and Wang, G and Xie, X and Liu, H and Liao, J and others,

Journal:

Oral neutrophils-the good, the bad, and the ugly (2023): 41

Preferential Gs protein coupling of the galanin Gal1 receptor in the $\mu$-opioid-Gal1 receptor heterotetramer

Authors:

De Oliveira, Paulo A and Moreno, Estefan{\'\i}a and Casajuana-Martin, Nil and Casad{\'o}-Anguera, Ver{\`o}nica and Cai, Ning-Sheng and Camacho-Hernandez, Gisela Andrea and Zhu, Hu and Bonifazi, Alessandro and Hall, Matthew D and Weinshenker, David and others,

Journal:

Pharmacological Research (2022): 106322

Selective activation of TRPA1 ion channels by nitrobenzene skin sensitizers DNFB and DNCB

Authors:

Wu, Han and Niu, Canyang and Qu, Yaxuan and Sun, Xiaoying and Wang, KeWei

Journal:

Journal of Biological Chemistry (2022)

Selective activation of TRPA1 ion channel by skin sensitizer nitrobenzene DNFB

Authors:

Wu, Han and Qu, Yaxuan and Sun, Xiaoying and Wang, Kewei

Journal:

Authorea Preprints (2021)

Diquafosol tetrasodium elicits total cholesterol release from rabbit meibomian gland cells via P2Y 2 purinergic receptor signalling

Authors:

Endo, Ken-ichi and Sakamoto, Asuka and Fujisawa, Koushi

Journal:

Scientific reports (2021): 1--8

References

View all 172 references: Citation Explorer

Calcium imaging of cortical neurons using Fura-2 AM

Authors:

Barreto-Chang OL, Dolmetsch RE.

Journal:

J Vis Exp. (2009)

A flow cytometric comparison of Indo-1 to fluo-3 and Fura Red excited with low power lasers for detecting Ca(2+) flux

Authors:

Bailey S, Macardle PJ.

Journal:

J Immunol Methods (2006): 220

Measurement of [Ca2+]i in whole cell suspensions using fura-2

Authors:

Hirst RA, Harrison C, Hirota K, Lambert DG.

Journal:

Methods Mol Biol (2006): 37

Load of calcium probe Fura -2/AM in Escherichia coli cells

Authors:

Shao M, Wang HM, Liu ZH, Shen P, Cai RX.

Journal:

Wei Sheng Wu Xue Bao (2005): 805

Fluorescence measurements of free [Mg2+] by use of mag-fura 2 in Salmonella enterica

Authors:

Froschauer EM, Kolisek M, Dieterich F, Schweigel M, Schweyen RJ.

Journal:

FEMS Microbiol Lett (2004): 49

Spectral properties

Excitation (nm)	493
Emission (nm)	515
Quantum yield	0.75 ¹

Storage, safety and handling

H-phrase	H303, H313, H333
Hazard symbol	XN
Intended use	Research Use Only (RUO)
R-phrase	R20, R21, R22
UNSPSC	12352200

Instrument settings

Fluorescence microplate reader
Excitation	490 nm
Emission	525 nm
Cutoff	515 nm
Recommended plate	Black wall/clear bottom
Instrument specification(s)	Bottom read mode/Programmable liquid handling

Other instruments	ArrayScan, FDSS, FLIPR, FlexStation, IN Cell Analyzer, NOVOStar, ViewLux

Documents

Protocol
Safety Data Sheet (Catalog 36317)
Safety Data Sheet (Catalog 36318)
Safety Data Sheet (Catalog 36319)
Certificate of Analysis

Telephone
Fax
Email	sales@aatbio.com
International	See distributors
Bulk request	Inquire
Custom size	Inquire
Technical Support	Contact us
Request quotation	Request
Purchase order	Send to sales@aatbio.com
Shipping	Standard overnight for United States, inquire for international

Page updated on October 18, 2025