Screen Quest™ Rhod-4 No Wash Calcium Assay Kit *Medium Removal*
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Catalog Number | |
Unit Size | |
Quantity |
Telephone | 1-800-990-8053 |
Fax | 1-800-609-2943 |
sales@aatbio.com | |
International | See distributors |
Bulk request | Inquire |
Custom size | Inquire |
Shipping | Standard overnight for United States, inquire for international |
Excitation (nm) | 523 |
Emission (nm) | 551 |
Quantum yield | 0.11 |
Certificate of Origin | Download PDF |
H-phrase | H303, H313, H333 |
Hazard symbol | XN |
Intended use | Research Use Only (RUO) |
R-phrase | R20, R21, R22 |
UNSPSC | 12352200 |
Screen Quest™ Rhod-4 No Wash Calcium Assay Kit |
Overview | ![]() ![]() |
Excitation (nm) 523 | Emission (nm) 551 | Quantum yield 0.11 |
Platform
Fluorescence microplate reader
Excitation | 540 nm |
Emission | 590 nm |
Cutoff | 570 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, ViewLuxComponents
Example protocol
AT A GLANCE
Protocol summary
- Prepare cells
- Remove the growth medium
- Add Rhod-4 NW dye-loading solution (100 µL/well for 96-well plate or 25 µL/well for 384-well plate)
- Incubate at room temperature for 1 hour
- Monitor fluorescence intensity at Ex/Em = 540/590 nm
Important notes
Do not add additional probenecid. Thaw 1 vial of Rhod-4 NW (Component A), 1 bottle of 10X Pluronic® F127 Plus (Component B), and 1 bottle of HHBS (Component C) at room temperature before use.
PREPARATION OF STOCK SOLUTION
1. Rhod-4 NW stock solution:
Add 100 µL of DMSO into the vial of Rhod-4 NW (Component A) and mix well. Protect from light. Note: 10 µL of Rhod-4 NW stock solution is enough for one plate.
2. Assay Buffer (1X):
a) For Cat. #36330 (1 plate kit) and # 36331 (10 plates kit) , make 1X Assay Buffer by adding 9 mL of HHBS (Component C) into 10X Pluronic® F127 Plus (1 mL, Component B), and mix them well.
b) For Cat. # 36332 (100 plates kit), make 1X Assay Buffer by adding 90 mL of HHBS (Not included) into 10X Pluronic® F127 Plus (10 mL, Component B), and mix them well. Note: 10 mL of 1X assay buffer is enough for one plate. Aliquot and store un-used 1X assay buffer at < -20 oC. Protect from light and avoid repeated freeze-thaw cycles.
PREPARATION OF WORKING SOLUTION
Rhod-4 NW dye-loading solution:
Add 10 µL of Rhod-4 NW stock solution into 10 mL of 1X assay buffer, and mix them well. Note: This working solution is stable for at least 2 hours at room temperature.
For guidelines on cell sample preparation, please visit
https://www.aatbio.com/resources/guides/cell-sample-preparation.html
SAMPLE EXPERIMENTAL PROTOCOL
- Remove the growth medium from the cell plate. Note: It is important to remove the growth medium in order to minimize background fluorescence, and compound interference with serum or culture media. Alternatively, grow the cells in growth medium with 1 - 5% FBS to avoid medium removal step. In this case, 2X dye loading solution in HHBS buffer is needed. [We offer 2 separate no wash calcium assay kits (Cat. #36334 and Cat. #36335) for those who use 0.5 - 1% FBS in growth medium to avoid the medium removal step].
- Add 100 µL/well (96-well plate) or 25 µL/well (384-well plate) of Rhod-4 NW dye-loading solution into the cell plates.
- Incubate the dye-loading plate in a cell incubator for 30 minutes, then incubate the plate at room temperature for another 30 minutes. Note: If the assay requires 37°C, perform the experiment immediately without further room temperature incubation. If the cells can function well at room temperature for longer time, incubate the cell plate at room temperature for 1 - 2 hours.
- Prepare the compound plate with HHBS or the desired buffer.
- Run the calcium flux assay by monitoring the fluorescence intensity at Ex/Em = 540/590 nm.
Spectrum

Spectral properties
Excitation (nm) | 523 |
Emission (nm) | 551 |
Quantum yield | 0.11 |
Product Family
Name | Excitation (nm) | Emission (nm) | Extinction coefficient (cm -1 M -1) | Quantum yield |
Screen Quest™ Fluo-4 No Wash Calcium Assay Kit | 495 | 528 | 82000 | 0.161 |
Images

References
Authors: Territo PR, Heil J, Bose S, Evans FJ, Balaban RS.
Journal: Appl Spectrosc (2007): 138
Authors: Stamm C, del Nido PJ.
Journal: Thorac Cardiovasc Surg (2004): 127
Authors: Martin VV, Beierlein M, Morgan JL, Rothe A, Gee KR.
Journal: Cell Calcium (2004): 509
Authors: Bednar B, Cunningham ME, Kiss L, Cheng G, McCauley JA, Liverton NJ, Koblan KS.
Journal: J Neurosci Methods (2004): 247
Authors: Stamm C, Friehs I, Choi YH, Zurakowski D, McGowan FX, del Nido PJ.
Journal: Cardiovasc Res (2003): 695
Authors: Du C, MacGowan GA, Farkas DL, Koretsky AP.
Journal: Biophys J (2001): 549
Authors: Du C, MacGowan GA, Farkas DL, Koretsky AP.
Journal: Cell Calcium (2001): 217
Authors: Lannergren J, Westerblad H, Bruton JD.
Journal: J Muscle Res Cell Motil (2001): 265
Authors: MacGowan GA, Du C, Glonty V, Suhan JP, Koretsky AP, Farkas DL.
Journal: J Biomed Opt (2001): 23
Authors: Muriel MP, Lambeng N, Darios F, Michel PP, Hirsch EC, Agid Y, Ruberg M.
Journal: J Comp Neurol (2000): 297
Application notes
A Meta-Analysis of Common Calcium Indicators
A New Red Fluorescent & Robust Screen Quest™ Rhod-4™ Ca2+Indicator for Screening GPCR & Ca2+ Channel Targets
A New Robust No-Wash FLIPR Calcium Assay Kit for Screening GPCR and Calcium Channel Targets
A Novel NO Wash Probeniceid-Free Calcium Assay for Functional Analysis of GPCR and Calcium Channel Targets
FAQ
Are there upgraded trypan blue derivatives for cell viability testing?
Can I intracellularly measure mitochondria calcium flux and changes in mitochondria membrane potential at the same time?
Do you offer any products for measuring intracellular calcium concentration or movement by flow cytometry?
Does EDTA inactivate proteinase K?