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Rhod-2 AM

Cat. No. M10357

All AbMole products are for research use only, cannot be used for human consumption.

Rhod-2 AM Structure
Size Price Availability Quantity
50ug USD 60  USD60 In stock
1mg USD 560  USD560 In stock
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Quality Control & Documentation
  • Purity: >98%, Biological Stain
  • COA
  • MSDS
Biological Activity

Rhod-2 AM is a fluorescent, mitochondrial probe (λex=552 nm, λem=581 nm). Rhod-2 AM is cell-permeable version of Rhod-2. The long-wavelength Rhod-2 Ca2+ indicators are valuable alternatives to Fluo-3 for experiments in cells and tissues that have high levels of autofluorescence. Fluorescent probes that show spectral responses upon binding Ca2+ have enabled researchers to investigate changes in intracellular free Ca2+ concentrations by using fluorescence microscopy, flow cytometry, fluorescence spectroscopy and fluorescence microplate readers.


PREPARATION OF STOCK SOLUTIONS

Unless otherwise noted, all unused stock solutions should be divided into single-use aliquots and stored at -20 °C after preparation. Avoid repeated freeze-thaw cycles.
Rhod-2 AM Stock Solution
Prepare a 2 to 5 mM stock solution of Rhod-2 AM in high-quality, anhydrous DMSO.

PREPARATION OF WORKING SOLUTION

Rhod-2 AM Working Solution
On the day of the experiment, either dissolve Rhod-2 AM in DMSO or thaw an aliquot of the indicator stock solution to room temperature. Prepare a dye working solution of 2 to 20 µM in a buffer of your choice (e.g., Hanks and Hepes buffer) with 0.04% Pluronic® F-127. For most cell lines, Rhod-2 AM at a final concentration of 4-5 μM is recommended. The exact concentration of indicators required for cell loading must be determined empirically.
Note The nonionic detergent Pluronic® F-127 is sometimes used to increase the aqueous solubility of Rhod-2 AM.
Note If your cells contain organic anion-transporters, probenecid (1-2 mM) may be added to the dye working solution (final in well concentration will be 0.5-1 mM) to reduce leakage of the de-esterified indicators.

SAMPLE EXPERIMENTAL PROTOCOL

Following is our recommended protocol for loading AM esters into live cells. This protocol only provides a guideline and should be modified according to your specific needs.
  1. Prepare cells in growth medium overnight.
  2. On the next day, add 1X Rhod-2 AM working solution into your cell plate.
    Note If your compound(s) interfere with the serum, replace the growth medium with fresh HHBS buffer before dye-loading.
  3. Incubate the dye-loaded plate in a cell incubator at 37 °C for 30 to 60 minutes.
    Note Incubating the dye for longer than 1 hour can improve signal intensities in certain cell lines.
  4. Replace the dye working solution with HHBS or buffer of your choice (containing an anion transporter inhibitor, such as 1 mM probenecid, if applicable) to remove any excess probes.
  5. Add the stimulant as desired and simultaneously measure fluorescence using either a fluorescence microscope equipped with a TRITC filter set or a fluorescence plate reader containing a programmable liquid handling system such as an FDSS, FLIPR, or FlexStation, at Ex/Em = 540/590 nm cutoff 570 nm.


Product Citations
Chemical Information
Molecular Weight 1123.96
Formula C52H59BrN4O
CAS Number 145037-81-6
Solubility (25°C) DMSO 1~5mM
Storage -20°C, protect from light, dry, sealed
References

[1] Lakshmini Balachandar, et al. Curr Protoc Neurosci. Simultaneous Ca2+ Imaging and Optogenetic Stimulation of Cortical Astrocytes in Adult Murine Brain Slices

[2] Xudong Peng, et al. BMC Ophthalmol. Phospholipase Cγ2 is critical for Ca 2+ flux and cytokine production in anti-fungal innate immunity of human corneal epithelial cells

[3] Cynthia Brisac, et al. J Virol. Calcium flux between the endoplasmic reticulum and mitochondrion contributes to poliovirus-induced apoptosis

[4] Xiaoyu D, et al. Plasma Science and Technology. Measurement of cytoplasmic Ca2+ concentration in Saccharomyces cerevisiae induced by air cold plasma

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