AAT Bioquest® Product Technical Information Sheet Last Updated July 2013 TMRE [Tetramethylrhodamine ethyl ester] Ordering Information Product Numbers: 22220 (25 mg) Storage Conditions Avoid exposure to light Keep at -20 °C and desiccated Chemical and Physical Properties Molecular Weight: 514.95 Appearance: Red powder Solvent: DMSO Spectral Properties: Excitation = 549 nm; Emission = 574 nm. Biological Applications Positively charged rhodamine dyes (such as rhodamine esters and rosamines) are selectively localized in mitochondria, thus they are widely used for labeling mitochondria of live cells. Like JC-10 and JC-1, TMRM and TMRE are widely used for measuring mitochondrial membrane potential besides their selective mitochondrial staining. These two particular rhodamine esters stain mitochondria in orange fluorescence. Their spectral properties are similar to those of TRITC, making the use of TMRM and TMRE quite convenient. TMRE is slightly more hydrophobic than TMRM. Sample Protocol for Staining Cells The following procedure can be adapted for most cell types. Growth medium, cell density, the presence of other cell types and other factors may influence staining. Residual detergent on glassware may also affect real or apparent staining of many organisms, causing brightly stained material to appear in solutions with or without cells present. 1). Make 1-10 mM DMSO stock solution. The DMSO stock solution is good for 6 months if stored at -20 °C. 2). Seed cell line(s) and run treatment(s) for desired amount of time. Optional: 10 min before adding TMRE (next step), add 100nM to 10 µM FCCP to one sample in media as positive control. 3). Add TMRE to cells in media to final concentration of 50-1000 nM. 4). Incubate 15 to 30 min at 37 °C 5). Measure the fluorescence intensities with ex/em = 540/590 nm (cut off at 570 nm) References 1. Criddle DN, Murphy J, Fistetto G, Barrow S, Tepikin AV, Neoptolemos JP, Sutton R, Petersen OH. (2006) Fatty acid ethyl esters cause pancreatic calcium toxicity via inositol trisphosphate receptors and loss of ATP synthesis. Gastroenterology, 130, 781. 2. Forster K, Paul I, Solenkova N, Staudt A, Cohen MV, Downey JM, Felix SB, Krieg T. (2006) NECA at reperfusion limits infarction and inhibits formation of the mitochondrial permeability transition pore by activating p70S6 kinase. Basic Res Cardiol, 101, 319. 3. Greco NJ, Seetharaman S, Kurtz J, Lee WR, Moroff G. (2006) Evaluation of the reactivity of apoptosis markers before and after cryopreservation in cord blood CD34(+) cells. Stem Cells Dev, 15, 124. ©2012 by AAT Bioquest®, Inc., 520 Mercury Drive, Sunnyvale, CA 94085 Ordering: [email protected]; Tel: 800-990-8053; Fax: 408-733-1304 Technical Support: [email protected]; Tel: 408-733-1055