BCECF-AM (Synonyms: BCECF Acetoxymethyl ester) |
Catalog No.GC14603
One-Click Copy Product Info
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Intracellular ratiometric pH indicator
Products are for research use only. Not for human use. We do not sell to patients.
Cas No.: 117464-70-7
Sample solution is provided at 25 µL, 10mM.
BCECF AM ester is a widely-used fluorescent indicator for estimating intracellular pH.[1],[2] This membrane-permeant compound is modified by intracellular esterases to produce BCECF, a polar fluorescein derivative that is retained by cells. Intracellular BCECF is stable and has an efflux half-life of greater than two hours.[3] Intracellular pH measurements are made by ratio imaging, which involves determining the pH-dependent ratio of emission intensity (detected at 535 nm) when the dye is excited at 490 nm vs. the emission intensity when excited at 440 nm.[2],[4] This approach is amenable to either spectrofluorometry or flow cytometry.[2],[5] BCECF AM can also be used to investigate intracellular changes in other ions, including potassium.[6]
Reference:
[1]. Rink, T.J., Tsien, R.Y., and Pozzan, T. Cytoplasmic pH and free Mg2+ in lymphocytes. Journal of Cell Biology 95(1), 189-196 (1982).
[2]. Ozkan, P., and Mutharasan, R. A rapid method for measuring intracellular pH using BCECF-AM. Biochimica et Biophysica Acta 1572(1), 143-148 (2002).
[3]. Dive, C., Cox, H., Watson, J.V., et al. Polar fluorescein derivatives as improved substrate probes for flow cytoenzymological assay of cellular esterases. Mol.Cell Probe 2(2), 131-145 (1988).
[4]. O'Connor, N., and Silver, R.B. Ratio imaging: Practical considerations for measuring intracellular Ca2+ and pH in living cells. Methods Cell Biol. 81, 415-433 (2007).
[5]. Chow, S., Hedley, D., and Tannock, I. Flow cytometric calibration of intracellular pH measurements in viable cells using mixtures of weak acids and bases. Cytometry 24, 360-367 (1996).
[6]. Balkay, L., Márián, T., Emri, M., et al. Flow cytometric determination of intracellular free potassium concentration. Cytometry 28, 42-49 (1997).
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Cell experiment: |
PASMCs are placed in a laminar flow cell chamber perfused with HBSS with pH adjusted to 7.4. pHi is measured in cells incubated with the membrane permeant (acetoxymethyl ester) form of the pH-sensitive fluorescent dye BCECF-AM for 60 min at 37°C under an atmosphere of 20% O2-5% CO2. Cells are then washed with HBSS for 15 min at 37°C to remove extracellular dye and allow complete de-esterification of cytosolic dye. Ratiometric measurement of BCECF fluorescence is performed on a workstation consisting of a Nikon TSE 100 Ellipse inverted microscope with epi-fluorescence attachments. The light beam from a xenon arc lamp is filtered by interference filters at 490 and 440 nm, and focused onto the PASMCS under examination via a 20× fluorescence objective. Light emitted from the cell at 530 nm is returned through the objective and detected by an imaging camera. An electronic shutter is used to minimize photobleaching of dye. Protocols are executed and data collected on-line with InCyte software. pHi is estimated from in situcalibration after each experiment. Cells are perfused with a solution containing (in mM): 105 KCl, 1 MgCl2, 1.5 CaCl2, 10 glucose, 20 HEPES-Tris and 0.01 nigericin to allow pHi to equilibrate to external pH. A two point calibration is created from fluorescence measured as pHi is adjusted with KOH from 6.5 to 7.5. Intracellular H+ ion concentration ([H+]i) is determined from pHi using the formula: pHi = −log ([H+]i). |
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References: [1]. Clark Undem, et al. Endothelin-1 Augments Na+/H+ Exchange Activity in Murine Pulmonary Arterial Smooth Muscle Cells via Rho Kinase. PLoS One. 2012; 7(9): e46303. |
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| Cas No. | 117464-70-7 | SDF | |
| Synonyms | BCECF Acetoxymethyl ester | ||
| Chemical Name | 3',6'-bis[(acetyloxy)methoxy]-5(or 6)-[[(acetyloxy)methoxy]carbonyl]-3-oxo-spiro[isobenzofuran-1(3H),9'-[9H]xanthene]-2',7'-dipropanoic acid, 2',7'-bis[(acetyloxy)methyl] ester | ||
| Canonical SMILES | O=C(C)OCOC1=C(CCC(OCOC(C)=O)=O)C=C(C2(C(C=CC(C(OCOC(C)=O)=O)=C3)=C3C(O2)=O)C(C(O4)=C5)=CC(CCC(OCOC(C)=O)=O)=C5OCOC(C)=O)C4=C1.O=C(C)OCOC6=C(CCC(OCOC(C)=O)=O)C=C(C7(C(C=C(C(OCOC(C)=O)=O)C=C8)=C8C(O7)=O)C(C(O9)=C%10)=CC(CCC(OCOC(C)=O)=O)=C%10OCOC(C)=O)C9=C6 | ||
| Formula | C42H40O21 | M.Wt | 880.7 |
| Solubility | Soluble in ,Acetonitrile, DMF, DMSO | Storage | Store at -20°C, protect from light,unstable in solution, ready to use. |
| General tips | Please select the appropriate solvent to prepare the stock solution according to the
solubility of the product in different solvents; once the solution is prepared, please store it in
separate packages to avoid product failure caused by repeated freezing and thawing.Storage method
and period of the stock solution: When stored at -80°C, please use it within 6 months; when stored
at -20°C, please use it within 1 month. To increase solubility, heat the tube to 37°C and then oscillate in an ultrasonic bath for some time. |
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| Shipping Condition | Evaluation sample solution: shipped with blue ice. All other sizes available: with RT, or with Blue Ice upon request. | ||
| Prepare stock solution | |||
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1 mg | 5 mg | 10 mg |
| 1 mM | 1.1355 mL | 5.6773 mL | 11.3546 mL |
| 5 mM | 227.1 μL | 1.1355 mL | 2.2709 mL |
| 10 mM | 113.5 μL | 567.7 μL | 1.1355 mL |
Step 1: Enter information below (Recommended: An additional animal making an allowance for loss during the experiment)
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Working concentration: mg/ml;
Method for preparing DMSO master liquid: mg drug pre-dissolved in μL DMSO ( Master liquid concentration mg/mL, Please contact us first if the concentration exceeds the DMSO solubility of the batch of drug. )
Method for preparing in vivo formulation: Take μL DMSO master liquid, next addμL PEG300, mix and clarify, next addμL Tween 80, mix and clarify, next add μL saline, mix and clarify.
Method for preparing in vivo formulation: Take μL DMSO master liquid, next add μL Corn oil, mix and clarify.
Note: 1. Please make sure the liquid is clear before adding the next solvent.
2. Be sure to add the solvent(s) in order. You must ensure that the solution obtained, in the previous addition, is a clear solution before proceeding to add the next solvent. Physical methods such as vortex, ultrasound or hot water bath can be used to aid dissolving.
3. All of the above co-solvents are available for purchase on the GlpBio website.
Quality Control & SDS
- View current batch:
- Purity: >95.00% Appearance: An oil
- COA (Certificate of Analysis)
- SDS (Safety Data Sheet)
- Datasheet
Average Rating: 5 (Based on Reviews and 33 reference(s) in Google Scholar.)