Description
BCECF AM ester is a widely-
used fluorescent indicator for estimating intracellular pH. 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. 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. This approach is amenable to either spectrofluorometry or flow cytometry. BCECF AM can also be used to investigate intracellular changes in other ions, including potassium.
Chemical Properties
pale yellow to yellow powder
Uses
Cell-permeable ester of BCECF that, on hydrolysis by cytosolic esterases, yields the intracellularly trapped pH-indicator BCECF. Allows simultaneous recording of cell volume changes and intracellular pH in single osteosarcoma cells.
General Description
Material is an isomeric mixture of lactones, acetates, and AM esters
Biological Activity
Non-charged, non-fluorescent pH indicator. Readily and rapidly diffuses through cell membranes.
References
[1] T J RINK T P R Y Tsien. Cytoplasmic pH and free Mg2+ in lymphocytes.[J]. Journal of Cell Biology, 1982, 95 1: 189-196. DOI:
10.1083/jcb.95.1.189[2] PINAR OZKAN Raj M. A rapid method for measuring intracellular pH using BCECF-AM[J]. Biochimica et biophysica acta. General subjects, 2002, 1572 1: Pages 143-148. DOI:
10.1016/s0304-4165(02)00303-3[3] CAROLINE DIVE. Polar fluorescein derivatives as improved substrate probes for flow cytoenzymological assay of cellular esterases[J]. Molecular and Cellular Probes, 1988, 2 2: Pages 131-145. DOI:
10.1016/0890-8508(88)90035-7[4] NATHAN O’CONNOR Randi B S. Ratio imaging: practical considerations for measuring intracellular Ca2+ and pH in living cells.[J]. Methods in cell biology, 2013, 114: 387-406. DOI:
10.1016/b978-0-12-407761-4.00016-6[5] SUE CHOW Ian T David Hedley. Flow cytometric calibration of intracellular pH measurements in viable cells using mixtures of weak acids and bases[J]. Cytometry Part B: Clinical Cytometry, 1996, 24 4: 360-367. DOI:
10.1002/(sici)1097-0320(19960801)24:4<360::aid-cyto7>3.0.co;2-j[6] LáSZLó BALKAY. Flow cytometric determination of intracellular free potassium concentration[J]. Cytometry Part B: Clinical Cytometry, 1998, 28 1: 42-49. DOI:
10.1002/(sici)1097-0320(19970501)28:1<42::aid-cyto5>3.0.co;2-j
Abs/Em Maxima
440; 490/530 nm
Excitation / Emission Maximum
440-490 nm/535 nm
