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Use of pluronic acid F‐127 with Fluo‐3/AM probe to determine intracellular calcium changes elicited in bean protoplasts
Author(s) -
Granados M. E.,
Soriano E.,
SaavedraMolina A.
Publication year - 1997
Publication title -
phytochemical analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.574
H-Index - 72
eISSN - 1099-1565
pISSN - 0958-0344
DOI - 10.1002/(sici)1099-1565(199707)8:4<204::aid-pca354>3.0.co;2-l
Subject(s) - chemistry , poloxamer , intracellular , vacuole , calcium , hydrolysis , protoplast , biophysics , cytosol , calcium in biology , fluorescence , chromatography , biochemistry , cytoplasm , organic chemistry , enzyme , physics , quantum mechanics , copolymer , biology , polymer
Measurement of cytosolic calcium concentration in protoplasts of bean leaf was carried out using Fluo‐3/AM indicator. The uptake of the fluorescent indicator, as its permeable ester, was achieved by employing pluronic acid F‐127 in the incubating solution. In the absence of pluronic acid F‐127 loading was two times lower in the same time period. A rapid intracellular location of the fluorescent dye after intracellular hydrolysis is reported, with no vacuole trapping of the fluorescent indicator. An increase from 54 to 91 n M free calcium ([Ca 2+ ] i ) was observed immediately after addition of a pectic elicitor. Advantages of the use of pluronic acid F‐127 are in the speed of the indicator entry, eliminating any possible hydrolysis by external esterases, and more efficient loading while maintaining cell integrity. © 1997 John Wiley & Sons, Ltd.