Use of pluronic acid F-127 with Fluo-3/AM probe to determine intracellular calcium changes elicited in bean protoplasts
✍ Scribed by M. E. Granados; E. Soriano; A. Saavedra-Molina
- Book ID
- 101284407
- Publisher
- John Wiley and Sons
- Year
- 1997
- Tongue
- English
- Weight
- 127 KB
- Volume
- 8
- Category
- Article
- ISSN
- 0958-0344
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✦ Synopsis
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 nM 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.