期刊
ANALYTICAL AND BIOANALYTICAL CHEMISTRY
卷 393, 期 4, 页码 1123-1133出版社
SPRINGER HEIDELBERG
DOI: 10.1007/s00216-008-2489-7
关键词
Biosensor; Endocytosis; GFP; In vivo imaging; pH measurement; Tat-HIV protein
资金
- Italian Ministry for University and Research [RBLA03ER38]
- Fondazione Monte dei Paschi di Siena
Genetically encoded fluorescent proteins are optimal reporters when used to monitor cellular processes as they can be targeted to any subcellular region by fusion to a protein of interest. Here, we present the pH-sensitive fluorescent protein E(1)GFP which is ideally suited to monitor pH changes in dynamic intracellular structures in real time with high spatio temporal resolution. E(1)GFP is a ratiometric pH indicator by emission with a pK close to 6.0. We describe an application of this novel pH reporter in the measurement of pH changes along the endo-lysosomal pathway. By fusing E(1)GFP to the HIV-Tat protein which is endowed with cell-penetrating properties, we were able to monitor multi-step endocytosis from the initial cell-surface binding through to the intracellular endocytic network in real time. This represents a framework for the application of E(1)GFP to the in situ detection of pH changes involved in dynamic biological phenomena.
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