期刊
METHODS
卷 46, 期 3, 页码 233-238出版社
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/j.ymeth.2008.09.016
关键词
Exocytosis; Fluorescent protein; Hormone secretion; Live cell imaging; Total internal reflection fluorescence microscopy
资金
- Wellcome Trust
- Medical Research Council (UK)
- National Institutes of Health (US)
- European Union (FP6)
- Juvenile Diabetes Research Foundation
- Human Frontiers Science Program
- Ministry of Education, Culture, Sports, and Technology of Japan [18689008]
- Mochida Memorial Foundation for Medical and Pharmaceutical Research
- Uehara Memorial Foundation
- Research Foundation for Opto-Science and Technology
- Diabetes UK
- Grants-in-Aid for Scientific Research [18689008] Funding Source: KAKEN
Ca2+ ions are the most ubiquitous second messenger found in all cells, and play a significant role in controlling regulated secretion from neurons, endocrine, neuroendocrine and exocrine cells. Here, we describe microscopic techniques to image regulated secretion, a target of Ca2+ signalling. The first of these, total internal reflection fluorescence (TIRF), is well suited for optical sectioning at cell-substrate regions with an unusually thin region of fluorescence excitation (<150 nm). It is thus particularly useful for studies of regulated hormone secretion. A brief summary of this approach is provided, as well as a description of the physical basis for the technique and the tools to implement TIRF using a standard fluorescence microscope. We also detail the different fluorescent probes which can be used to detect secretion and how to analyze the data obtained. A comparison between TIRF and other imaging modalities including confocal and multiphoton microscopy is also included. (C) 2008 Elsevier Inc. All rights reserved.
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