Author/Editor		Mason, WT; Hoyland, J; Neylon, CB; Kato, M; Akerman, S; Bunting, R; Tregear, RT; Zorec, R
Title		Dynamic, real time imaging of fluorescent probes of biological activity in living cells
Type		članek
Source		J Med Lab Sciences
Vol. and No.		Letnik 5, št. 1
Publication year		1991
Volume		str. 41-52
Language		eng
Abstract		The development of ion-sensitive fluorochromes has given rise to an excitting new field of cell biology. These probes can be introduced into the cytoplasm of living cells and as the ionic concentration of the cell interior changes, the dyes undergo changes in fluorescence consisting of wavelength shifts and changes in quantum efficiency. The preferred fluorochromes undergo changes at two or more walvelenghts, and this property can be utilised with ratio measurements of fluorescence intensity to eliminate artivacts due to dye loading or cell tickness. The availability of these fluorochromes has led to the development of low light level detection technologies for capturing and analysing the resulting signals. The most powerful approaoch is the use of low light level charge coupled device cameras, which effectively provides an array of several hundred thousand photodetectors. Such measurements can be made at high speeds, up to the video frame rate of about 25 Hz or one image grame every 40 miliseconds. When interfaced to a computer and the analogue camera signals thus digitised, poweful image processing and image analysis techniques can be employed to yield spatial and temporal estimates of ion concentrations in single or multiple living cells. This article will describe the technology required to perform these experiment, the advantages and disadvantages of the approach and some applications from our laboratory using dynamic ratio imaging.
Descriptors		CELL CYCLE IONS RECEPTORS, CELL SURFACE ANTIBODIES FLUORESCENT DYES MICROSCOPY, FLUORESCENCE IMAGE PROCESSING, COMPUTER-ASSISTED TISSUE CULTURE