Spectrally And Temporally Resolved Low-Light Level Video Microscopy

John E. Wampler; Ruth Furukawa; Marcus Fechheimer

doi:10.1117/12.962696

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22 December 1989 Spectrally And Temporally Resolved Low-Light Level Video Microscopy

John E. Wampler, Ruth Furukawa, Marcus Fechheimer

Proceedings Volume 1161, New Methods in Microscopy and Low Light Imaging; (1989) https://doi.org/10.1117/12.962696
Event: 33rd Annual Technical Symposium, 1989, San Diego, United States

Abstract

The IDG law-light video microscope system was designed to aid studies of localization of subcellular luminescence sources and stimulus/response coupling in single living cells using luminescent probes. Much of the motivation for design of this instrument system came from the pioneering efforts of Dr. Reynolds (Reynolds, Q. Rev. Biophys. 5, 295-347; Reynolds and Taylor, Bioscience 30, 586-592) who showed the value of intensified video camera systems for detection and localizion of fluorescence and bioluminescence signals from biological tissues. Our instrument system has essentially two roles, 1) localization and quantitation of very weak bioluminescence signals and 2) quantitation of intracellular environmental characteristics such as pH and calcium ion concentrations using fluorescent and bioluminescent probes. The instrument system exhibits over one million fold operating range allowing visualization and enhancement of quantum limited images with quantum limited response, spectral analysis of fluorescence signals, and transmitted light imaging. The computer control of the system implements rapid switching between light regimes, spatially resolved spectral scanning, and digital data processing for spectral shape analysis and for detailed analysis of the statistical distribution of single cell measurements. The system design and software algorithms used by the system are summarized. These design criteria are illustrated with examples taken from studies of bioluminescence, applications of bioluminescence to study developmental processes and gene expression in single living cells, and applications of fluorescent probes to study stimulus/response coupling in living cells.

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John E. Wampler, Ruth Furukawa, and Marcus Fechheimer "Spectrally And Temporally Resolved Low-Light Level Video Microscopy", Proc. SPIE 1161, New Methods in Microscopy and Low Light Imaging, (22 December 1989); https://doi.org/10.1117/12.962696

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