infocus #30 June 2013 Live-Cell Fluorescence Microscopy

DOI: 10.22443/rms.inf.1.93

Single-molecule fluorescence microscopy provides a window into the molecular activity of the living cell. Through genetically fusing molecules of interest with fluorescent proteins, specific biological mechanisms can be elucidated. While the approach is maturing rapidly with the application of numerous inventive optical and
analytical techniques, it is still a challenge to generate adequate imaging conditions.
This article addresses the question of how an experiment can be optimally configured in order to produce the best possible data. We present development of a model
of the combined instrumentation and sample that concurrently and accurately computes with the relevant quantitative information. This is used to explore the effect of
parameter changes on system-level properties, such as image contrast, that are critical to achieving our experimental goals.