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Since the production of Robert Hooke’s intricate diagrams of the microcomos in the mid-seventeenth century (Hooke, 1665), the use of the light microscope has undergone a technological revolution. Techniques and optics have greatly advanced, allowing us not only to describe the morphology of a specimen but also to probe the movement and dynamics of proteins and organelles within the cell. One of the most significant molecular and genetic advancements has been the isolation, engineering, and use of green fluorescent protein (GFP) to allow the visualization of protein fusions. In 2008, the impact GFP has had on cell biology was recognized by awarding the Nobel prize in Chemistry to the scientists involved in the pioneering initial discovery and development of its use as a fluorescent molecular tag. GFP was isolated from the jellyfish Aequorea victoria and has been expressed in a wide range of organisms including several species of plant. Subsequent engineering of GFP has resulted in multiple fluorophores with differing excitation/emission spectra allowing the visualization of two protein fusions (dual imaging) in the same cell (Shaner et al., 2007). There are numerous fluorescent protein fusions readily available to light up any organelle (Nelson et al., 2007; Geldner et al., 2009), and the generation of fusions can easily be produced using the available binary vectors (Karimi et al., 2007). This commentary briefly summarizes laser-based microscopy techniques which have expanded beyond the pure analysis of protein localization and steady-state levels, gene expression or organelle movement to allow the quantitative studies of protein and organelle dynamics. Bleach it, switch it: photobleaching, photoactivation, and photoconvertible proteins

Bleach it, switch it, bounce it, pull it: using lasers to reveal plant cell dynamics