Single-cell quantification of molecules and rates using open-source microscope-based cytometry

摘要: Microscope-based cytometry provides a powerful means to study cells in high throughput. Here we present set of refined methods for making sensitive measurements large numbers individual Saccharomyces cerevisiae over time. The consists relatively simple 'wet' methods, microscope procedures, open-source software tools and statistical routines. This combination is very sensitive, allowing detection measurement fewer than 350 fluorescent protein molecules per living yeast cell. These enabled new protocols, including 'snapshot' protocols calculate rates maturation degradation molecular species, GFP derivative native mRNA, unperturbed, exponentially growing cells. Owing their sensitivity, accuracy ability track changes time, these may complement flow-cytometric studies the quantitative physiology cellular systems.