Bacterial pathogens deliver effector proteins into host cells by deploying sophisticated secretion systems. This effector translocation during host-pathogen interactions is a prerequisite for the manipulation of host cells and organisms and is important for pathogenesis. Analyses of dynamics and kinetics of translocation, subcellular localization, and cellular targets of effector proteins lead to understanding the mode of action and function of effector proteins in host-pathogen interplay. This review provides an overview of biochemical and genetic tools that have been developed to study protein effector translocation qualitatively or quantitatively. After introducing the challenges of analyses of effector translocation during host-pathogen interaction, we describe various methods ranging from static visualization in fixed cells to dynamic live-cell imaging of effector protein translocation. We show the main findings enabled by the approaches, emphasize the advantages and limitations of the methods, describe recent approaches that allow real-time tracking of effector proteins in living cells on a single molecule level, and highlight open questions in the field to be addressed by application of new methods.
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