Measurement of Effector Protein Translocation Using Phosphorylatable Epitope Tags and Phospho-Specific Antibodies.

Numerous bacterial pathogens employ specialized protein secretion machineries to directly inject anti-host proteins, termed effector proteins, into eukaryotic cells. Effector proteins carrying small phosphorylatable tags can be used to detect and quantify effector protein injection. Here, we describe the use of the ELK- and GSK-tags to detect the translocation of the Y. pestis YopE effector protein into RAW 264.7 macrophage-like cells using immunoblot analysis with phospho-specific antibodies.