| Fluorescence has become an invaluable tool for cell biologists to address a wide variety of biological questions in live cells. Although fluorescent proteins provide a means to study proteins in their native environment, they have shortcomings. In this thesis, I describe a recently developed technology based on fluorogen activating protein as an alternative to fluorescent proteins and its unique advantages in a number of applications. In Chapter I, I present an overview of the chemical tagging strategies. At the end of chapter I, I describe the characteristics of FAPs and fluorogen, the distinct features of a few tandem dyes and the use of FAP as a hybrid probe. In Chapter II, I present FAP based super-resolution imaging techniques, in two imaging modalities: STED microscopy and localization based single molecule imaging. In Chapter III, I present an improved cell surface labeling strategy based on FAP and a recently developed fluorogen. Lastly, in Chapter IV, I present a new FAP based tandem dye and its applications in quantification of receptor trafficking and pH sensing. |
