Labeling of platelet-derived growth factor by reversible biotinylation to visualize its endocytosis by microscopy

Abstract

Microscopical analyses of endocytic trafficking require tools for efficient detection of internalized cargo. Due to the lack of suitable reagents and limitations related to its biological properties, visualization of platelet-derived growth factor (PDGF) by microscopy remained a challenge. To overcome these restrictions, we generated a biologically active PDGF labeled with up to five biotins on cleavable linkers. Subsequently, we stimulated cells with such ligand followed by removal of extracellular biotins. PDGF captured in endocytic vesicles was successfully detected with antibiotin antibodies with parallel detection of PDGF receptor, as well as other markers of endocytic compartments. Labeled PDGF was successfully validated and can be utilized in various microscopical techniques.

Keywords: Endocytosis; Labeling; Microscopy; PDGF; Reversible biotinylation.