Confocal FRET Microscopy: Study of Clustered Distribution of Receptor–Ligand Complexes in Endocytic Membranes
Year: 2005
Type: book-chapter
Abstract: This chapter discusses the use of confocal Forster resonance (radiationless) energy transfer (C-FRET) microscopic techniques to study the clustered distribution of receptor–ligand complexes in endocytic membranes. Confocal microscopy is capable of producing images free of out-of-focus information. It provides a significant improvement in lateral resolution and in the capacity for direct, noninvasive serial optical sectioning of intact, thick living specimens. FRET microscopy helps to follow the endocytic and transcytotic transport of membrane-bound receptors internalized from opposite membranes in the epithelial polarized MDCK cell. In FRET-based assay of receptor distribution and sorting, pIgA-R ligands, labeled with different fluorophores, are internalized from opposite PMs in the MDCK cells, which are stably transfected with rabbit pIgA-R. Upon binding, these receptor–ligand complexes are transcytosed from the basolateral PM and endocytosed from the apical PM to eventually co-localize in the apical recycling endosomal compartment. The receptor distribution and sorting the FRET-based assay provides a snapshot of receptor distribution and sorting throughout the endocytic pathway of polarized cells.
Source: Molecular Imaging
Authors: HORST WALLRABE, MARGARIDA BARROSO
Language: English
Subfield: Cell Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $2,175
OpenAlex ID: W137816197