title: Using Fluorescence Recovery After Photobleaching to Study Gap Junctional Communication In Vitro.
abstract: Fluorescence recovery after photobleaching (FRAP) is a microscopy-based technique to study the movement of fluorescent molecules inside a cell. Although initially developed to investigate intracellular mobility, FRAP can be also used to measure intercellular dynamics. This chapter describes how to perform FRAP experiment to study gap junctional communication in living cells. The procedures described here can be carried out with a laser-scanning confocal microscope and any in vitro cultured cells known to communicate via gap junctions. In addition, the method can be easily adjusted to measure gap junction function in 3D cell cultures as well as ex vivo tissue.
authors.affiliation: Nuffield Department of Orthopaedics, Rheumatology and Musculoskeletal Sciences, University of Oxford, Windmill Road, Oxford, OX3 7HE, UK. maria.kuzmakuzniarska@gmail.com.
authors.forename: Maria
authors.initials: M
authors.lastname: Kuzma-Kuzniarska
authors.affiliation: Structural Genomics Consortium, University of Oxford, Oxford, UK.
authors.forename: Clarence
authors.initials: C
authors.lastname: Yapp
authors.affiliation: Nuffield Department of Orthopaedics, Rheumatology and Musculoskeletal Sciences, University of Oxford, Windmill Road, Oxford, OX3 7HE, UK.
authors.forename: Philippa A
authors.initials: PA
authors.lastname: Hulley
mesh_terms: D002450:Cell Communication / Q000502:physiology*; D018929:Cell Culture Techniques / Q000379:methods; D002478:Cells, Cultured; D005452:Fluoresceins / Q000737:chemistry; D036681:Fluorescence Recovery After Photobleaching / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry; D017629:Gap Junctions / Q000502:physiology*; D006801:Humans; D018613:Microscopy, Confocal; D013194:Staining and Labeling / Q000379:methods; D000070916:Tenocytes
chemical_list: D005452:Fluoresceins; D005456:Fluorescent Dyes; C085925:calcein AM
keywords: Calcein; Confocal microscopy; Fluorescence recovery after photobleaching; Fluorescence recovery curve; Gap junctions; In vitro; Intercellular communication; Mobile fraction percentage; Monolayer; Tenocytes