title: Probing Protein Distribution Along the Nuclear Envelope In Vivo by Using Single-Point FRAP.
abstract: Determining the locations of nuclear envelope transmembrane proteins and their concentrations across the outer and inner nuclear membranes has been a challenging and time-consuming process. Typically, this required the week-long process of fixing and immunogold staining of cells prior to analysis by electron microscopy. Here, we describe a method, single-point fluorescence recovery after photobleaching (spFRAP), which is able to quickly determine the localization and distribution of nuclear membrane proteins along the double nuclear envelope membranes with a precision of 10-15 nm in a matter of 10-20 min the day after transfection.
authors.affiliation: Department of Biology, Temple University, 1900 North 12th St., Philadelphia, PA, 19122, USA.
authors.forename: Krishna C
authors.initials: KC
authors.lastname: Mudumbi
authors.affiliation: Department of Biology, Temple University, 1900 North 12th St., Philadelphia, PA, 19122, USA. weidong.yang@temple.edu.
authors.forename: Weidong
authors.initials: W
authors.lastname: Yang
mesh_terms: D036681:Fluorescence Recovery After Photobleaching; D015870:Gene Expression; D017930:Genes, Reporter; D006367:HeLa Cells; D006801:Humans; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009685:Nuclear Envelope / Q000378:metabolism*; D021381:Protein Transport; D014162:Transfection
chemical_list: D008565:Membrane Proteins
keywords: Fluorescence recovery after photobleaching; Nuclear envelope transmembrane proteins; Single molecule; Super-resolution