title: Standing-wave-excited multiplanar fluorescence in a laser scanning microscope reveals 3D information on red blood cells.
abstract: Standing-wave excitation of fluorescence is highly desirable in optical microscopy because it improves the axial resolution. We demonstrate here that multiplanar excitation of fluorescence by a standing wave can be produced in a single-spot laser scanning microscope by placing a plane reflector close to the specimen. We report here a variation in the intensity of fluorescence of successive planes related to the Stokes shift of the dye. We show by the use of dyes specific for the cell membrane how standing-wave excitation can be exploited to generate precise contour maps of the surface membrane of red blood cells, with an axial resolution of ≈90 nm. The method, which requires only the addition of a plane mirror to an existing confocal laser scanning microscope, may well prove useful in studying diseases which involve the red cell membrane, such as malaria.
authors.affiliation: Centre for Biophotonics, Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, 161 Cathedral Street, Glasgow G4 0RE, United Kingdom.
authors.forename: Rumelo
authors.initials: R
authors.lastname: Amor
authors.affiliation: Institute of Life Sciences and Department of Chemistry, University of Southampton, Highfield Campus, Southampton SO17 1BJ, United Kingdom.
authors.forename: Sumeet
authors.initials: S
authors.lastname: Mahajan
authors.affiliation: MRC Laboratory of Molecular Biology, Francis Crick Avenue, Cambridge Biomedical Campus, Cambridge CB2 2QH, United Kingdom.
authors.forename: William Bradshaw
authors.initials: WB
authors.lastname: Amos
authors.affiliation: Centre for Biophotonics, Strathclyde Institute of Pharmacy and Biomedical Sciences, University of Strathclyde, 161 Cathedral Street, Glasgow G4 0RE, United Kingdom.
authors.forename: Gail
authors.initials: G
authors.lastname: McConnell
mesh_terms: D000818:Animals; D004912:Erythrocytes / Q000166:cytology*; D051379:Mice; D018613:Microscopy, Confocal / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods