title: Imaging callose at plasmodesmata using aniline blue: quantitative confocal microscopy.
abstract: Callose (β-1,3-glucan) is both structural and functional component of plasmodesmata (Pd). The turnover of callose at Pd controls the cell-to-cell diffusion rate of molecules through Pd. An accurate assessment of changes in levels of Pd-associated callose has become a first-choice experimental approach in the research of intercellular communication in plants.Here we describe a detailed and easy-to-perform procedure for imaging and quantification of Pd-associated callose using fixed plant tissue stained with aniline blue. We also introduce an automated image analysis protocol for non-biased quantification of callose levels at Pd from fluorescence images using ImageJ. Two experimental examples of Pd-callose quantification using the automated method are provided as well.
authors.affiliation: Department of Molecular Biology and Ecology of Plants, George S. Wise Faculty of Life Sciences, Tel Aviv University, Tel Aviv, 69978, Israel.
authors.forename: Raul
authors.initials: R
authors.lastname: Zavaliev
authors.forename: Bernard L
authors.initials: BL
authors.lastname: Epel
mesh_terms: D000814:Aniline Compounds / Q000737:chemistry*; D017360:Arabidopsis / Q000737:chemistry / Q000648:ultrastructure*; D001692:Biological Transport; D002473:Cell Wall / Q000737:chemistry / Q000648:ultrastructure*; D044602:Cellulases / Q000737:chemistry; D005404:Fixatives / Q000737:chemistry; D005936:Glucans / Q000032:analysis* / Q000096:biosynthesis; D007091:Image Processing, Computer-Assisted / Q000379:methods; D018613:Microscopy, Confocal / Q000379:methods; D008867:Microtomy; D018515:Plant Leaves / Q000737:chemistry / Q000648:ultrastructure*; D031425:Plasmodesmata / Q000737:chemistry / Q000648:ultrastructure*; D012984:Software; D016707:Tissue Fixation; D014026:Nicotiana / Q000737:chemistry / Q000648:ultrastructure*
chemical_list: D000814:Aniline Compounds; D005404:Fixatives; D005936:Glucans; C017006:aniline blue; C048306:callose; D044602:Cellulases