title: Single-cell investigation by laser scanning confocal microscopy of cytochemical alterations resulting from extracellular oxidant challenge.
abstract: Confocal laser scanning fluorescence microscopy coupled to image analysis was employed in order to develop and evaluate procedures for the appraisal at the single-cell level of: (1) protein-bound 4-hydroxynonenal, the specific product of membrane peroxidation (by means of immunocytochemistry with biotin-avidin revelation); (2) protein oxidation (by reaction of protein carbonyls with 2, 4-dinitrophenyl-hydrazine followed by immunocytochemistry of dinitrophenyl moieties); and (3) cellular protein thiols (by direct alkylation of sulfhydryl groups with thiol-specific fluorescent reagents possessing different cell permeabilities). The procedures proved able to reveal the subcellular distribution of cytochemical parameters useful as indices of oxidative stress conditions, and may allow "redox phenotyping" of isolated cells, which would provide an efficient tool in selected experimental models.
authors.affiliation: Istituto di Patologia Generale, Università di Siena, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Pompella
authors.forename: C
authors.initials: C
authors.lastname: Cambiaggi
authors.forename: S
authors.initials: S
authors.lastname: Dominici
authors.forename: A
authors.initials: A
authors.lastname: Paolicchi
authors.forename: R
authors.initials: R
authors.lastname: Tongiani
authors.forename: M
authors.initials: M
authors.lastname: Comporti
mesh_terms: D000447:Aldehydes / Q000276:immunology / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D002478:Cells, Cultured; D019084:Fluorescent Antibody Technique, Indirect; D006801:Humans; D007091:Image Processing, Computer-Assisted; D008214:Lymphocytes / Q000166:cytology / Q000187:drug effects* / Q000648:ultrastructure; D008297:Male; D008565:Membrane Proteins / Q000378:metabolism; D018613:Microscopy, Confocal; D009504:Neutrophils / Q000166:cytology / Q000187:drug effects* / Q000648:ultrastructure; D016877:Oxidants / Q000494:pharmacology*; D011817:Rabbits; D051381:Rats; D017207:Rats, Sprague-Dawley; D013438:Sulfhydryl Compounds; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured
chemical_list: D000447:Aldehydes; D008565:Membrane Proteins; D016877:Oxidants; D013438:Sulfhydryl Compounds; C027576:4-hydroxy-2-nonenal; D013755:Tetradecanoylphorbol Acetate