title: Recombinant differential anchorage probes that tower over the spatial dimension of intracellular signals for high content screening and analysis.
abstract: Recombinant fluorescent probes allow the detection of molecular events inside living cells. Many of them exploit the intracellular space to provide positional signals and, thus, require detection by single cell imaging. We describe here a novel strategy based on probes capable of encoding the spatial dimension of intracellular signals into "all-or-none" fluorescence intensity changes (differential anchorage probes, DAPs). The resulting signals can be acquired in single cells at high throughput by automated flow cytometry, (i) bypassing image acquisition and analysis, (ii) providing a direct quantitative readout, and (iii) allowing the exploration of large experimental series. We illustrate our purpose with DAPs for Bax and the effector caspases 3 and 7, which are keys players in apoptotic cell death, and show applications in basic research, high content multiplexed library screening, compound characterization, and drug profiling.
authors.affiliation: INSERM U916, Institut Bergonié, Université Victor Segalen, 229 cours de l'Argonne, 33076 Bordeaux, France.
authors.forename: Laura
authors.initials: L
authors.lastname: Schembri
authors.forename: Marion
authors.initials: M
authors.lastname: Zanese
authors.forename: Gaelle
authors.initials: G
authors.lastname: Depierre-Plinet
authors.forename: Muriel
authors.initials: M
authors.lastname: Petit
authors.forename: Assia
authors.initials: A
authors.lastname: Elkaoukabi-Chaibi
authors.forename: Loic
authors.initials: L
authors.lastname: Tauzin
authors.forename: Cristina
authors.initials: C
authors.lastname: Florean
authors.forename: Lydia
authors.initials: L
authors.lastname: Lartigue
authors.forename: Chantal
authors.initials: C
authors.lastname: Medina
authors.forename: Christophe
authors.initials: C
authors.lastname: Rey
authors.forename: Francis
authors.initials: F
authors.lastname: Belloc
authors.forename: Josy
authors.initials: J
authors.lastname: Reiffers
authors.forename: François
authors.initials: F
authors.lastname: Ichas
authors.forename: Francesca
authors.initials: F
authors.lastname: De Giorgi
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D017209:Apoptosis; D053148:Caspase 3 / Q000378:metabolism; D053179:Caspase 7 / Q000378:metabolism; D002453:Cell Cycle; D016923:Cell Death; D045744:Cell Line, Tumor; D002463:Cell Membrane Permeability; D049109:Cell Proliferation; D055808:Drug Discovery; D005434:Flow Cytometry; D005456:Fluorescent Dyes / Q000378:metabolism*; D057166:High-Throughput Screening Assays; D006801:Humans; D042541:Intracellular Space / Q000378:metabolism*; D057054:Molecular Imaging; D008969:Molecular Sequence Data; D010447:Peptide Hydrolases / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism*; D051028:bcl-2-Associated X Protein / Q000378:metabolism
chemical_list: D005456:Fluorescent Dyes; D011994:Recombinant Proteins; D051028:bcl-2-Associated X Protein; D010447:Peptide Hydrolases; D053148:Caspase 3; D053179:Caspase 7