ONE- OR TWO-PHOTON STREAK-FLIM

FLIM

A unique tool for characterizing interactions

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Telephone : +33 5 34 32 38 37
Streak-FLIM (Fluorescence Lifetime Imaging Microscopy) is a unique technique developed by the FRAIB imaging platform, which makes it possible to reveal protein-protein and protein-DNA interactions, or changes in spatial conformation. If they are close enough to interact, molecules transfer energy to each other and this can be measured through fluorescence lifetime, providing a wealth of useful information. These measures are nevertheless independent of the concentration of the molecule and its fluorescence intensity.

Services:

FLIM

  • Detection of molecular interactions (protein – protein, protein – DNA, etc.) by measuring the fluorescence lifetime of the donor in the absence or presence of an acceptor.
  • Detection by a Streak camera (Hamamatsu Streakscope) for measuring the lifetime, with a resolution on the order of a picosecond.
  • Variable time acquisition (from seconds to tens of seconds) according to the fluorescence intensity and the type of information sought (lifetime by ROI, or pixel-based).

Access mode:

  • With assistance
  • As collaborative project
  • As service delivery

Technical characteristics:

    • Nikon TE2000 inverted microscope
    • Images in x, t (t in variable full scale according to the voltage ramp applied to the deflection electrodes from 1 ns to 1 µs).
    • Images in x, t with stacks of images in y.
    • Obtaining the fluorescence decay curves for one ROI or per pixel.
    • Determining the ai and ti parameters by least squares adjustment of the obtained experimental curves (nonlinear least squares or maximum likelihood).

Available resourcesClic to contact

Name
Contact
Location

Two-photon FLIM

  • Tsunami Spectra Physics infrared laser, femtosecond tunable with excitation between 700-1000 nm. Pulse selector for adapting the pulse frequency to the sensor operating frequency.
  • Picosecond pulsed diode laser, 403 nm (for DAPI one-photon excitation).
  • Picosecond pulsed diode laser, 445 nm (for CFP one-photon excitation).

 

One-photon FLIM

  • Picosecond pulsed diode laser 442nm.
  • Picosecond pulsed diode laser 464nm.
Alain Jauneau / Cécile Pouzet

+33 5 34 32 38 37

imageriefraib@lrsv.ups-tlse.fr

FRAIB – Campus INRA Auzeville

Software manual

 

Some publications made thanks to these resources:

  • Dorian Farache, Alain Jauneau, Cécile Chemin, Marine Chartrain, Marie-Hélène Rémy, Andreas Merdes and Laurence Haren. Functional analysis of Gamma-tubulin Complex Proteins indicates specific lateral association via their N-terminal domains. JBC, 2016
  • A. Testard, D. Da Silva, M Ormancey, C. Pichereaux, C. Pouzet, A. Jauneau, S. Grat, E. Robe, Brière, V. Cotelle, C. Mazars and P. Thulleau. Calcium- and Nitric Oxide-Dependent Nuclear Accumulation of Cytosolic Glyceraldehyde-3-Phosphate Dehydrogenase in Response to Long Chain Bases in Tobacco BY-2 Cell. Plant Cell Physiol, 2016
  • Marçal Soler, Anna Plasencia, Romain Larbat, Cécile Pouzet, Alain Jauneau, Susana Rivas, Edouard Pesquet, Catherine Lapierre, Isabelle Truchet and Jacqueline Grima-Pettenati. The Eucalyptus linker histone variant EgH1.3 cooperates with the transcription factor EgMYB1 to control lignin biosynthesis during wood formation. New Phytologist, 2016
  • Judith Fliegmann, Alain Jauneau, Carole Pichereaux, Charles Rosenberg, Virginie Gasciolli, Antonius C.J. Timmers, Odile Burlet-Schiltz, Julie Cullimore and Jean-Jacques Bono. LYR3, a high-affinity LCO-binding protein of Medicago truncatula, interacts with LYK3, a key symbiotic receptor. Febs Letters, 2016
  • Clementine Le Roux,Gälle Huet, Alain Jauneau, Laurent Camborde, Dominique Tremousaygue, Alexandra Kraut, Binbin Zhou, Marie Levaillant, Hiroaki Adachi, Hirofumi Yoshioka, Sylvain Raffaele, Richard Berthome, Yohann Coute, Jane E. Parker,and Laurent Deslandes. A Receptor Pair with an Integrated Decoy Converts Pathogen Disabling of Transcription Factors to Immunity. Cell, 2015
  • Josep Vilarrasa-Blasi, Mary-Paz Gonzalez-Garcıa, David Frigola, Norma Fabregas-Vallve, Konstantinos G. Alexiou, Nuria Lopez-Bigas, Susana Rivas, Alain Jauneau, Jan U. Lohmann, Philip N. Benfey, Marta Ibanes, and Ana I. Cano-Delgado Regulation of Plant Stem Cell Quiescence by a Brassinosteroid Signaling Module. Developmental Cell, 2014
  • Stella Cesari Gaëtan Thilliez, Cécile Ribot, Véronique Chalvon, Corinne Michel, Alain Jauneau, Susana Rivas, Ludovic Alaux, Hiroyuki Kanzaki, Yudai Okuyama, Jean-Benoit Morel, Elisabeth Fournier, Didier Tharreau, Ryohei Terauchi and Thomas Kroj. The Rice Resistance Protein Pair RGA4/RGA5 Recognizes the Magnaporthe oryzae Effectors AVR-Pia and AVR1-CO39 by Direct Binding. The Plant Cell, 2013, 25:1463-1481
  • Daniel Marino, Solène Froidure, Joanne Canonne, Sara Ben Khaled, Mehdi Khafif, Cécile Pouzet, Alain Jauneau, Dominique Roby, Susana Rivas. Arabidopsis ubiquitin ligase MIEL1 mediates degradation of the transcription factor MYB30 weakening plant defence. Nature Communications, 2013, 4: 1476 .
  • Céline Courilleau, Catherine Chailleux, Alain Jauneau, Fanny Grimal, Sébastien Briois, Elisa Boutet-Robinet, François Boudsocq, Didier Trouche, and Yvan Canitrot. The chromatin remodeler p400 ATPase facilitates Rad51-mediated repair of DNA double-strand breaks J Cell Biol, 2012 199:1067-1081.
  • Joanne Canonne, Daniel Marino, Alain Jauneau, Cecile Pouzet, Christian Brière, Dominique Roby and Susana Rivas. The Xanthomonas Type III Effector XopD Targets the Arabidopsis Transcription Factor MYB30 to Suppress Plant Defense. Plant Cell, 2011.
  • Katharina Heidrich, Lennart Wirthmueller, Céline Tasset, Cécile Pouzet, Laurent Deslandes and Jane E. Parker. Arabidopsis EDS1 connects pathogen effector recognition to cell compartment specific immune responses. Science 2011.
  • Rose Boutros, Corinne Lorenzo, Odile Mondesert, Alain Jauneau, Vanessa Oakes, Christine Dozier, Brian Gabrielli, Bernard Ducommun. CDC25B associates with a centrin 2-containing complex and is involved in maintaining centrosome integrity.Biol Cell., 2011.
  • Solène Froidure, Xavier Daniel, Joanne Canonne, Alain Jauneau, Christian Brière, Dominique Roby, Susana Rivas. AtsPLA2-α negatively regulates hypersensitive cell death and plant defence through its interaction with the Arabidopsis transcription factor AtMYB30. PNAS, 2010, 107 (34)15281-15286
  • Céline Tasset, Maud Bernoux, Alain Jauneau, Cécile Pouzet, Christian Brière, Sylvie Kieffer-Jacquino, Yves Marco and Laurent Deslandes. The Ralstonia solanacearum effector PopP2 autoacetylates on a lysine residue that is required to trigger plant immune response. PlosPathogens. 2010.
  • Alexandre Perochon, Stefan Dieterle, Cecile Pouzet, Didier Aldon, Jean-Philippe Galaud, Benoit, Ranty. Interaction of a plant pseudo-response regulator with a calmodulin-like protein. BBRC 2010.
  • Bernoux M, Timmers T, Jauneau A, Brière C, de Wit PJ, Marco Y, Deslandes L. RD19, an Arabidopsis Cysteine Protease Required for RRS1-R-Mediated Resistance, Is Relocalized to the Nucleus by the Ralstonia solanacearum PopP2 Effector. Plant Cell., 2008. 20: 2252-2264.

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