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7. A microfluidic platform towards automated multiplexed in situ sequencing

Author

Maïno, Nicolas, Hauling, Thomas, Cappi, G., Madaboosi, Narayanan, Dupouy, D. G., Nilsson, Mats, Maïno, Nicolas, Hauling, Thomas, Cappi, G., Madaboosi, Narayanan, Dupouy, D. G., and Nilsson, Mats

Abstract

Advancements in multiplexed in situ RNA profiling techniques have given unprecedented insight into spatial organization of tissues by enabling single-molecule quantification and sub-micron localization of dozens to thousands of RNA species simultaneously in cells and entire tissue sections. However, the lack of automation of the associated complex experimental procedures represents a potential hurdle towards their routine use in laboratories. Here, we demonstrate an approach towards automated generation and sequencing of barcoded mRNA amplicons in situ, directly in fixed cells. This is achieved through adaptation of a microfluidic tool compatible with standard microscope slides and cover glasses. The adapted tool combines a programmable reagent delivery system with temperature controller and flow cell to perform established in situ sequencing protocols, comprising hybridization and ligation of gene-specific padlock probes, rolling circle amplification of the probes yielding barcoded amplicons and identification of amplicons through barcode sequencing. By adapting assay parameters (e.g. enzyme concentration and temperature), we achieve a near-identical performance in identifying mouse beta-actin transcripts, in comparison with the conventional manual protocol. The technically adapted assay features i) higher detection efficiency, ii) shorter protocol time, iii) lower consumption of oligonucleotide reagents but slightly more enzyme. Such an automated microfluidic tissue processor for in situ sequencing studies would greatly enhance its research potentials especially for cancer diagnostics, thus paving way to rapid and effective therapies.

Published
2019
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10. Projet RESPIRATION : Création de ressources pédagogiques numériques accessibles pour une Université inclusive : Collection d’interviews interdisciplinaires signéeset filmées d’experts du colloque international de recherche « Agricultures Urbaines & Transition Ecologique, Toulouse 2017 »

Author

Dumat, Camille, Combes, S., Michaud, N, Laffont, L., Saucian, M, Leroy, A, Plessis-Fraissard, N, Ardourel, Y, Aubry, C, Champoux, F, Toutut-Picard, E, Consales, Jn, Dupouy, D, Lepengue, N, Centre d'Etude et de Recherche Travail Organisation Pouvoir (CERTOP), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Université Toulouse - Jean Jaurès (UT2J), Chimie, biologie et radicaux libres - UMR 6517 (CBRL), Université de la Méditerranée - Aix-Marseille 2-Université Paul Cézanne - Aix-Marseille 3-Université de Provence - Aix-Marseille 1-Centre National de la Recherche Scientifique (CNRS), Centre interuniversitaire de recherche et d'ingenierie des matériaux (CIRIMAT), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie du CNRS (INC), Université Toulouse - Jean Jaurès (UT2J), Université de Toulouse (UT)-Université de Toulouse (UT)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), and Université de Toulouse (UT)

Subjects
[SDE]Environmental Sciences, ComputingMilieux_MISCELLANEOUS
Abstract

International audience

Published
2017

12. Voltage polarity effects in GST-based phase change memory: Physical origins and implications

Author

Padilla, A., primary, Burr, G. W., additional, Virwani, K., additional, Debunne, A., additional, Rettner, C. T., additional, Topuria, T., additional, Rice, P. M., additional, Jackson, B., additional, Dupouy, D., additional, Kellock, A. J., additional, Shelby, R. M., additional, Gopalakrishnan, K., additional, Shenoy, R. S., additional, and Kurdi, B. N., additional

Published
2010
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18. Early neonatal diagnosis of congenital toxoplasmosis: value of comparative enzyme-linked immunofiltration assay immunological profiles and anti-Toxoplasma gondii immunoglobulin M (IgM) or IgA immunocapture and implications for postnatal therapeutic strategies

Author

Pinon, J M, primary, Chemla, C, additional, Villena, I, additional, Foudrinier, F, additional, Aubert, D, additional, Puygauthier-Toubas, D, additional, Leroux, B, additional, Dupouy, D, additional, Quereux, C, additional, Talmud, M, additional, Trenque, T, additional, Potron, G, additional, Pluot, M, additional, Remy, G, additional, and Bonhomme, A, additional

Published
1996
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30. Pyrimethamine-sulfadoxine Treatment of Congenital Toxoplasmosis: Follow-up of 78 Cases Between 1980 and 1997.

Author

Villena, I., Aubert, D., Leroux, B., Dupouy, D., Talmud, M., Chemla, C., Trenque, T., Schmit, G., Quereux, C., Guenounou, M., Pluot, M., Bonhomme, A., and Pinon, J. M.

Subjects
CONGENITAL toxoplasmosis, PEDIATRIC drug therapy, DRUG therapy
Abstract

The purpose of this study was to determine the clinical and immunological outcome of 78 children with congenital toxoplasmosis treated with the pyrimethamine-sulfadoxine combination between 1980 and 1997. Methods. Children were divided into 3 groups according to the initial duration of treatment (always including folinic acid, 5 mg/week by mouth), as follows: pyrimethamine (1.25 mg/kg every 15 d)+sulfadoxine (25 mg/kg every 15 d) for 12 months (Group 1, 47 children), or for 24 months, with or without prenatal therapy (respectively, Group 2, 19 children, and Group 3, 12 children). Results. Chorioretinitis occurred in 23% of these 78 children. Four children had unilateral blindness, 1 had mild epileptic fits and 1 had psychomotor retardation. The lowest rate of sequelae were in Groups 2 and 3. Immunological rebounds, generally without clinical repercussions, occurred frequently (90% of cases on average) during, or more often after therapy, regardless of the treatment duration. Treatment was always well tolerated. Conclusions. Our current treatment strategy for congenital toxoplasmosis consists of a 24-month course of pyrimethamine-sulfadoxine (Fansidar[sup ®]) combined with folinic acid (Lederfoline[sup ®]). If the prenatal diagnosis is positive, we also prescribe this treatment to the mother until delivery. This combination offers satisfactory compliance, adequate serum concentrations, and good preventive efficacy. [ABSTRACT FROM AUTHOR]

Published
1998
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36. Demonstration of a Biological Effect of Natural Ionizing Radiations

Author

Conter, A., Dupouy, D., and Planel, H.

Abstract

The blue-green alga, Synechococcus lividus, was grown under various levels of radiations. Shielding of cultures with lead resulted in a lower cell growth rate: the reduction disappeared when a normal radiation level was restored in the lead chamber. Irradiations from a thorium source at a dose-rate 14 times higher than that of natural irradiation stimulated the growth of the algae. These results are a new demonstration of a biological effect of background radiations.

Published
1983
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37. Pharmacodynamics and pharmacokinetics of dermatan sulfate in humans

Author

Dol, F, Houin, G, Rostin, M, Montastruc, JL, Dupouy, D, Gianese, F, Sie, P, and Boneu, B

Abstract

Dermatan sulfate (DS), a catalyst of the thrombin-heparin cofactor II interaction, has antithrombotic activity and is devoid of significant hemorrhagic risk in several animal models. We investigated the pharmacodynamic and pharmacokinetic properties of DS in humans. DS was injected in single bolus intravenous injections of four increasing doses (0.5, 1, 1.5, 2 mg/kg) to six healthy volunteers. The resulting anticoagulant activities were assessed by the activated partial thromboplastin time (APTT) and the thrombin clotting time (TCT). There were dose-dependent prolongations of the APTT and TCT, and the anticoagulant activities disappeared in less than three hours. The pharmacokinetic parameters were calculated from the plasma concentrations of DS measured with a new chromogenic assay. The volume of distribution was approximately 1.8 times greater than the theoretical plasma volume and was independent of dose. In contrast, the clearance decreased with dose and the terminal half-life ranged from 0.45 +/- 0.08 hours at 0.5 mg/kg to 0.72 +/- 0.11 hours (mean +/- SD) at 2 mg/kg. The bioavailabilities of subcutaneous (SC) and intramuscular (IM) administration relative to those of intravenous administration were determined in 12 other volunteers. The respective bioavailabilities were 24.7% +/- 12.9% and 12.4% +/- 9.2% for SC and IM administration. There was no detectable change in the APTT and the TCT when the volunteers were injected with 1.5 mg/kg SC or IM. In addition, the pharmacokinetic parameters derived from plasma concentrations of DS showed considerable interindividual variations by the two later routes of administration. Peak concentrations were noted 2.7 +/- 1.3 hours after SC injection and 4.3 +/- 4.9 hours after IM injection. The average peak concentrations were 0.7 +/- 0.3 and 0.4 +/- 0.2 mg/L after SC and IM injections, respectively. The half-lives of DS were 7.9 +/- 6.5 hours (SC) and 6.3 +/- 7.4 hours (IM). No adverse reaction to DS was recorded during this study.

Published
1989
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