WO2018085557A1 - Formation de pores cellulaire à l'aide d'un rayonnement laser pulsé - Google Patents

Formation de pores cellulaire à l'aide d'un rayonnement laser pulsé Download PDF

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Publication number
WO2018085557A1
WO2018085557A1 PCT/US2017/059743 US2017059743W WO2018085557A1 WO 2018085557 A1 WO2018085557 A1 WO 2018085557A1 US 2017059743 W US2017059743 W US 2017059743W WO 2018085557 A1 WO2018085557 A1 WO 2018085557A1
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WIPO (PCT)
Prior art keywords
cells
substrate
projections
range
cell
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PCT/US2017/059743
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English (en)
Inventor
Eric Mazur
Nabiha SAKLAYEN
Marinna MADRID
Marinus HUBER
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President And Fellows Of Harvard College
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Application filed by President And Fellows Of Harvard College filed Critical President And Fellows Of Harvard College
Publication of WO2018085557A1 publication Critical patent/WO2018085557A1/fr
Priority to US16/250,714 priority Critical patent/US10829729B2/en

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    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
    • C12M35/00—Means for application of stress for stimulating the growth of microorganisms or the generation of fermentation or metabolic products; Means for electroporation or cell fusion
    • C12M35/02—Electrical or electromagnetic means, e.g. for electroporation or for cell fusion

Definitions

  • FIG. 6A is a schematic representation of an experimental set-up for irradiating a substrate with one or more laser pulses to deliver cargos to cells in accordance with aspects of the present disclosure
  • FIG. 12D is a fluorescent image of cells depicting cell viability after delivery of calcein red-orange AM
  • FIG. 13A is a scanning electron microscopy image of a top view of a
  • FIG. 13D is a Scanning Electron Microscopy image of a top view of a
  • FIG. 19 is a table depicting flow cytometry settings on BD LSRFortessaSORPTM cell analyser running BD FACSDiva software version 6.1.3;
  • the present disclosure relates generally to substrates for use in processing cells and method of cells processing that allow causing changes in cells that facilitate uptake of cargos by the cells.
  • a medium containing one or more cells and one or more cargos to be internalized by the cells is disposed on a substrate surface having a plurality of metalized projections.
  • the application of a plurality of nanosecond laser pulses to the cells can mediate the uptake of the cargo(s) by the cells. It has been discovered that the use of nanosecond pulses in combination with a substrate having metalized projections as discussed herein leads to efficient introduction of many types of cargos into cells at a high cell viability.
  • FIGs. 8B and 8C show, respectively, a scanning electron microscopy (SEM) image of a top view, and an SEM image of a side view, of thermoplasmonic substrates formed using the above process. Characterization through SEM confirms that the pyramids of the template-stripped thermoplasmonic substrates are highly uniform in base lengths (2.4 ⁇ ), spacings (1.4 ⁇ ), and heights (1.4 ⁇ ), and consistent from batch to batch.
  • SEM scanning electron microscopy

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  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Engineering & Computer Science (AREA)
  • Wood Science & Technology (AREA)
  • Organic Chemistry (AREA)
  • Biotechnology (AREA)
  • Chemical & Material Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Zoology (AREA)
  • Microbiology (AREA)
  • Sustainable Development (AREA)
  • Physics & Mathematics (AREA)
  • Biomedical Technology (AREA)
  • Cell Biology (AREA)
  • Biochemistry (AREA)
  • General Engineering & Computer Science (AREA)
  • General Health & Medical Sciences (AREA)
  • Electromagnetism (AREA)
  • Apparatus Associated With Microorganisms And Enzymes (AREA)

Abstract

Selon un aspect, l'invention concerne un procédé de traitement de cellules, qui consiste à disposer une pluralité de cellules sur un substrat à travers lequel une pluralité de saillies sont distribuées et une couche électroconductrice recouvrant au moins partiellement lesdites saillies, à exposer les cellules à une cargaison devant être internalisée par les cellules, à exposer la surface du substrat (et en particulier les saillies) à une ou plusieurs impulsions laser ayant une largeur d'impulsion dans une plage d'environ 1 ns à environ 1 000 ns de manière à faciliter l'absorption de la cargaison par au moins une partie des cellules (par exemple, les cellules positionnées à proximité des saillies (par exemple, à quelques centaines de nanomètres (comme moins de 100 nm) des saillies)). Selon certains modes de réalisation, les impulsions laser ont une largeur d'impulsion dans une plage d'environ 10 ns à environ 500 ns, par exemple, dans une plage d'environ 5 ns à environ 50 ns.
PCT/US2017/059743 2016-11-03 2017-11-02 Formation de pores cellulaire à l'aide d'un rayonnement laser pulsé WO2018085557A1 (fr)

Priority Applications (1)

Application Number Priority Date Filing Date Title
US16/250,714 US10829729B2 (en) 2016-11-03 2019-01-17 Cellular poration using laser radiation

Applications Claiming Priority (4)

Application Number Priority Date Filing Date Title
US201662416789P 2016-11-03 2016-11-03
US62/416,789 2016-11-03
US201762442293P 2017-01-04 2017-01-04
US62/442,293 2017-01-04

Related Parent Applications (1)

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PCT/US2017/059720 Continuation-In-Part WO2018085542A1 (fr) 2016-11-03 2017-11-02 Perméabilisation cellulaire par rayonnement laser continu

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US16/250,714 Continuation-In-Part US10829729B2 (en) 2016-11-03 2019-01-17 Cellular poration using laser radiation

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Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP3833959A4 (fr) * 2018-08-10 2022-06-29 Cellino Biotech, Inc. Système de fabrication de cellules commandée par l'image
US11643667B2 (en) 2017-08-28 2023-05-09 Cellino Biotech, Inc. Microfluidic laser-activated intracellular delivery systems and methods

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009017695A1 (fr) * 2007-07-26 2009-02-05 The Regents Of The University Of California Outil chirurgical pour cellule unique et dispositif de transfection cellulaire utilisant les propriétés photothermiques de films minces et/ou de nanoparticules métalliques
EP2272945A1 (fr) * 2009-07-06 2011-01-12 Gottfried Wilhelm Leibniz Universität Hannover Procédé et système pour la manipulation de cellules
WO2016127069A1 (fr) * 2015-02-06 2016-08-11 President And Fellows Of Harvard College Procédé et système plasmoniques de thérapie cellulaire, basés sur des nanocavités

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2009017695A1 (fr) * 2007-07-26 2009-02-05 The Regents Of The University Of California Outil chirurgical pour cellule unique et dispositif de transfection cellulaire utilisant les propriétés photothermiques de films minces et/ou de nanoparticules métalliques
EP2272945A1 (fr) * 2009-07-06 2011-01-12 Gottfried Wilhelm Leibniz Universität Hannover Procédé et système pour la manipulation de cellules
WO2016127069A1 (fr) * 2015-02-06 2016-08-11 President And Fellows Of Harvard College Procédé et système plasmoniques de thérapie cellulaire, basés sur des nanocavités

Non-Patent Citations (8)

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Title
DEMESY ET AL.: "Tridimensional Multiphysics Model for the Study of Photo-induced Thermal Effects in Arbitrary Nano-structures", JEOS:RP, vol. 6, 2011, pages 11037
EKICI ET AL.: "Thermal Analysis of Gold Nanorods Heated with Femtosecond Laser Pulses", J. PHYS. D APPL. PHYS., vol. 41, no. 18, 2008, pages 185501, XP020140817, DOI: doi:10.1088/0022-3727/41/18/185501
ERIC DIEBOLD: "Plasmon-enhanced nonlinear optics for applications in sensing and biology", PH. D. THESIS, May 2010 (2010-05-01), XP055248702, Retrieved from the Internet <URL:http://mazur.harvard.edu/publications/Pub_679.pdf> [retrieved on 20160209] *
NABIHA SAKLAYEN ET AL: "Intracellular Delivery Using Nanosecond-Laser Excitation of Large-Area Plasmonic Substrates", ACS NANO, vol. 11, no. 4, 25 April 2017 (2017-04-25), US, pages 3671 - 3680, XP055460666, ISSN: 1936-0851, DOI: 10.1021/acsnano.6b08162 *
RANHUA XIONG ET AL: "Laser-assisted photoporation: fundamentals, technological advances and applications", ADVANCES IN PHYSICS: X, vol. 1, no. 4, 3 July 2016 (2016-07-03), pages 596 - 620, XP055466643, ISSN: 2374-6149, DOI: 10.1080/23746149.2016.1228476 *
S?BASTIEN COURVOISIER ET AL: "Plasmonic Tipless Pyramid Arrays for Cell Poration", NANO LETTERS, vol. 15, no. 7, 8 July 2015 (2015-07-08), pages 4461 - 4466, XP055248701, ISSN: 1530-6984, DOI: 10.1021/acs.nanolett.5b01697 *
YI-CHIEN WU ET AL: "Massively parallel delivery of large cargo into mammalian cells with light pulses", NATURE METHODS, vol. 12, no. 5, May 2015 (2015-05-01), pages 439 - 444, XP055248844, ISSN: 1548-7091, DOI: 10.1038/nmeth.3357 *
YI-CHIEN WU ET AL: "Supplementary information.-Massively parallel delivery of large cargo into mammalian cells with light pulses", NATURE METHODS, vol. 12, no. 5, 6 April 2015 (2015-04-06), pages 439 - 444, XP055276556, ISSN: 1548-7091, DOI: 10.1038/nmeth.3357 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US11643667B2 (en) 2017-08-28 2023-05-09 Cellino Biotech, Inc. Microfluidic laser-activated intracellular delivery systems and methods
EP3833959A4 (fr) * 2018-08-10 2022-06-29 Cellino Biotech, Inc. Système de fabrication de cellules commandée par l'image

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