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Biomicrofluidics / Volume 6 / Issue 1 / REGULAR ARTICLES

Biomicrofluidics 6, 014109 (2012); http://dx.doi.org/10.1063/1.3683161 (9 pages)

High-throughput study of alpha-synuclein expression in yeast using microfluidics for control of local cellular microenvironment

Patrícia Rosa1, Sandra Tenreiro2, Virginia Chu1, Tiago F. Outeiro2,3, and João Pedro Conde1,4

1INESC Microsistemas e Nanotecnologias (INESC MN) and IN-Institute of Nanoscience and Nanotechnology, Rua Alves Redol, 9, Lisbon 1000-029, Portugal
2Instituto de Medicina Molecular, Faculdade de Medicina da Universidade de Lisboa Av. Prof. Egas Moniz, 1649-028 Lisboa, Portugal
3Department of Neurodegeneration and Restaurative Research, University Medizin Goettingen, Goettingen, Germany
4Department of Bioengineering, Instituto Superior Técnico, Av Rovisco Pais 1, Lisbon 1049-001, Portugal

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(Received 24 October 2011; accepted 12 January 2012; published online 9 February 2012)

Microfluidics is an emerging technology which allows the miniaturization, integration, and automation of fluid handling processes. Microfluidic systems offer low sample consumption, significantly reduced processing time, and the prospect of massive parallelization. A microfluidic platform was developed for the control of the soluble cellular microenvironment of Saccharomyces cerevisiae cells, which enabled high-throughput monitoring of the controlled expression of alpha-synuclein (aSyn), a protein involved in Parkinson’s disease. Y-shaped structures were fabricated using particle desorption mass spectrometry-based soft-lithography techniques to generate biomolecular gradients along a microchannel. Cell traps integrated along the microchannel allowed the positioning and monitoring of cells in precise locations, where different, well-controlled chemical environments were established. S. cerevisiae cells genetically engineered to encode the fusion protein aSyn-GFP (green fluorescent protein) under the control of GAL1, a galactose inducible promoter, were loaded in the microfluidic structure. A galactose concentration gradient was established in the channel and a time-dependent aSyn-GFP expression was obtained as a function of the positioning of cells along the galactose gradient. Our results demonstrate the applicability of this microfluidic platform to the spatiotemporal control of cellular microenvironment and open a range of possibilities for the study of cellular processes based on single-cell analysis.

© 2012 American Institute of Physics

Article Outline

  1. INTRODUCTION
  2. MATERIALS AND METHODS
    1. Microfluidic structure fabrication
    2. Generation of stable gradients and further validation by fluorescence microscopy
    3. Cell media and growth conditions
    4. Monitoring of Syn-GFP expression induction as a function of the local concentration of galactose
  3. RESULTS AND DISCUSSION
    1. Generation of a stable GAL gradient inside of the microchannel
    2. ASyn-GFP expression induction as a function of the generation of GAL and RAF gradients within microchannels
  4. CONCLUSIONS

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KEYWORDS, PACS, and IPC

Keywords

bioMEMS, cellular biophysics, diseases, genetic engineering, medical control systems, microchannel flow, microorganisms, molecular biophysics, patient monitoring, proteins, soft lithography, spatiotemporal phenomena

PACS

International Patent Classification (IPC)

  • B81B

    Micro-structural devices or systems, e.g. micro-mechanical devices

  • F15D

    Fluid dynamics, i.e. methods or means for influencing the flow of gases or liquids

  • H01L21/02

    Manufacture or treatment of semiconductor devices or of parts thereof

  • H01L21/70

    Manufacture or treatment of devices consisting of a plurality of solid state components or integrated circuits formed in or on a common substrate or of specific parts thereof; Manufacture of integrated circuit devices or of specific parts thereof

  • G03F7/00

    Photomechanical, e.g. photolithographic, production of textured or patterned surfaces, e.g. printed surfaces; Materials therefor, e.g. comprising photoresists; Apparatus specially adapted therefor

  • C12N

    Micro-organisms or enzymes; Compositions thereof; Propagating, preserving, or maintaining micro-organisms; Mutation or genetic engineering; Culture media

ARTICLE DATA

Digital Object Identifier
http://dx.doi.org/10.1063/1.3683161

PUBLICATION DATA

ISSN

1932-1058 (online)

Publisher

$abstract.society.value is a Member of CrossRef American Institute of Physics

For access to fully linked references, you need to log in.
    A. Seidi, H. Kaji, N. Annabi, S. Ostrovidov, M. Ramalingam, and A. Khademhosseini, Biomicrofluidics 5, 22214 (2011)BIOMGB000005000002022214000001.


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