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Replica-moulded polydimethylsiloxane culture vessel lids attenuate osmotic drift in long-term cell cultures

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Abstract

An imbalance in medium osmolarity is a determinant that affects cell culture longevity. Even in humidified incubators, evaporation of water leads to a gradual increase in osmolarity over time. We present a simple replica-moulding strategy for producing self-sealing lids adaptable to standard, small-size cell-culture vessels. They are made of polydimethylsiloxane (PDMS), a flexible, transparent and biocompatible material, which is gas-permeable but largely impermeable to water. Keeping cell cultures in a humidified 5% CO2 incubator at 37°C, medium osmolarity increased by +6.86 mosmol/kg/day in standard 35 mm Petri dishes, while PDMS lids attenuated its rise by a factor of four to changes of +1.72 mosmol/kg/day. Depending on the lid membrane thickness, pH drifts at ambient CO2 levels were attenuated by a factor of 4 to 9. Comparative evaporation studies at temperatures below 60°C yielded a 10-fold reduced water vapour flux of 1.75 g/day/dm2 through PDMS lids as compared with 18.69 g/day/dm2 with conventional Petri dishes. Using such PDMS lids, about 2/3 of the cell cultures grew longer than 30 days in vitro. Among these, the average survival time was 69 days with the longest survival being 284 days under otherwise conventional cell culture conditions.

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Abbreviations

DIV:

days in vitro

FEP:

fluorinated ethylene propylene

HBSS:

Hanks balanced salt solution

ITO:

indium tin oxide

MEA:

microelectrode array

PDMS:

polydimethylsiloxane

PMMA:

polymethyl methacrylate

PS:

polystyrene

PTFE:

polytetrafluorethylene

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Authors and Affiliations

  1. Department of Neuroscience and Brain Technologies, The Italian Institute of Technology, Via Morego 30, 16163, Genoa, Italy

    Axel Blau & Fabio Benfenati

  2. Department of Physics and Biophysics, University of Kaiserslautern, Erwin-Schroedinger-Str. 56, 67663, Kaiserslautern, Germany

    Tanja Neumann & Christiane Ziegler

Authors
  1. Axel Blau
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  2. Tanja Neumann
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  3. Christiane Ziegler
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  4. Fabio Benfenati
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Correspondence to Axel Blau.

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Supplementary material pertaining to this article is available on the Journal of Biosciences Website at http://www.ias.ac.in/jbiosci/mar2009/pp59-69-suppl.pdf

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Blau, A., Neumann, T., Ziegler, C. et al. Replica-moulded polydimethylsiloxane culture vessel lids attenuate osmotic drift in long-term cell cultures. J Biosci 34, 59–69 (2009). https://doi.org/10.1007/s12038-009-0009-3

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