Micropatterning of substrates for the culture of cell networks by stencil-assisted additive nanofabrication

Anita Previdi; Claudio Piazzoni; Francesca Borghi; Carsten Schulte; Leandro Lorenzelli; Flavio Giacomozzi; Alessio Bucciarelli; Antonio Malgaroli; Jacopo Lamanna; Andrea Moro; Gabriella Racchetti; Alessandro Podestà; Cristina Lenardi; Paolo Milani

doi:10.3390/mi12010094

Micropatterning of substrates for the culture of cell networks by stencil-assisted additive nanofabrication

Anita Previdi
, Claudio Piazzoni
, Francesca Borghi
, Carsten Schulte
, Leandro Lorenzelli
, Flavio Giacomozzi
, Alessio Bucciarelli
, Antonio Malgaroli
, Jacopo Lamanna
, Andrea Moro
, Gabriella Racchetti
, Alessandro Podestà
, Cristina Lenardi
, Paolo Milani^*

^*Corresponding author for this work

Biomedical Engineering

Research output: Contribution to journal › Article › peer-review

4 Citations (Scopus)

7 Downloads (Pure)

Abstract

The fabrication of in vitro neuronal cell networks where cells are chemically or electrically connected to form functional circuits with useful properties is of great interest. Standard cell culture substrates provide ensembles of cells that scarcely reproduce physiological structures since their spatial organization and connectivity cannot be controlled. Supersonic Cluster Beam Deposition (SCBD) has been used as an effective additive method for the large-scale fabrication of interfaces with extracellular matrix-mimicking surface nanotopography and reproducible morphological properties for cell culture. Due to the high collimation of SCBD, it is possible to exploit stencil masks for the fabrication of patterned films and reproduce features as small as tens of micrometers. Here, we present a protocol to fabricate micropatterned cell culture substrates based on the deposition of nanostructured cluster-assembled zirconia films by stencil-assisted SCBD. The effectiveness of this approach is demonstrated by the fabrication of micrometric patterns able to confine primary astrocytes. Calcium waves propagating in the astrocyte networks are shown.

Original language	English
Article number	94
Number of pages	18
Journal	Micromachines
Volume	12
Issue number	1
Early online date	18 Jan 2021
DOIs	https://doi.org/10.3390/mi12010094
Publication status	Published - 18 Jan 2021

Funding

F.B. thanks Centro Universitario Cattolico (CUC) for partially supporting this work and Roberto Presilla and Alessandro Tredicucci for stimulating discussions. C.P. thanks Daniele Viganò and Federico Pezzotta for their technical assistance in sample holders’ machining. C.S. thanks Francesco Mario Esposito for his contribution in the development of the protocol for the antifouling functionalization of the substrates. C.S. thanks the support of the UNITECH NOLIMITS Imaging facility at the University of Milan for the confocal microscopy imaging.

Keywords

astrocytes
cell confinement
micropatterns
nanofabrication
nanostructured zirconia
primary cell networks

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10.3390/mi12010094Licence: CC BY 4.0

Previdi-etal-MM-2021-Micropatterning-of-substrates-for-the-culture-of-cell-networks-by-stencil-assistedFinal published version, 2.57 MBLicence: CC BY 4.0

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Previdi, A., Piazzoni, C., Borghi, F., Schulte, C., Lorenzelli, L., Giacomozzi, F., Bucciarelli, A., Malgaroli, A., Lamanna, J., Moro, A., Racchetti, G., Podestà, A., Lenardi, C., & Milani, P. (2021). Micropatterning of substrates for the culture of cell networks by stencil-assisted additive nanofabrication. Micromachines, 12(1), Article 94. https://doi.org/10.3390/mi12010094

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abstract = "The fabrication of in vitro neuronal cell networks where cells are chemically or electrically connected to form functional circuits with useful properties is of great interest. Standard cell culture substrates provide ensembles of cells that scarcely reproduce physiological structures since their spatial organization and connectivity cannot be controlled. Supersonic Cluster Beam Deposition (SCBD) has been used as an effective additive method for the large-scale fabrication of interfaces with extracellular matrix-mimicking surface nanotopography and reproducible morphological properties for cell culture. Due to the high collimation of SCBD, it is possible to exploit stencil masks for the fabrication of patterned films and reproduce features as small as tens of micrometers. Here, we present a protocol to fabricate micropatterned cell culture substrates based on the deposition of nanostructured cluster-assembled zirconia films by stencil-assisted SCBD. The effectiveness of this approach is demonstrated by the fabrication of micrometric patterns able to confine primary astrocytes. Calcium waves propagating in the astrocyte networks are shown.",

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AU - Lorenzelli, Leandro

AU - Giacomozzi, Flavio

AU - Bucciarelli, Alessio

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AU - Lamanna, Jacopo

AU - Moro, Andrea

AU - Racchetti, Gabriella

AU - Podestà, Alessandro

AU - Lenardi, Cristina

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N2 - The fabrication of in vitro neuronal cell networks where cells are chemically or electrically connected to form functional circuits with useful properties is of great interest. Standard cell culture substrates provide ensembles of cells that scarcely reproduce physiological structures since their spatial organization and connectivity cannot be controlled. Supersonic Cluster Beam Deposition (SCBD) has been used as an effective additive method for the large-scale fabrication of interfaces with extracellular matrix-mimicking surface nanotopography and reproducible morphological properties for cell culture. Due to the high collimation of SCBD, it is possible to exploit stencil masks for the fabrication of patterned films and reproduce features as small as tens of micrometers. Here, we present a protocol to fabricate micropatterned cell culture substrates based on the deposition of nanostructured cluster-assembled zirconia films by stencil-assisted SCBD. The effectiveness of this approach is demonstrated by the fabrication of micrometric patterns able to confine primary astrocytes. Calcium waves propagating in the astrocyte networks are shown.

AB - The fabrication of in vitro neuronal cell networks where cells are chemically or electrically connected to form functional circuits with useful properties is of great interest. Standard cell culture substrates provide ensembles of cells that scarcely reproduce physiological structures since their spatial organization and connectivity cannot be controlled. Supersonic Cluster Beam Deposition (SCBD) has been used as an effective additive method for the large-scale fabrication of interfaces with extracellular matrix-mimicking surface nanotopography and reproducible morphological properties for cell culture. Due to the high collimation of SCBD, it is possible to exploit stencil masks for the fabrication of patterned films and reproduce features as small as tens of micrometers. Here, we present a protocol to fabricate micropatterned cell culture substrates based on the deposition of nanostructured cluster-assembled zirconia films by stencil-assisted SCBD. The effectiveness of this approach is demonstrated by the fabrication of micrometric patterns able to confine primary astrocytes. Calcium waves propagating in the astrocyte networks are shown.

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KW - micropatterns

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ER -

Micropatterning of substrates for the culture of cell networks by stencil-assisted additive nanofabrication

Abstract

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Keywords

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