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Co-culture Synaptogenic Assay: A New Look at Fluorescence Reporters and Technological Devices

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Synapse Development

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1538))

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Abstract

The mechanism underlying the differentiation of pre- and postsynaptic specifications involves the sequential and dynamic recruitment of specific molecules coordinated by bidirectional signaling across the synaptic cleft. In this chapter, we describe the co-culture assay, a useful method to evaluate cell-surface molecules through its ability to promote the recruitment of proteins required for synapse structure and function. The versatility of this simple and reliable method is illustrated by the wide variety of applications ranging from analysis of synaptogenic activity to evaluation of soluble compounds with therapeutic potential. In addition, we provide a framework to enable the co-culture assay as a tool for high-throughput studies, thereby improving the efficiency and sensitivity of this classic method in neuroscience.

Authors contributed equally, correspondence may be addressed to either.

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Abbreviations

HTPS:

High-throughput studies

CAMs:

Cell adhesion molecules

NLG1:

Neuroligin 1

EphBs:

Ephrin B

LRRTM1:

Leucine rich repeat transmembrane neuronal 1

SynCAMs/Necls:

Synaptic adhesion molecule

HEK 293:

Human embryonic kidney

COS:

Cercopithecus aethiops kidney

MFD:

Microfluidic devices

TIRF:

Total internal reflection fluorescence

CFP:

Cyan fluorescent protein

P0:

Postnatal day 0

Fiji:

ImageJ image processing package (fiji.sc/Fiji)

Amaxa:

Electroporation system for gene delivering (bio.lonza.com)

MATLAB:

Matrix laboratory technical computing language (mathworks.com)

GNU Octave:

Matrix laboratory technical computing language (gnu.org/software/octave/)

Imaris:

Interactive microscopy image analysis software (bitplane.com)

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Acknowledgments

Thank you to Dr. Laura Swan for reading and suggestions to the manuscript.

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Author notes

    Authors and Affiliations

    1. Department of Neuroscience, School of Medicine, Tufts University, Boston, 02111, MA, USA

      Karen Perez de Arce

    2. Department of Cellular and Molecular Physiology, Institute of Translational Medicine, University of Liverpool, Crown Street, Liverpool, L69 3BX, UK

      Massimiliano Stagi

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    1. Karen Perez de Arce
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    2. Massimiliano Stagi
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    Correspondence to Massimiliano Stagi .

    Editor information

    Editors and Affiliations

    1. Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, Massachusetts, USA

      Alexandros Poulopoulos

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    Perez de Arce, K., Stagi, M. (2017). Co-culture Synaptogenic Assay: A New Look at Fluorescence Reporters and Technological Devices. In: Poulopoulos, A. (eds) Synapse Development. Methods in Molecular Biology, vol 1538. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6688-2_2

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    • DOI: https://doi.org/10.1007/978-1-4939-6688-2_2

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    • Publisher Name: Humana Press, New York, NY

    • Print ISBN: 978-1-4939-6686-8

    • Online ISBN: 978-1-4939-6688-2

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