High-content neurite development study using optically patterned substrates.

PloS One
Jonathan M BélisleSantiago Costantino

Abstract

The study of neurite guidance in vitro relies on the ability to reproduce the distribution of attractive and repulsive guidance molecules normally expressed in vivo. The identification of subtle variations in the neurite response to changes in the spatial distribution of extracellular molecules can be achieved by monitoring the behavior of cells on protein gradients. To do this, automated high-content screening assays are needed to quantify the morphological changes resulting from growth on gradients of guidance molecules. Here, we present the use of laser-assisted protein adsorption by photobleaching (LAPAP) to allow the fabrication of large-scale substrate-bound laminin-1 gradients to study neurite extension. We produced thousands of gradients of different slopes and analyzed the variations in neurite attraction of neuron-like cells (RGC-5). An image analysis algorithm processed bright field microscopy images, detecting each cell and quantifying the soma centroid and the initiation, terminal and turning angles of the longest neurite.

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Citations

Feb 7, 2014·Interface Focus·Beatrice ChelliFabio Biscarini
Sep 18, 2015·Frontiers in Bioengineering and Biotechnology·Anna LagunasJosep Samitier
Jan 13, 2018·Journal of Ocular Pharmacology and Therapeutics : the Official Journal of the Association for Ocular Pharmacology and Therapeutics·Najam A Sharif
May 8, 2017·Microsystems & Nanoengineering·Nirveek Bhattacharjee, Albert Folch
Nov 20, 2016·Journal of Cell Science·Verena RuprechtVirgile Viasnoff

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Methods Mentioned

BETA
fluorescence microscopy

Software Mentioned

Image Processing
LAPAP
MATLAB
Labview

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