Rapid Prototyping of 3D Biochips for Cell Motility Studies Using Two-Photon Polymerization.

Federico Sala,Rebeca Martínez Vázquez,Josef A Käs,Carlotta Ficorella,Hannah Marie Eichholz,Roberto Osellame

doi:10.3389/fbioe.2021.664094

Abstract

The study of cellular migration dynamics and strategies plays a relevant role in the understanding of both physiological and pathological processes. An important example could be the link between cancer cell motility and tumor evolution into metastatic stage. These strategies can be strongly influenced by the extracellular environment and the consequent mechanical constrains. In this framework, the possibility to study the behavior of single cells when subject to specific topological constraints could be an important tool in the hands of biologists. Two-photon polymerization is a sub-micrometric additive manufacturing technique that allows the fabrication of 3D structures in biocompatible resins, enabling the realization of ad hoc biochips for cell motility analyses, providing different types of mechanical stimuli. In our work, we present a new strategy for the realization of multilayer microfluidic lab-on-a-chip constructs for the study of cell motility which guarantees complete optical accessibility and the possibility to freely shape the migration area, to tailor it to the requirements of the specific cell type or experiment. The device includes a series of micro-constrictions that induce different types of mechanical stress on the cells during their migration. We show the realization of different possible geometries, in order to prove the versatility of the technique. As a proof of concept, we present the use of one of these devices for the study of the motility of murine neuronal cancer cells under high physical confinement, highlighting their peculiar migration mechanisms.

Highlights

Additive manufacturing, known as “3D printing,” is nowadays a standard technique for the realization of 3D objects
The migration area fits into a 5× objective field of view, it can be used for cell tracking experiments without the need of device realignment when monitored by microscope
The constriction region geometry can be reorganized depending on the experimental cell migration behavior under study

Summary

Introduction

Known as “3D printing,” is nowadays a standard technique for the realization of 3D objects. The sub-micron counterpart of 3D printing is direct laser writing, called twophoton polymerization (2PP) (Lee et al, 2006). This technique makes use of ultrashort laser pulses, usually in the femtosecond regime, to trigger the polymerization of specific liquid-phase photosensitive resists by multiphoton absorption. The use of this non-linear absorption process guarantees that only within the focal volume (voxel) of the laser beam the energy intensity. Thanks to the combination of small feature size and biocompatibility of some resists, one of the most interesting application is the realization of structures for biological studies (Nguyen and Narayan, 2017).

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