Non-linear excitation microscopy of live cells by using an implantable microscope objective-on-a-chip

A. Nardini; R. Martínez Vázquez; C. Conci; M. Grassi; M. Marini; M. Bouzin; M. Collini; R. Osellame; G. N. Cerullo; B. S. Kariman; M. T. Raimondi; G. Chirico

doi:10.1117/12.3001604

12 March 2024 Non-linear excitation microscopy of live cells by using an implantable microscope objective-on-a-chip

A. Nardini, R. Martínez Vázquez, C. Conci, M. Grassi, M. Marini, M. Bouzin, M. Collini, R. Osellame, G. N. Cerullo, B. S. Kariman, M. T. Raimondi, G. Chirico

Author Affiliations +

Proceedings Volume 12846, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XXII; 1284607 (2024) https://doi.org/10.1117/12.3001604
Event: SPIE BiOS, 2024, San Francisco, California, United States

Abstract

In the context of biomaterials, small-molecules and drugs testing, intravital microscopy allows to quantify in-vivo the immune reaction, reducing the number of laboratory animals required to statistically validate the product. However, fluorescence microscopy is affected by limited tissue penetration due to light scattering and by optical aberrations, induced on focused beams, by the animal tissue surrounding the implant. In this framework, we developed a system of microlenses coupled to microscaffolds, both incorporated in a miniaturized imaging window. The chip is designed to act as an in-situ microscope objective with the aim to overcome the restrictions of in-vivo imaging (i.e. spherical aberrations) and to allow multiple biological observations in the same animal (by including fluorescent beacons). The device is fabricated by two-photon polymerizing a biocompatible photoresist called SZ2080. The microlenses are manufactured by the concentric polar scanning of the laser beam to realize their outer surface, followed by the UV bulk polymerization of their inner SZ2080. We preliminarily characterized the imaging capabilities of our implantable system on live cells cultured on flat substrates and 3D microscaffolds by coupling it to low magnification objectives. The microlenses optical quality is sufficient to induce non-linear excitation and collect two-photon excitation images with the same level of laser intensity and signal-to-noise ratio. Remarkably, they allow to efficiently excite the fluorescence of labelled human fibroblasts collecting high resolution magnified images. These results will open the way to the application of implanted micro-optics for the real-time and continuous in-vivo observation of complex biological processes.

Conference Presentation

(2024) Published by SPIE. Downloading of the abstract is permitted for personal use only.

Citation Download Citation

A. Nardini, R. Martínez Vázquez, C. Conci, M. Grassi, M. Marini, M. Bouzin, M. Collini, R. Osellame, G. N. Cerullo, B. S. Kariman, M. T. Raimondi, and G. Chirico "Non-linear excitation microscopy of live cells by using an implantable microscope objective-on-a-chip", Proc. SPIE 12846, Imaging, Manipulation, and Analysis of Biomolecules, Cells, and Tissues XXII, 1284607 (12 March 2024); https://doi.org/10.1117/12.3001604

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