Polyethylene glycol molecular weight influences the ClearT2 optical clearing method for spheroids imaging by confocal laser scanning microscopy

Elisa C. Costa; André F. Moreira; Duarte de Melo-Diogo; Ilidio J. Correia

doi:10.1117/1.JBO.23.5.055003

11 May 2018 Polyethylene glycol molecular weight influences the Clear^T2 optical clearing method for spheroids imaging by confocal laser scanning microscopy

Elisa C. Costa, André F. Moreira, Duarte de Melo-Diogo, Ilidio J. Correia

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Journal of Biomedical Optics, Vol. 23, Issue 5, 055003 (May 2018). https://doi.org/10.1117/1.JBO.23.5.055003

Abstract

Some fluorescence microscopy techniques, such as confocal laser scanning microscopy (CLSM), have a limited penetration depth. Consequently, the visualization and imaging of three-dimensional (3-D) cell cultures, such as spheroids, using these methods can be a significant challenge. Therefore, to improve the imaging of 3-D tissues, optical clearing methods have been optimized to render transparency to the opaque spheroids. The influence of the polyethylene glycol (PEG) molecular weight (MW) used in the Clear^T2 method for the imaging of propidium iodide (PI)-stained spheroids was investigated. The results demonstrated that the Clear^T2 clearing method contributes to spheroids transparency and to the preservation of PI fluorescence intensity for all the PEG MW used (4000, 8000, and 10,000 Da). Furthermore, the Clear^T2 method performed using PEG 4000 Da allowed a better PI signal penetration depth and cross-section depth. Overall, the optimization of PEG MW can improve the imaging of intact spheroids by CLSM. Furthermore, this work may also contribute to increase the application of 3-D cell culture models by the pharmaceutical industry for the high-throughput screening of therapeutics.

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Elisa C. Costa, André F. Moreira, Duarte de Melo-Diogo, and Ilidio J. Correia "Polyethylene glycol molecular weight influences the Clear^T2 optical clearing method for spheroids imaging by confocal laser scanning microscopy," Journal of Biomedical Optics 23(5), 055003 (11 May 2018). https://doi.org/10.1117/1.JBO.23.5.055003

Received: 3 February 2018; Accepted: 17 April 2018; Published: 11 May 2018

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