Use of High-Refractive Index Hydrogels and Tissue Clearing for Large Biological Sample Imaging

Gels

卷:8

Sandra Fok¹  Nick Di Girolamo²  Blake J. Cochran²  Kerry-Anne Rye²  Victoria Lee²  Alexander Richardson² 

[1] Katharina Gaus Light Microscopy Facility, Mark Wainwright Analytical Centre, UNSW Sydney, Sydney, NSW 2052, Australia;

[2] School of Medical Sciences, UNSW Sydney, Sydney, NSW 2052, Australia;

关键词: hydrogel;    tissue clearing;    sample preparation;    light sheet microscopy;   

DOI  :  10.3390/gels8010032

来源: DOAJ

【 摘 要 】

Recent advances in tissue clearing and light sheet fluorescence microscopy have improved insights into and understanding of tissue morphology and disease pathology by imaging large samples without the requirement of histological sectioning. However, sample handling and conservation of sample integrity during lengthy staining and acquisition protocols remains a challenge. This study overcomes these challenges with acrylamide hydrogels synthesised to match the refractive index of solutions typically utilised in aqueous tissue clearing protocols. These hydrogels have a high-water content (82.0 ± 3.7% by weight). The gels are stable over time and FITC-IgG readily permeated into and effluxed out of them. Whilst the gels deformed and/or swelled over time in some commonly used solutions, this was overcome by using a previously described custom refractive index matched solution. To validate their use, CUBIC cleared mouse tissues and whole embryos were embedded in hydrogels, stained using fluorescent small molecule dyes, labels and antibodies and successfully imaged using light sheet fluorescence microscopy. In conclusion, the high water content, high refractive index hydrogels described in this study have broad applicability to research that delves into pathophysiological processes by stabilising and protecting large and fragile samples.

【 授权许可】

Unknown