Abstract
The delicate structure of murine lungs poses many challenges for acquiring high-quality images that truly represent the living lung. Here, we describe several optimized procedures for obtaining and imaging murine lung tissue. Compared to traditional paraffin cross-section and optimal cutting temperature (OCT), agarose-inflated vibratome sections (aka precision-cut lung slices), combines comparable structural preservation with experimental flexibility. In particular, we discuss an optimized procedure to precision-cut lung slices that can be used to visualize three-dimensional cell-cell interactions beyond the limitations of two-dimensional imaging. Super-resolution microscopy can then be used to reveal the fine structure of lung tissue's cellular bodies and processes that regular confocal cannot. Lastly, we evaluate the entire lung vasculature with clearing technology that allows imaging of the entire volume of the lung without sectioning. In this manuscript, we combine the above procedures to create a novel and evolutionary method to study cell behavior ex vivo, trace and reconstruct pulmonary vasculature, address fundamental questions relevant to a wide variety of vascular disorders, and perceive implications to better imaging clinical tissue.
Highlights
High resolution imaging of intact tissue is vital to understanding the anatomy and physiology underlying human disease
Detailed imaging of murine lung tissue that preserves the structure and cellular components helps advance the knowledge and understanding of diseases so therapeutic approaches can be applied to human subjects
Some antigens can be destroyed by the high-temperature processing used in “unmasking” and are no longer recognized by their designated antibodies
Summary
High resolution imaging of intact tissue is vital to understanding the anatomy and physiology underlying human disease. Detailed imaging of murine lung tissue that preserves the structure and cellular components helps advance the knowledge and understanding of diseases so therapeutic approaches can be applied to human subjects. The ability to label protein, RNA, and other biological compounds with a high signal-to-noise ratio while maintaining physiological structure broadens the understanding of pathology and can demonstrate the spatial approximation of different biological compounds and cell types in situ. The inherent difficulties in lung tissue preparation are the technical bottleneck. Sub-optimally prepared lung tissue can lead to inaccurate alveolar architecture, light scattering and difficulties with fluorescent staining, which may potentially produce an inaccurate model
Sign-in/Register to view highlights & summary Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
