Abstract
Spatial information of cells in their tissue microenvironment is necessary to understand the complexity of pathophysiological processes. Volumetric imaging of cleared organs provides this information; however, current protocols are often elaborate, expensive, and organ specific. We developed a simplified, cost-effective, non-hazardous approach for efficient tissue clearing and multi-organ volumetric imaging (EMOVI). EMOVI enabled multiplexed antibody-based immunolabeling, provided adequate tissue transparency, maintained cellular morphology and preserved fluorochromes. Exemplarily, EMOVI allowed the detection and quantification of scarce cell populations during pneumonitis. EMOVI also permitted histo-cytometric analysis of MHC-II expressing cells, revealing distinct populations surrounding or infiltrating glomeruli of nephritic kidneys. Using EMOVI, we found widefield microscopy with real-time computational clearing as a valuable option for rapid image acquisition and detection of rare cellular events in cleared organs. EMOVI has the potential to make tissue clearing and volumetric imaging of immune cells applicable for a broad audience by facilitating flexibility in organ, fluorochrome and microscopy usage.
Highlights
During immune homeostasis and inflammatory challenges, the communication between cells is crucial for a concerted immune response
Our results demonstrate that efficient tissue clearing and multi-organ volumetric imaging (EMOVI) enables multiplexed antibody-based immunolabeling, provides adequate tissue transparency, and maintains cellular morphology
We further demonstrate that EMOVI can be used to image and quantify changes in sparse cell populations in the lung during systemic inflammation
Summary
During immune homeostasis and inflammatory challenges, the communication between cells is crucial for a concerted immune response. This communication relies to a large part on the precise positioning of individual cell types in specific anatomical locations (the immune niche) [1, 2]. Our current understanding of these processes still mainly relies on single-cell analysis of cells that are extracted from dissociated or lysed tissue. In this process, information such as cellular interactions, behavior, and localization is lost.
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