Clearing method adapted to FFPE tissues for 3D imaging of nerve fibers, B cells, and tertiary lymphoid structures
Biomedical samples are commonly used for histological examination after their inclusion in paraffin (Formalin-Fixed Paraffin-Embedded (FFPE) biopsy). However, they provide minimal information about the interactions between different tissue components such as blood vessels, nerves, and cellular aggregates. Here we present a modified iDISCO tissue-clearing method that we term miDISCO+. miDISCO+ can be used for analyzing FFPE samples, requires the use of only one single FFPE sample, and allows to acquire a detailed image of the 3D tissue/organ architecture and the relevant cellular interactions. The addition of an antigen retrieval step to the protocol enables to use antibodies whose binding is sensitive to formalin fixation due to antigen masking. This method enabled us to detect CD20+ B cell follicles and to show that they are in close contact with TH+ sympathetic nerve fibers in FFPE biopsies of human palatine tonsils, and to detect CD20+ B cells in lung tumors and observe their 3D organization within tertiary lymphoid structures. Thus, miDISCO+ could be a potent tool for clinicians to refine diagnosis and to select optimal personalized treatment by giving an integrated 3D view of the tissue structures and cellular interactions in single FFPE samples.