A novel cryopreservation and biobanking strategy to study lymphoid tissue stromal cells in human disease

Non-hematopoietic lymph node stromal cells (LNSCs) regulate lymphocyte trafficking, survival, and function for key roles in host defense, autoimmunity, alloimmunity, and lymphoproliferative disorders. However, study of LNSCs in human diseases is complicated by a dependence on viable lymphoid tissues, which are most often excised prior to establishment of a specific diagnosis. Here, we demonstrate that cryopreservation can be used to bank lymphoid tissue for the study of LNSCs in human disease. Using human tonsils, lymphoid tissue fragments were cryopreserved for subsequent enzymatic digestion and recovery of viable non-hematopoietic cells. Flow cytometry and single-cell transcriptomics identified comparable proportions of LNSC cell types in fresh and cryopreserved tissue. Moreover, cryopreservation had little effect on transcriptional profiles, which showed significant overlap between tonsils and lymph nodes. The presence and spatial distribution of transcriptionally defined cell types was confirmed by in situ analyses. Our broadly applicable approach promises to greatly enable research into the roles of LNSC in human disease. ### Competing Interest Statement JDB has consulted for EUSA Pharma. IM has received research funding from Genentech and Regeneron, and he is a member of Garuda Therapeutics scientific advisory board. JCA has consulted for Ayala Pharmaceuticals, Cellestia, Inc., and Remix Therapeutics. All conflicts are unrelated to the contents of this manuscript. * LNSC : Lymph node stromal cell LN : Lymph node VSMC : Vascular smooth muscle cell TRC : T cell zone reticular cell PRCs : Perivascular reticular cell BRC : B cell-interacting reticular cell FDC : Follicular dendritic cell scRNAseq : Single-cell RNA sequencing