Grafting of iLNs or reaggregate LNs results in the formation of lymphoid structures. (A,B) Grafting of a P1 iLN under the kidney capsule of an adult mouse results in the formation of a lymphoid structure after 2 weeks. The red rectangle in panel A (25×/1.4) contains the graft that is enlarged in panel B (40×/1.4). (C) FACS analysis of an E15 embryonic iLN grafted for 2 weeks under the kidney capsule of an adult mouse and the iLN of the host. As shown by forward/side-scatter profile, the E15 grafts contained only dead cells. Results are representative of 6 independent experiments. (D) E17 and P1 iLN grafts contained CD4⁺ and CD8⁺ T cells as well as CD19⁺ B cells. Percentages shown in dot plots correspond to single positive CD4, CD8, and CD4⁻8⁻ cells. Histograms show the percentage of CD19⁺ B cells. Results shown are representative of 5 independent assays. (E) Bone marrow–derived cells colonizing the kidney-grafted iLNs are of host origin. Single-cell suspensions of wild-type congenic (CD45.1) iLNs grafted for 2 weeks under the kidney capsule of C57Bl6J (CD45.2) mice were analyzed for the expression of CD45.1/CD45.2 markers to assess their origin. Percentages shown correspond to donor iLN CD45.1⁺ cells and host CD45.2⁺ cells. As shown here, more than 98% of the cells are of host origin. (F) Reaggregate iLNs before being grafted under the kidney capsule of an adult recipient mouse (40×/1.4). (G) The iLN reaggregate (blue circle) shown on the kidney 2 weeks after grafting (25×/1.4). (H) Reaggregate iLNs recruited B and T cells similar to the LNs of the host. Percentages shown correspond to B220⁺ B cells and CD3⁺ T cells in host iLNs and kidney-grafted reaggregate iLNs. (I) Reaggregate iLNs were colonized by lymphocytes that migrated to specific areas in the organ. Immunofluorescence staining of sections of an adult host iLN and a P1 reaggregate iLN that had been grafted for 2 weeks under the kidney capsule of the host showing B220⁺ B-cell follicular areas (red) in the cortex and the CD3⁺ T-cell areas (green) located in the paracortex. The organization of B- and T-cell areas appeared similar between the host iLN and the reaggregate organ, although B cells appear less packed in the case of the latter (10×/1.4 NA). Results shown are representative of 6 to 8 independent experiments. (A,B,F,G,I) Images were viewed with a Zeiss Stemi SVII dissecting microscope.