Fig. 1

MBCs scan SCS macrophages for antigen. a Tiled maximum intensity projection (3072 × 2048 × 141 μm) of lymph node 70 days after HEL-OVA primary immunisation. MBC spots (white), CD169⁺ SCS macrophages (pink), SHG (blue). See also Supplementary Movie 1. Scale bar = 500 μm. b Maximum intensity projection (300 × 300 × 48 μm) of follicle 32 days after HEL-OVA immunisation. MBC SW_HEL tracks (red), naive SW_HEL B cell tracks (green), CD169⁺ SCS macrophages (pink), SHG (blue). See also Supplementary Movie 2. Scale bar = 50 μm. c Proportion of naive B cells (blue) or MBCs (red) in proximity (<20 μm) to CD169⁺ SCS macrophage surface at any given time point, number time points = 46, quantified from b where number naive cells = 74, number MBCs = 40. Box plots show centre line as median, box limits as upper and lower quartiles, whiskers as minimum to maximum values. d Maximum intensity projection of follicle 54 days after HEL-OVA primary immunisation, showing colocalisation (white) of a MBC (red) with CD169⁺ SCS macrophages (pink) over time. Interactions with three different SCS macrophages are highlighted in shades of pink. See also Supplementary Movie 3. Scale bar = 5 μm. e Speed of MBCs segmented based on distance from CD169⁺ SCS macrophage surface: far (<20 μm) (red) or close (>20 μm) (blue). Number of data points = 1827. Data are combined from 4 movies. Comparison between groups utilised one-tailed unpaired Student’s t-test. f Arrest coefficient of naive B cells (blue) or MBCs (red), defined as percentage of time a cell slowed down to <3μm/min. Number of naive cells = 733, MBCs = 202. Data are pooled from 4 movies. Comparison between groups utilised one-tailed unpaired Student’s t-test. g Number of donor-derived MBCs in draining inguinal lymph node (dLN) (red) compared to non-draining popliteal lymph node (ndLN) (blue). Data are from 15 mice and representative of 2 independent experiments. Comparison between groups utilised one-tailed unpaired Student’s t-test. h Identification of unphotoconverted Kaede green MBCs, photoconverted Kaede red MBCs and endogenous B cells in the draining lymph node by FACs analysis, example shown is 2 days after photoconversion. Box plots show centre line as median, box limits as upper and lower quartiles, whiskers as minimum to maximum values. i Kaede MBCs in the draining lymph node were photoconverted, allowed to recover from anesthesia and then harvested 1–4 days later. Graph shows proportion of photoconverted cells to non-photoconverted cells in the lymph node at indicated time points