Extended Data Fig. 3: Confetti analyses of the differentiation of monocytes and dendritic cells.

a, Histograms showing the observed (colour) and random (white) distribution of distances from each MOP to the closest indicated cells (MOP–Ly6C^hi monocyte, n = 137 MOPs from a total of 3 sternum sections from 3 mice; MOP–Ly6C^lo monocyte, n = 171 MOPs from a total of 4 sternum sections from 4 mice; MOP–cDCs, n = 200 MOPs from a total of 5 sternum sections from 4 mice). b, Maps showing the location of Gfi1^hi and Gfi1^lo MOPs, MDPs, Ly6C^hi monocytes and Ly6C^lo monocytes in a sternum segment from Gfi1–tomato mice. Scale bar, 200 μm. c, Number of Gfi1^hi and Gfi1^lo MOPs per sternum segment. Each dot represents one sternum segment from three Gfi1–tomato mice in two experiments. d, Histograms showing the observed (colour) and random (white) distribution of distances from each Gfi1^hi MOP (orange) and Gfi1^loMOP (purple) to the closest indicated cells. n = 113 Gfi1^hi MOPs and n = 72 Gfi1^lo MOPs from a total of 3 sternum segments from 3 mice. e, Representative image showing the lack of contribution of a confetti-labelled MOP to surrounding monocytes. Tracked cells are YFP⁺, CFP⁺ or unlabelled CD117⁺ CD115⁺ CD11b⁻ Ly6C⁺ MOPs, CD117⁻ CD115⁺ CD11b⁺ Ly6C^hi monocytes and CD117⁻ CD115⁺ CD11b⁺ Ly6C^lo monocytes. Scale bar, 20 μm. f, Quantification of cell numbers for CFP⁻ (white) and CFP⁺ (blue) Ly6C^hi monocytes (top) or Ly6C^lo monocytes (bottom) found within the indicated distances to the closest CFP⁺ MOP cell. Each dot represents one CFP⁺ MOP (n = 9 MOPs from a total of 6 sternum segments from 4 confetti mice). g, h, Histograms showing the observed (colour) and random (white) distribution of distances from each Ly6C^lo monocyte (yellow) and cDC (pink) to the six closest indicated neighbour cells. n = 1,603 Ly6C^lo monocytes from a total of 3 sternum segments from 3 mice; n = 1,228 cDCs from a total of 6 sternum segments from 6 mice. i, Images showing the lack of contribution of a CFP⁺ MDP to surrounding Ly6C^lo monocytes. Tracked cells are YFP⁺, CFP⁺ or unlabelled CD117⁺ CD115⁺ CD11b⁻ Ly6C⁻ MDPs, CD117⁺ CD115⁺ CD11b⁻ Ly6C⁺ MOPs, CD117⁻ CD115⁺ CD11b⁺ Ly6C^hi monocytes and CD117⁻ CD115⁺ CD11b⁺ Ly6C^lomonocytes. Scale bar, 20 μm. j, Images showing a lack of association between confetti-labelled Ly6C^hi and Ly6C^lo monocytes. Tracked cells are YFP⁺, CFP⁺ or unlabelled CD11b⁺ CD115⁺ CD117⁻ Ly6C^hi monocytes and CD11b⁺ CD115⁺ CD117⁻ Ly6C^lo monocytes. Scale bars for main and zoomed-in images, 40 μm and 10 μm respectively. The histograms show the observed (colour) and random (white) distribution of distances from each confetti-labelled Ly6C^hi (blue) or Ly6C^lo (yellow) monocytes to the closest Ly6C^hi or Ly6C^lo monocyte in the same colour. n = 48 confetti-labelled Ly6C^hi monocytes, and n = 32 confetti-labelled Ly6C^lo monocytes, from a total of 3 sternum sections from 3 mice. k, Images showing a lack of association between confetti labelled cDCs. Tracked cells are RFP⁺, GFP⁺, YFP⁺, CFP⁺ or unlabelled MCH II⁺ reticulated cDCs. Scale bars for main and zoomed-in images, 40 μm and 10 μm respectively. The histogram shows the observed (colour) and random (white) distribution of distances from each confetti-labelled cDC (pink) to the closest cDC in the same colour. n = 80 confetti-labelled cDCs from a total of 3 sternum sections from 3 mice. Unless otherwise indicated, for all graphs one dot corresponds to one cell. Horizontal blue bars indicate the median distance. Statistical differences were calculated using two-tailed Student’s t-tests; P values are shown.

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