Fig. 2

MitoCeption protocol update. (1) Schema of the updated protocol. Unattached recipient cells can be MitoCepted in a microcentrifuge tube of 1.5 ml and then centrifuged at 500 x g at during 5 min and immediately put in incubation for at least 1 h with mitochondria. Then, cells can be washed of excess mitochondria and used for downstream applications. Graphs show the proportional increase of the transfer of fluorescent mitochondria in percentages and MFIs due to the uptake in relation to the control of non-MitoCepted mitochondria. PAMM is composed of the PBMCs of at least three donors. (2, 3) MitoCepted CD3+ Lymphocytes and CD14+ Monocytes (n = 3, 3 PBMC donors, 3 PBMC donors for PAMM). Fresh PBMCs were MitoCepted with isolated mitochondria labeled with MitoTracker® Green from PBMCs; lymphocytes and monocytes were selected by their size, granularity, singles, alive cells (− for Annexin and 7AAD), and CD3+ and CD14+ identification. (2.2a, 2.2c, 2.2d) Flow cytometry of CD3+ cells after 1 h of MitoCeption. (2.2b, 2.2e, 2.2f) CD3+ after wash of excess mitochondria and 18 h of MitoCeption. (2.2c) CD3+ percentage of MitoCeption cells after 1 h. (2.2d) CD3+ MFI of the MitoCepted cells after 1 h. (2.2e) CD3+ Percentage of MitoCeption cells after 18 h. (2.2f) CD3+ MFI of the MitoCepted cells after 18 h. (2.3a, 2.3c, 2.3d) Flow cytometry of CD14+ cells after 1 h of MitoCeption. (2.3b, 2.3e, 2.3f) Flow cytometry of CD14+ cells after 18 h of MitoCeption. (2.3c) CD14+ percentage of MitoCeption cells after 1 h. (2.3d) CD14+ MFI of the MitoCeption cells after 1 h. (2.3e) CD14+ Percentage of MitoCeption cells after 18 h. (2.3f) CD14+ MFI of the MitoCepted cells after 18 h. Statistical analysis for all conditions: Mean ± SEM one-way ANOVA and Tuckey’s pot-test. (*p < 0.05, **p < 0.01, ***p < 0.001)