Anti gr 1

In the BQ123-induced PMN-MDSC depletion experiment, 10 µg/g of anti-Gr1 (BioXcell, West Lebanon, New Hampshire, and USA) was administered intraperitoneally to WT mice. C57BL/6 mice were administered BQ123 or PBS via the same route as mentioned above before model induction. For the colitis mouse model, Gr1ab was injected into recipient mice on days 2 and 5 of the model. For the lung inflammation mouse model, Gr1ab was injected into recipient mice on days 0 and 3, for the acute hepatitis model, Gr1ab was injected into recipient mice 1 h before receiving the ConA injection on day 0. The rat IgG isotype was used as a control.

Cellular subsets and cytokines were depleted by intraperitoneally administering depleting antibodies (BioXCell) beginning 1 d before therapy as we previously reported (Moynihan et al., 2016 (link)): CD8 T cells with anti-CD8-α (clone 2.43, 400 μg every 3 days), CD4 T cells with anti-CD4 (clone GK1.5, 400 μg every 3 days), NK cells with anti-NK1.1 (clone PK136, 400 μg every 3 days), neutrophils with anti-Gr-1 (clone RB6–8C5, 400 μg every 2 days), macrophages with anti-F4/80 (clone CI:A3–1, 200 μg every day) (Lin et al., 2017 (link)), IFN-γ with anti- IFN-γ (clone XT3.11, 200 μg every 3 days), TNF-α with anti-TNF-α (clone XMG1.2, 500 μg every 2 days) and CXCL9 with anti-CXCL9 (clone MIG-2F5.5, 300 μg every 2 days). VEGFR2 was blocked by anti-VEGFR2 (clone DC101, 500 μg every 3 days). Apoptosis of intratumoral T cells were induced with anti-CD3ε F(ab’)2 (clone 145–2C11, 50 μg for intratumoral injection or 100 μg for systematic i.p. injection every day) (Besançon et al., 2017 (link)) to avoid toxicity associated with full anti-CD3 antibodies in treated mice (data not shown). Cellular depletions of CD3⁺ T cells, CD8⁺ T cells, CD4⁺ T cells, neutrophils, macrophages and NK cells were confirmed by flow cytometry of PBMCs (Figures S3A and S4A).

One hour before administration, clodronate (CLO) liposomes or control liposomes (Liposoma, Amsterdam, The Netherlands, #CP-005-005) were kept at room temperature. Per mouse, 200 µL liposomes (50 µg/g) was administered retro-orbitally after vortexing, using a 29 G insulin syringe (VWR, Leuven, Belgium,). Likewise, labeled BMDMs were vortexed prior to injection with a 29 G insulin syringe. Unless noted otherwise, 100 µL of cells suspended in DPBS were injected in the right orbital plexus of anesthetized mice at a concentration of 10 × 10⁶/mL [42 (link),43 (link)]. Anti-Gr-1 (Bioxcell, Lebanon, USA, #BE0075) or anti-Ly6G (Bioxcell, Lebanon, USA, #BE0075-1) depleting monoclonal antibodies were administered intraperitoneally at a dose of 250 µg per mouse in 100 µL DPBS [44 (link)].