Additional file 1 of Mn-single-atom nano-multizyme enabled NIR-II photoacoustically monitored, photothermally enhanced ROS storm for combined cancer therapy

Additional file 1: Figure S1. TEM image of N-HCN. Figure S2. SEM image of Mn/N-HCN. Figure S3. The average size (A) and polydispersity index (B) of Mn/N-HCN in water at different time. Figure S4. Zeta potential of Mn/N-HCN. Figure S5. XPS spectra of Mn/N-HCN. Figure S6. PEXAFS fitting curves of (A) Mn2O3, (B) MnO, and (C) Mn foil in R space. Figure S7. The proposed Mn-N4-C local environment of Mn/N-HCN. Figure S8. ESR spectra of •OH with characteristic quartet signal (1:2:2:1) of DMPO-OH. Figure S9. Time-course absorbance changes of TMB with the addition of different concentrations of H2O2. Figure S10. TMB assay for measuring POD-mimic activity of Mn/N-HCN from pH 4.6 to pH. Figure S11. (A) Michaelis-Menten kinetics curves for the POD-mimic activity of Mn/N-HCN, (B) corresponding Lineweaver-Burk plots. Figure S12. TMB assay for measuring POD-mimic activity of Mn/N-HCN. Figure S13. pH-dependent CAT-mimic activity of Mn/N-HCN. Figure S14. ESR spectra of •O2- with characteristic peaks 1:1:1:1 of DMPO-OOH. Figure S15. ABDA assay for 1O2 generation by Mn/N-HCN, in the presence of SOD. Figure S16. GSH depletion by Mn/N-HCN during different times. Figure S17. UV-Vis-NIR spectra of N-HCN and Mn/N-HCN. Figure S18. Calculation of photothermal conversion efficiency of Mn/N-HCN (100 µg/mL) under irradiation with 1064 nm laser (1 W/cm2). Red curve: Photothermal profile of Mn/N-HCN irradiated for 7 min. Blue curve: followed by nature cooling. Linear time data versus -ln(θ) obtained from the cooling period. Figure S19. Temperature of Mn/N-HCN containing PBS solution subjecting to on/off cycling of laser irradiation. Figure S20. Uptake of FITC-label Mn/N-HCNs by 4T1 cells at different times. Figure S21. (A) Cell viability of L929 cells was detected by the CCK-8 method after 12 h-treatment of Mn/N-HCN at different concentrations. (B) Cell viability of 4T1 cells after 12 h-treatment of Mn/N-HCN at different concentrations and conditions. Figure S22. Live/dead cell assay. The green signal from Calcein-AM indicates live cells and the red signal from PI indicates dead cells. Scale bars=50 μm. Figure S23. Flow cytometric measurement on cells co-stained with Annexin V-FITC and PI. Figure S24. Intracellular O2 generation using [Ru(dpp)3]Cl2 as a probe: (I) Control; (II) Mn/N-HCN+H2O2 (100 μM); (III) Mn/N-HCN+H2O2+L. Scale bars=50μm. Figure S25. The photoacoustic spectra of the Mn/N-HCN. The wavelength range is from 1200 nm to 1600 nm. Figure S26. (A) NIR-II (1200 nm) PA signal of Mn/N-HCN at different concentrations in PBS. (B) PA intensity to nanoparticle concentration with a linear fitting. Figure S27. In vitro haemocompatibility assay of Mn/N-HCN with different concentrations ranges from 0 to 200 µg/mL. Figure S28. Biodistribution of Mn/N-HCN in various organs after 6 h post-injection. Figure S29. Photographs of tumors at 1, 7, and 14 day after different treatments (I Control, II PBS+laser, III Mn/N-HCN, IV Mn/N-HCN+Laser). Figure S30. Tumor Inhibition Rate after different treatments (I Control, II PBS+laser, III Mn/N-HCNs, IV Mn/N-HCN+Laser). Figure S31. Body weight during 14 days (I Control, II PBS+laser, III Mn/N-HCNs, IV Mn/N-HCN+Laser). Figure S32. H&E staining of major organs (heart, liver, spleen, lung, and kidney) dissected from the BALB/c mice with different treated groups (I Control, II PBS+laser, III Mn/N-HCN, IV Mn/N-HCN+Laser) for post 14 days injection (Scale bar: 50 μm). Table S1. Parameters of EXAFS fits for Mn foil, MnO, Mn2O3, and Mn/N-HCN