Supporting Information

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1 Supporting Information Detection and Imaging of Hydrogen Sulfide in Lysosome of Living Cells with an Activatable Fluorescence Quantum Dots Yiwei Wu a, Qiuyue Wang a, Tingting Wu b, Wei Liu a, Hexin Nan a, Shenghao Xu c, Yizhong Shen b, a Hubei Key Laboratory of Pollutant Analysis & Reuse Technology, College of Chemistry and Chemical Engineering, Hubei Normal University, Huangshi , China b School of Food & Biological Engineering, Engineering Research Center of Bio-Process, Ministry of Education, Hefei University of Technology, Hefei, , China c Key Laboratory of Sensor Analysis of Tumor Marker, Ministry of Education, Qingdao University of Science and Technology, Qingdao , China Corresponding author. address: yzshen@hfut.edu.cn (Y. Shen) S-1

2 Table of contents Page 1 Figure S1-7 & Table S1, S2 S3-9 2 Supplementary experiments S10 3 References S10-12 S-2

3 SUPPLEMENTARY FIGURES Figure S1. DLS of (a) QDs, (b) AgNPs, and (c, d) QDs/AgNPs nanocomplexes before and after incubation with H 2 S in D.I. water. S-3

4 Figure S2. Fluorescence characterization of the assembled QDs/AgNPs nanocomplexes. (a) Fluorescence spectra of QDs (3.5 μm) following added with varying concentrations (0, 28, 56, 70, 84, 98, 140, 154, 196, 210, 224, 238, 252 and 266 pm) of AgNPs in HEPES buffer solution at ph = 7.0. (b) The corresponding fluorescence quenching efficiency (%) of QDs at 530 nm in the absence and presence of AgNPs at different concentrations as described Fig. S2a. (c) Fluoresce spectra of QDs (3.5 μm) following added with 238 pm AgNPs complex for different times (0, 1, 2, 3, 4, 5, 9, 10, 11, 12, 13, 14, 15, 16 and 17 min) in HEPES buffer solution at ph = 7.0. (d) The corresponding fluorescence quenching efficiency (%) of QDs at 530 nm vs incubation time as described Figure S2c. All fluorescence data were measured in HEPES buffers (ph 7.0) under the excitation of 450 nm. S-4

5 Table S1. Stern-Volmer quenching constants (K sv ) for the interaction of QDs with AgNPs solutions at three different temperatures ph 7.0 Temperature Correlation Stern Volmer linear equation (K) coefficient K sv (L mol -1 ) 293 F 0 /F = [Q] F 0 /F = [Q] F 0 /F = [Q] Figure S3. UV-vis absorption spectra of (a) QDs (3.5 μm), (b) AgNPs (238 pm), (c) the assembled QDs/AgNPs nanocomplexes (3.5 μm determined by QDs), and (d) the assembled QDs/AgNPs nanocomplexes (3.5 μm) + H 2 S (5.2 μm) in HEPES buffer solution at ph = 7.0. The purple arrow indicated the red shift from 502 nm to 508 nm. Figure S4. (a) Time-dependent UV-vis absorption spectra of 238 pm AgNPs incubation with 5.2 μm H 2 S in HEPES buffer solution at ph=7.0. (b) The corresponding curve of the OD value at ~ 400 nm vs incubation time as depicted in Figure S4a. S-5

6 Table S2. Comparison of the sensitivity of this proposed activatable fluorescent QDs/AgNPs nanoprobe with those of other methods for the determination of H 2 S. System Methods LOD (μm) Ref. Nanoporous gold-based microbial Electrochemical method biosensor Sodium 1,2-naphthoquinone-4-sulfonate Spectrophotometric method and response surface methodology Ciprofloxacin capped silver nanoparticles Colorimetric method Gold nanorods 24 4 Automated multi-syringe flow Chemiluminescence injection analysis system Sulfide in aqueous samples ICP-AES Mercaptopropionic acid-capped CdTe quantum dots Core-shell Au@Ag nanoclusters Functionalized CdS quantum dots Ligand-functionalized carbon dots Cyclam-functionalized carbon dots/cu 2+ Fluorescence Fluorescein/chelator/Cu 2+ complex Fluorescent polymeric nanoparticles/cu 2+ carbon dots/hg The assembled QDs/AgNPs nanoprobe This work S-6

7 Figure S5. Effect of ph on the stability of the assembled QDs/AgNPs nanocomplexes. (a-e) Fluorescence spectra of 3.5 μm QDs/AgNPs nanocomplexes incubation with 5.2 μm H 2 S in HEPES buffer solution at ph = 5, ph = 6, ph = 7, ph = 8, and ph = 9 for 0 s and 300 s. (d) The fluorescence activation fold (F/F 0 ) of 3.5 μm QDs/AgNPs nanocomplexes incubation with 5.2 μm H 2 S in HEPES buffer solution at ph = 5, ph = 6, ph = 7, ph = 8, and ph = 9 for 300 s, respectively. F 0 and F indicate the fluorescence intensity of 3.5 μm QDs/AgNPs nanocomplexes at 530 nm in the absence and presence of 5.2 μm H 2 S for 300 s, respectively. S-7

8 Figure S6. (a-d) Fluorescence spectra of 3.5 μm QDs/AgNPs nanocomplexes incubation with 5.2 μm H 2 S in HEPES buffer (ph = 7) solution and different cellular culture medium for 0 s and 300 s. (e) The fluorescence activation fold (F/F 0 ) of 3.5 μm QDs/AgNPs nanocomplexes incubation with 5.2 μm H 2 S in HEPES buffer solution and different cellular culture medium for 300 s, respectively. F 0 and F indicate the fluorescence intensity of 3.5 μm QDs/AgNPs nanocomplexes at 530 nm in the absence and presence of 5.2 μm H 2 S for 300 s, respectively. (f) The fluorescence stability of 3.5 μm QDs/AgNPs at 530 nm vs incubation time (0, 1.0, 2.0, 4.0, 8.0, 16, 32, 48, 64 and 72 h) in cellular culture medium (DMEM) under the excitation of 450 nm. All of cellular culture mediums were supplemented with fetal bovine serum (10%). S-8

9 Figure S7. Cell viability of HeLa cells after treatment with the assembled QDs/AgNPs nanocomplexes at different concentrations from 0 to 5.0 M (determined by QDs) for 24 h. Data shown were collected with the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Briefly, the HeLa cells were seeded into a 96-well plate at a density of ~ cells and incubated with 100 μl DMEM culture medium containing the assembled QDs/AgNPs nanocomplexes at different concentrations from 0 to 5.0 M (determined by QDs) for 24 h. Then, 50 μl of MTT (1 mg/ml) in PBS solution was added into each well. After incubation at 37 C for 4 h, the culture medium was removed carefully, and 150 L DMSO was added to each well. The absorbance (OD) at 490 nm in each well was acquired by a Bio-Rad microplate reader. The absorbance of blank cells (OD control ) were used as the control. The percentage of cell viability was calculated by dividing OD to OD control. Figure S8. Effects of different percentage alcohol (V/V) on the fluorescence stability of 3.5 μm QDs/AgNPs at 530 nm in HEPES buffer (ph 7.0) under the excitation of 450 nm. S-9

10 SUPPLEMENTARY EXPERIMENTS Investigation of Stern-Volmer quenching constant. By the titration experiments, 3.5 M QDs incubated with different concentrations of AgNPs including 0, 26, 52, 79, 105, and 131 pm in HEPES buffers (ph 7.0) at 293 K, 313 K, or 323 K for 5 min, respectively. Afterward, the fluorescence intensity at 530 nm originated from all samples were collected with excitation of 450 nm. All of these experiments were performed by a recycled water system that could effectively regulate the incubation temperature keep in constant. Hence, the temperature effect on the quenching process was explored through the well-known Stern-Volmer equation (1) 15,16 : F0 1 [ ] Eq. 1 SV F K Q In the expression, F 0 and F indicate the fluorescence intensities of QDs at 530 nm before and after incubation with AgNPs acted as a quencher, respectively. [Q] represent the concentration of the AgNPs, and K sv represent the Stern-Volmer quenching constant, which could be aquired from the linear-fitting curve of F 0 /F versus [Q] at three different temperatures (293 K, 313 K, and 323 K). REFERENCES (1) Liu, Z.; Ma, H. Y.; Sun, H. H, Gao, R.; Liu, H. L.; Wang, X., Xu, P.; Xun, L. Y. Nanoporous Gold-Based Microbial Biosensor for Direct Determination of Sulfide. Biosens. Bioelectron. 2017, 98, (2) Shariati-Rad, M.; Irandoust, M.; Jalilvand, F. Spectrophotometric Determination of Hydrogen Sulfide in Environmental Samples Using Sodium 1,2-naphthoquinone-4-sulfonate and Response Surface Methodology. Int. J. Environ. Sci. Technol. 2016, 13, (3) Ahmed, K. B. A.; Mariappan, M.; Veerappan, A. Nanosilver Cotton Swabs for Highly Sensitive and Selective Colorimetric Detection of Sulfide Ions at Nanomolar Level. Sens. Actuat. B-Chem. 2017, 244, (4) Liu J. M.; Wang, X. X.; Li, F. M.; Lin, L. P.; Cai, W. L.; Lin, X.; Zhang, L. H.; Li, Z. M.; Lin S. Q. A Colorimetric Probe for Online Analysis of Sulfide Based on the S-10

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