Nanozyme sensor arrays for detecting versatile analytes from small molecules to proteins and cells

Transcription

1 Supporting Information Nanozyme sensor arrays for detecting versatile analytes from small molecules to proteins and cells Xiaoyu Wang, a, Li Qin, a, Min Zhou, a Zhangping Lou, a and Hui Wei a,b,* a Department of Biomedical Engineering, College of Engineering and Applied Sciences, Nanjing National Laboratory of Microstructures, Nanjing University, Nanjing, Jiangsu , China. b State Key Laboratory of Analytical Chemistry for Life Science and State Key Laboratory of Coordination Chemistry, School of Chemistry and Chemical Engineering, Collaborative Innovation Center of Chemistry for Life Sciences, Nanjing University, Nanjing, Jiangsu , China. * weihui@nju.edu.cn; Web: S1

2 Table of contents Supplementary Figure 1. Representative TEM images of Pt, Ru, and Ir nanoparticles. Supplementary Figure 2. (a) XPS spectrum of Pt 4f in Pt NPs. (b) XPS spectrum of Ru 3p in Ru NPs. (c) XPS spectrum of Ir 4f in Ir NPs. Supplementary Figure 3. Zeta potentials of Pt, Ru, and Ir nanozymes. Supplementary Figure 4. (a-c) Typical absorption spectra of different reaction systems of OPD, OPD + H 2O 2, OPD + nanozymes, and OPD + H 2O 2 + nanozymes after catalytic oxidation in ph 4.5, 0.2 M acetate buffer at 37 C. (d-f) Typical absorption spectra of 1 mm TMB, 2 mm OPD, and 1 mm ABTS after catalytic oxidation with 10 mm H 2O 2 in ph 4.5 acetate buffer at 37 C, in the presence of 10 μg/ml of Pt, Ru, and Ir nanozymes. Supplementary Figure 5. (a) Kinetic curves of A 450 for monitoring the catalytic reation of 2 mm OPD with various concentration of Pt nanozymes in the presence of 10 mm H 2O 2. (b) Kinetic curves of A 450 for monitoring the catalytic reation of 2 mm OPD with various concentration of H 2O 2 in the presence of 5 μg/ml Pt nanozymes. (c, d) ph and temperaturedependent peroxidase mimicking activities of Pt nanozymes. Supplementary Figure 6. The steady-state kinetics assays of Pt nanozyme. Supplementary Figure 7. The steady-state kinetics assays of Ru nanozyme. Supplementary Figure 8. The steady-state kinetics assays of Ir nanozyme. Supplementary Figure 9. Chemical structure of six biothiols studied in this work. Supplementary Figure 10. Typical absorption spectra for monitoring the catalytic oxidation of OPD in the presence of Pt nanozyme with various concentration of biothiols. Supplementary Figure 11. Typical absorption spectra for monitoring the catalytic oxidation of OPD in the presence of Ru nanozyme with various concentration of biothiols. Supplementary Figure 12. Typical absorption spectra for monitoring the catalytic oxidation of OPD in the presence of Ir nanozyme with various concentration of biothiols. Supplementary Figure 13. Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 5 μm of biothiols. Supplementary Figure 14. (a) Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 10 μm of biothiols. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 10 μm of biothiols. Supplementary Figure 15. (a) Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 100 μm of biothiols. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 100 μm of biothiols. Supplementary Figure 16. Canonical variable 1 of the sensor arrays plotted versus different concentrations of Cys. Supplementary Figure 17. (a) Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 100 μm of biothiols in the presence of 1% FBS without pretreament. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 100 μm of biothiols in the presence of 1% FBS. Supplementary Figure 18. (a) Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 20 μm of biothiols in the presence of 1% FBS. The sample pretreatment procedures were conducted by a centrifugal filter unit. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 20 μm of biothiols in the presence of 1% FBS. S2

3 Supplementary Figure 19. Kinetic curves of A 450 for monitoring the catalytic oxidation of 2 mm OPD with 40 mm H 2O 2 in the presence of 1 μg/ml of (a) Pt, (b) Ru, and 0.25 μg/ml of (c) Ir nanozymes with various proteins. Supplementary Figure 20. Dependence of colorimetric response (A/A 0) in the presence of Pt, Ru, and Ir nanozymes on the concentration of (a, b, c) Cyt and (d, e, f) HSA. Supplementary Figure 21. Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards 20 nm of proteins. Supplementary Figure 22. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards 50 nm of proteins. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 50 nm of proteins. Supplementary Figure 23. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards 100 nm of proteins. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 100 nm of proteins. Supplementary Figure 24. 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 2 μm of proteins. Supplementary Figure 25. Canonical variable 1 of the sensor array plotted versus different concentrations of HSA. Supplementary Figure 26. Concentration-dependent response of the as-prepared nanozyme sensor arrays. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards various concentrations of Cyt. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against six different concentrations of Cyt. (c) Canonical variable 1 of the sensor array plotted versus different concentrations of Cyt. Supplementary Figure 27. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards 100 nm of proteins in the presence of human urine. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 100 nm of proteins in the presence of human urine. Supplementary Figure 28. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards the mixture of HSA and GSH. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against the mixture of HSA and GSH. Supplementary Table 1. Kinetics parameters of Pt, Ru, and Ir nanozymes as well as HRP. Supplementary Table 2. Identification of 30 unknown biothiol samples. Supplementary Table 3. Basic physical properties of protein analytes. Supplementary Table 4. Kinetics parameters of Ir, Ir-GSH, Ir-CYS, and Ir-HSA. Supplementary Table 5. Identification of 45 unknown protein samples. S3

4 Supplementary Figure 1. Representative TEM images of (a, b) Pt nanoparticles, (c, d) Ru nanoparticles, and (e, f) Ir nanoparticles. S4

5 Supplementary Figure 2. (a) XPS spectrum of Pt 4f in Pt NPs. (b) XPS spectrum of Ru 3p in Ru NPs. (c) XPS spectrum of Ir 4f in Ir NPs. S5

6 Supplementary Figure 3. Zeta potentials of Pt, Ru, and Ir nanozymes. Each error bar shows the standard deviation of three independent measurements. We measured the zeta potentials of Pt, Ru, and Ir nanoparticles because the surface charges would affect the interaction between nanozymes and analytes. As shown in Figure S2, all the three nanozymes showed negative zeta potentials. S6

7 Supplementary Figure 4. (a-c) Typical absorption spectra of different reaction systems of OPD, OPD + H 2O 2, OPD + nanozymes, and OPD + H 2O 2 + nanozymes after catalytic oxidation in ph 4.5, 0.2 M acetate buffer at 37 C. (d-f) Typical absorption spectra of 1 mm TMB, 2 mm OPD, and 1 mm ABTS after catalytic oxidation with 10 mm H 2O 2 in ph 4.5 acetate buffer at 37 C, in the presence of 10 μg/ml of Pt, Ru, and Ir nanozymes. S7

8 Supplementary Figure 5. (a) Kinetic curves of A 450 for monitoring the catalytic reation of 2 mm OPD with various concentration of Pt nanozymes in the presence of 10 mm H 2O 2. (b) Kinetic curves of A450 for monitoring the catalytic reation of 2 mm OPD with various concentration of H 2O 2 in the presence of 5 μg/ml Pt nanozymes. (c, d) ph and temperaturedependent peroxidase mimicking activities of Pt nanozymes. Each error bar shows the standard deviation of three independent measurements. As shown in Figure S5, the Pt nanozymes exhibited H 2O 2 concentrations, catalyst concentrations, and ph-dependent peroxidase mimicking activity, which is similar with natural peroxidases. However, the catalytic activity of Pt nanozymes slightly increased as the temperature increased, demonstrating the higher thermal stability of Pt nanozymes than natural peroxidases. S8

9 Supplementary Figure 6. The steady-state kinetics assays of Pt nanozyme. Plots of the velocity of the reaction versus different concentrations of H 2O 2 (a, 0.5 mm TMB) or TMB (b, 50 mm H 2O 2). Double reciprocal plots of the velocity versus varying concentration of (c) H 2O 2 and (d) TMB. Each error bar shows the standard deviation of three independent measurements. S9

10 Supplementary Figure 7. The steady-state kinetics assays of Ru nanozyme. Plots of the velocity of the reaction versus different concentrations of H 2O 2 (a, 0.5 mm TMB) or TMB (b, 50 mm H 2O 2). Double reciprocal plots of the velocity versus varying concentration of (c) H 2O 2 and (d) TMB. Each error bar shows the standard deviation of three independent measurements. S10

11 Supplementary Figure 8. The steady-state kinetics assays of Ir nanozyme. Plots of the velocity of the reaction versus different concentrations of H 2O 2 (a, 0.5 mm TMB) or TMB (b, 50 mm H 2O 2). Double reciprocal plots of the velocity versus varying concentration of (c) H 2O 2 and (d) TMB. Each error bar shows the standard deviation of three independent measurements. S11

12 Supplementary Figure 9. Chemical structure of six biothiols studied in this work. S12

13 Supplementary Figure 10. Typical absorption spectra for monitoring the catalytic oxidation of OPD in the presence of Pt nanozyme with various concentration of biothiols. (a) Cys, (b) DTT, (c) MA, (d) ME, and (e) MS. S13

14 Supplementary Figure 11. Typical absorption spectra for monitoring the catalytic oxidation of OPD in the presence of Ru nanozyme with various concentration of biothiols. (a) Cys, (b) DTT, (c) MA, (d) ME, and (e) MS. S14

15 Supplementary Figure 12. Typical absorption spectra for monitoring the catalytic oxidation of OPD in the presence of Ir nanozyme with various concentration of biothiols. (a) Cys, (b) DTT, (c) MA, (d) ME, and (e) MS. S15

16 Supplementary Figure 13. Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 5 μm of biothiols. Each error bar shows the standard deviation of five measurements. A and A 0 were the absorption of oxopd in the presence and absence of biothiols in the nanozyme reaction systems, respectively. S16

17 Supplementary Figure 14. (a) Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 10 μm of biothiols. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 10 μm of biothiols. The canonical scores were calculated by LDA for the identification of six biothiols. S17

18 Supplementary Figure 15. (a) Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 100 μm of biothiols. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 100 μm of biothiols. The canonical scores were calculated by LDA for the identification of six biothiols. S18

19 Supplementary Figure 16. Canonical variable 1 of the sensor arrays plotted versus different concentrations of Cys. S19

20 Supplementary Figure 17. (a) Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 100 μm of biothiols in the presence of 1% FBS without pretreament. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 100 μm of biothiols in the presence of 1% FBS. The canonical scores were calculated by LDA for the identification of six biothiols and serum. S20

21 Supplementary Figure 18. (a) Colorimetric response patterns ((A 0-A)/A 0) of nanozyme sensor arrays towards 20 μm of biothiols in the presence of 1% FBS. The sample pretreatment procedures were conducted by a centrifugal filter unit. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 20 μm of biothiols in the presence of 1% FBS. The canonical scores were calculated by LDA for the identification of six biothiols and serum. S21

22 Supplementary Figure 19. Kinetic curves of A 450 for monitoring the catalytic oxidation of 2 mm OPD with 40 mm H 2O 2 in the presence of 1 μg/ml of (a) Pt, (b) Ru, and 0.25 μg/ml of (c) Ir nanozymes with various proteins. Bla: blank; 1: Lys; 2: BSA; 3: Tfn; 4: Amy; 5: HSA; 6: Try; 7: Cyt; 8: GOx; 9: Hem. S22

23 Supplementary Figure 20. Dependence of colorimetric response (A/A 0) in the presence of Pt, Ru, and Ir nanozymes on the concentration of (a, b, c) Cyt and (d, e, f) HSA. Each error bar shows the standard deviation of three independent measurements. S23

24 Supplementary Figure 21. Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards 20 nm of proteins. Each error bar shows the standard deviation of five measurements. A and A 0 were the absorption of oxopd in the presence and absence of proteins in the nanozyme reaction systems, respectively. S24

25 Supplementary Figure 22. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards 50 nm of proteins. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 50 nm of proteins. The canonical scores were calculated by LDA for the identification of nine proteins. S25

26 Supplementary Figure 23. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards 100 nm of proteins. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 100 nm of proteins. The canonical scores were calculated by LDA for the identification of nine proteins. S26

27 Supplementary Figure 24. 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 2 μm of proteins. The canonical scores were calculated by LDA for the identification of nine proteins. S27

28 Supplementary Figure 25. Canonical variable 1 of the sensor array plotted versus different concentrations of HSA. S28

29 Supplementary Figure 26. Concentration-dependent response of the as-prepared nanozyme sensor arrays. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards various concentrations of Cyt. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against six different concentrations of Cyt. The canonical scores were calculated by LDA for the identification of nine proteins. (c) Canonical variable 1 of the sensor array plotted versus different concentrations of Cyt. S29

30 Supplementary Figure 27. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards 100 nm of proteins in the presence of human urine. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against 100 nm of proteins in the presence of human urine. The canonical scores were calculated by LDA for the identification of nine proteins and urine. S30

31 Supplementary Figure 28. (a) Colorimetric response patterns (A/A 0) of nanozyme sensor arrays towards the mixtures of 200 nm HSA and 200 μm GSH at different ratios. Each error bar shows the standard deviation of five measurements. (b) 2D canonical score plot for the first two factors of colorimetric response patterns obtained against the mixtures of HSA and GSH. The canonical scores were calculated by LDA for the identification of nine proteins. As we know, cell lysate consists of biothiols, proteins, etc. Before cells discrimination assays, we examined the ability of sensor arrays to differentiate the mixture of GSH and HSA. As shown in Figure S28, these mixtures with different ratios were clustered into different groups with no errors or misclassifications, suggesting that the sensor arrays could discriminate the mixture of GSH and HSA. The result above demonstrated the possibility for discrimination of cells using their lysate solutions. S31

32 Supplementary Table 1. Kinetics parameters of Pt, Ru, and Ir nanozymes as well as HRP. Catalyst Substrate K m (mm) V max (Ms -1 ) Ref. Pt TMB This work Pt H 2 O This work Ir TMB This work Ir H 2 O This work Ru TMB This work Ru H 2 O This work HRP TMB [1] HRP H 2 O [1] S32

33 Supplementary Table 2. Identification of 30 unknown biothiol samples. No. Concentration Source Data Identification Verification 1 5 μm DTT DTT 2 5 μm ME ME 3 5 μm MA MA 4 5 μm MS MS 5 5 μm Cys Cys 6 5 μm GSH GSH 7 10 μm DTT DTT 8 10 μm ME ME 9 10 μm MA MA μm MS MS μm Cys Cys μm GSH GSH μm DTT DTT μm ME ME μm MA MA μm MS MS μm Cys Cys μm GSH GSH μm DTT DTT μm DTT ME μm MA MA μm MS MS S33

34 23 50 μm Cys Cys μm GSH GSH μm in serum DTT DTT μm in serum ME ME μm in serum MA MA μm in serum GSH MS μm in serum Cys Cys μm in serum GSH GSH The samples in red were mis-identified. S34

35 Supplementary Table 3. Basic physical properties of protein analytes. Protein MW (kda) pi α-amylase (Amy) Bovine serum albumin (BSA) Trypsin (Try) Cytochrome c (Cyt) Transferrin (Tfn) Human serum albumin (HSA) Hemoglobin (Hem) Lysozyme (Lys) Glucose oxidase (GOx) Nine kinds of proteins with different characteristics including molecular weight and isoelectric point (pi) were chosen to study the discrimination ability of the nanozyme sensor arrays. S35

36 Supplementary Table 4. Kinetics parameters of Ir, Ir-GSH, Ir-CYS, and Ir-HSA. Catalyst Substrate K m (mm) V max (Ms -1 ) Ref. Ir TMB This work Ir H 2 O This work Ir-GSH TMB This work Ir-GSH H 2 O This work Ir-Cys TMB This work Ir-Cys H 2 O This work Ir-HSA TMB This work Ir-HSA H 2 O This work S36

37 Supplementary Table 5. Identification of 45 unknown protein samples. No. Concentration Source Data Identification Verification 1 20 nm Lys Lys 2 20 nm BSA BSA 3 20 nm Tra Tra 4 20 nm Amy Amy 5 20 nm HSA HSA 6 20 nm Tfn Try 7 20 nm Cyt Cyt 8 20 nm GOx GOx 9 20 nm Hem Hem nm Lys Lys nm BSA BSA nm Tfn Tra nm Amy Amy nm HSA HSA nm Try Try nm Cyt Cyt nm GOx GOx nm Hem Hem nm Lys Lys nm BSA HSA nm Tfn Tra nm Amy Amy nm HSA HSA nm Try Try nm Cyt Cyt nm GOx GOx nm Hem Hem nm Lys Lys nm BSA HSA nm Tfn Tra nm Amy Amy nm HSA HSA nm Try Try nm Cyt Cyt nm GOx GOx S37

38 nm Hem Hem 37 1 μm Lys Lys 38 1 μm BSA Cyt 39 1 μm Tfn Tra 40 1 μm Amy Amy 41 1 μm HSA HSA 42 1 μm Try Try 43 1 μm Cyt Cyt 44 1 μm GOx GOx 45 1 μm Hem Hem The samples in red were mis-identified. S38