Additional file 1 of Quantifying intracellular trafficking of silica-coated magnetic nanoparticles in live single cells by site-specific direct stochastic optical reconstruction microscopy
Creators: name: Chakkarapani, Suresh Kumar
name: Shin, Tae Hwan
name: Lee, Seungah
name: Park, Kyung-Soo
name: Lee, Gwang
name: Kang, Seong Ho
Publisher: name: figshare
Year: 2021
Subjects: subject: Biophysics
subject: Space Science
subject: Cell Biology
subject: Genetics
subject: FOS: Biological sciences
subject: Biotechnology
subject: 39999 Chemical Sciences not elsewhere classified
subject: FOS: Chemical sciences
subject: 69999 Biological Sciences not elsewhere classified
subject: Developmental Biology
subject: Cancer
subject: Hematology

description: Additional file 1: Table S1. Ingenuity Pathway Analysis-based profiles of DNA repair-related genes in HEK293 cells treated with MNPs@SiO2(RITC). Table S2. ICP-QMS and AAS data for Co and Fe quantification in 3 × 106 cells of HEK293, NIH3T3 and RAW 264.7 at 0.01 µg/mL MNPs@SiO2(RITC). Table S3. Quantitative real time PCR primer sequences for genes encoding DNA repair related genes. Fig. S1. Characterization of mouse primary liver cells. a Morphological analysis of isolated Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells. Scale bar = 20 μm. Cell specific marker protein expression analysis for b F4/80 using flow cytometry, c albumin, and CD31 using immunoblotting. β-actin was used as the internal control. Fig. S2. (a) Physical layout and (b) schematic representation of the lab-made dSTORM setup. Following acronyms were used; L, laser; M, mirror; DCM, dichroic mirror; MS, mechanical shutter; NP, Nomarski prism; EM-CCD, electron-multiplying cooled charge-coupled device. Fig. S3. Plot of drift correction on the x- and y-axes with respect to frame illustrated with a fiducial marker. Images of raw, before drift correction and after drift correction. Fig. S4. a. The photo-switching rate constant koff was obtained by irradiation of 532 nm laser and monitoring the decrease in the fluorescence intensity over time. b. The photo-switching rate constant kon was obtained by simultaneous irradiation of the sample with 405 nm and 532 nm (200 mW/cm2) and monitoring the increase in the fluorescence intensity with time. Both koff and kon shows a linear dependence with illuminated intensities. Error bars represent the standard deviations of five measurements. c. Photoinduced restoration of RITC to fluorescence state with 405 nm activation laser. Fig. S5. Wide-field and dSTORM images of HEK293, NIH3T3, and RAW 264.7 cells incubated for 1 h with three different concentrations a. 0.1 µg/mL, b. 0.01 µg/mL, and c. 0.003 µg/mL of 50-nm MNPs@SiO2(RITC). Fig. S6. Wide-field and dSTORM images of HEK293, NIH3T3, and RAW 264.7 cells incubated for 3 h with three different concentrations a. 0.1 µg/mL, b. 0.01 µg/mL, and c. 0.003 µg/mL of 50-nm MNPs@SiO2(RITC). Fig. S7. The wide-field TIRF images of Fig. 1b. A single emitter is magnified and the consecutive images show the photo-switching events in the same spot and the corresponding Gaussian fitting for localizing the centroid of the individual NPs within diffraction-limited images. Fig. S8. The wide-field TIRF images of the RAW 264.7 cell incubated with 0.01 µg/mL of 50-nm MNPs@SiO2(RITC) for 5 h with a. serum free DMEM and b. PBS buffer. Fig. S9. Evaluation of cytotoxicity in MNPs@SiO2(RITC)-treated cells. MTS assay of MNPs@SiO2(RITC)-treated HEK293, NIH3T3, and RAW 264.7 cells. Cells were treated with 0.1 µg/mL, 0.3 µg/mL and 1.0 µg/mL MNPs@SiO2(RITC) for 12 h, followed by cell viability assessment using MTS assay. Fig. S10. Sixty-nine DNA-repair-related genes were constructed algorithmically using Ingenuity Pathway Analysis (IPA). Red and green in the genetic network indicate up- and down-regulated genes, respectively, in HEK293 cells treated with 1.0 mg/mL MNPs@SiO2(RITC), when compared with non-treated controls for 12 h. The data set of differentially expressed genes obtained from microarray data with a greater than threefold change is shown. Fig. S11. Sixty-nine DNA repair related genes showing significantly altered expression by MNPs@SiO2(RITC) on microarray analysis. DNA repair-related genes were constructed algorithmically using IPA. Red and green in the genetic network indicate up- and down-regulated genes, respectively, in HEK293 cells treated with 0.1 mg/mL MNPs@SiO2(RITC), when compared with non-treated controls for 12 h. The data set of differentially expressed genes obtained from microarray data with &gt; threefold change is shown. Thirty-six genes were significantly changed. Fig. S12. DIC, wide-field, and dSTORM images of HEK293, NIH3T3, and RAW 264.7 cell respectively. Images were acquired after incubation of 50-nm MNPs@SiO2(RITC) for 3 h with each cell. Merged images of DIC and dSTORM for site-specific analysis of the internalized NPs. Fig. S13. DIC, wide-field, and dSTORM images of HEK293, NIH3T3, and RAW 264.7 cell respectively. Images were acquired after incubation of 50-nm MNPs@SiO2(RITC) for 5 h with each cell. Merged images of DIC and dSTORM for site-specific analysis of the internalized NPs. Fig. S14. DIC, wide-field, and dSTORM images of HEK293, NIH3T3, and RAW 264.7 cell respectively. Images were acquired after incubation of 50-nm MNPs@SiO2(RITC) for 12 h with each cell. Merged images of DIC and dSTORM for site-specific analysis of the internalized NPs. Fig. S15. DIC and super-resolution images of Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells incubated for 5 h with 2 µg/mL concentration of 50-nm MNPs@SiO2(RITC). Fig. S16. DIC and super-resolution images of Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells incubated for 24 h with 2 µg/mL concentration of 50-nm MNPs@SiO2(RITC). Fig. S17. DIC and super-resolution images of Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells incubated for 5 h with 2 µg/mL concentration of 10-nm SiO2 NPs. Fig. S18. DIC and super-resolution images of Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells incubated for 5 h with 2 µg/mL concentration of 50-nm SiO2 NPs. Fig. S19. DIC and super-resolution images of Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells incubated for 5 h with 2 µg/mL concentration of 200-nm SiO2 NPs. Fig. S20. DIC and super-resolution images of Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells incubated for 24 h with 2 µg/mL concentration of 10-nm SiO2 NPs. Fig. S21. DIC and super-resolution images of Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells incubated for 24 h with 2 µg/mL concentration of 50-nm SiO2 NPs. Fig. S22. DIC and super-resolution images of Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells incubated for 24 h with 2 µg/mL concentration of 200-nm SiO2 NPs. Fig. S23. Evaluation of cytotoxicity in SiO2 NPs-treated cells. MTS assay of a. 10-nm, b. 50-nm, and c. 200-nm SiO2 NPs-treated Kupffer cells, hepatocytes, and liver sinusoidal endothelial cells. Cells were treated with 0.1 mg/mL, 0.3 mg/mL and 1.0 mg/mL SiO2 NPs for 12 h, followed by cell viability assessment using MTS assay.