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Tianhui Maria Ma, J. Scott VanEpps, Michael J. Solomon 

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1 Structure, Mechanics, and Instability of Fibrin Clot Infected with Staphylococcus epidermidis 
Tianhui Maria Ma, J. Scott VanEpps, Michael J. Solomon  Biophysical Journal  Volume 113, Issue 9, Pages (November 2017) DOI: /j.bpj Copyright © 2017 Biophysical Society Terms and Conditions

2 Figure 1 S. epidermidis alters fibrin elasticity. Shown here is the elastic modulus, G′, of a pure fibrin clot (0.5 mg/mL) and infected fibrin clots (0.5 mg/mL fibrin with 1 × 109, 2 × 109, 4 × 109, and 6 × 109 cells/mL) with variation with time. Error bars denote SD with n = 3. To see this figure in color, go online. Biophysical Journal  , DOI: ( /j.bpj ) Copyright © 2017 Biophysical Society Terms and Conditions

3 Figure 2 Steady-state microstructure of a fibrin clot and an infected fibrin clot. (a) Given here are axial projections of a pure fibrin clot (0.5 mg/mL) and (b) an infected clot (with 4 × 109 cells/mL in 0.5 mg/mL fibrin) showing the existence of large pores. Fibrin is red and bacterial cells are blue. (c) Shown here is a high-resolution image from the identified region of interest in (a). (d) Shown here is a high-resolution image from the region of interest in (b). The scale bar represents 10 μm in (a–d). (e) Shown here is mesh size characterization of a steady-state pure fibrin clot and an infected fibrin clot. Error bars denote standard deviations of all the image stacks in all five replicates (see Materials and Methods). To see this figure in color, go online. Biophysical Journal  , DOI: ( /j.bpj ) Copyright © 2017 Biophysical Society Terms and Conditions

4 Figure 3 MPT microrheology in fibrin and infected clots. (a) Given here is one image stack from a set of time-series images of MPT in a pure fibrin and in an infected clot. Fibrin is red; bacteria are blue; probes bound to fibrin are yellow; freely diffusing probes are green (see Materials and Methods). The zoomed-in plots show the trajectories of the bound probes in pure fibrin and infected fibrin, respectively. The fluctuation of probes bound to fibrin is inversely proportional to the elastic modulus. (b) Shown here is a comparison between the steady-state elastic modulus G′ obtained from bulk rheology and G′ calculated from MPT (see Materials and Methods). Error bars denote SD with n = 3. To see this figure in color, go online. Biophysical Journal  , DOI: ( /j.bpj ) Copyright © 2017 Biophysical Society Terms and Conditions

5 Figure 4 Infected fibrin clot formation and rupture. (a) Axial projections of an infected clot (4 × 109 cells/mL in 0.5 mg/mL fibrin) rendered from 20 to 30 μm above the coverslip are given at 1-μm intervals at different time points. Fibrin is stained red and bacteria are stained blue. The scale bar represents 10 μm in all images. (b and c) Given here are renderings of the 3D image volume of (b) a pure fibrin clot and (c) the region in the infected clot showed in (a), captured 150 min after adding thrombin. The width and height of volumes in (b) and (c) are 127 and 50 μm, respectively. To see this figure in color, go online. Biophysical Journal  , DOI: ( /j.bpj ) Copyright © 2017 Biophysical Society Terms and Conditions

6 Figure 5 Microstructural change during the infected fibrin clotting process. Given here is the variance of (a) the normalized fiber area, (b) the bacteria fibrin overlap percentage, and (c) mesh sizes during the infected fibrin and the pure fibrin clotting process. Error bars denote SD of n = 5. To see this figure in color, go online. Biophysical Journal  , DOI: ( /j.bpj ) Copyright © 2017 Biophysical Society Terms and Conditions

7 Figure 6 Multiscale imaging of infected fibrin clots. Given here are confocal microscopy 3D volumes of (a) a pure fibrin clot and (b) an infected clot, imaged 5 h. Fibrin is stained red; cells are stained blue. The width and height values of the volumes are 254 and 10 μm, respectively. (c) Given here are triplicate time-lapse photos of infected fibrin clots with 4 × 109 cells/mL (top three rows) compared to a pure fibrin clot (bottom row) in chambered cover glass wells. Scale bar in the first photo = 3 mm. To see this figure in color, go online. Biophysical Journal  , DOI: ( /j.bpj ) Copyright © 2017 Biophysical Society Terms and Conditions


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