Volume 111, Issue 3, Pages 577-588 (August 2016) The Effect of Hematocrit on Platelet Adhesion: Experiments and Simulations  Andrew P. Spann, James E. Campbell, Sean R. Fitzgibbon, Armando Rodriguez, Andrew P. Cap, Lorne H. Blackbourne, Eric S.G. Shaqfeh  Biophysical Journal  Volume 111, Issue 3, Pages 577-588 (August 2016) DOI: 10.1016/j.bpj.2016.06.024 Copyright © 2016 Biophysical Society Terms and Conditions

Figure 1 Parameters in receptor bond model. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 2 Schematic of simulation initialization, example shown for 25% hematocrit. (a) RBCs are introduced to fill a gridlike pattern with a uniform probability distribution. (b) The simulation is run with just RBCs to equilibration. (c) Platelets are inserted inside the channel region containing RBCs. (d) The simulation is run with bond formation disabled to allow margination. The state is then labeled time 0 for bond formation simulations. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 3 The microfluidic channel used in the experimental apparatus is shown here. Most notably, the image capture region lies ∼165.5-mm downstream from the input well. Images were processed showing platelets adhering to a 300 × 300 pixel (190 × 190 μm) region of interest. The channel is 350-μm wide in the region of data capture. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 4 The platelet surface area probability distribution at each hematocrit after platelets have marginated but before platelet bonding is enabled according to Fig. 2 d. The capillary number is Ca = 2 (corresponding to a characteristic shear rate of γ˙=4000s−1). To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 5 (a) Images of the marginated state comparable to Fig. 4 with capillary number is Ca = 0.5 (corresponding to a characteristic shear rate of γ˙=1000s−1). (b) The RBC surface area probability distribution, normalized so that the total area under the curve (including the middle channel region not shown) is equal for 15 and 30% hematocrits. In contrast with Fig. 4, this distribution has been symmetrized so that it measures platelet distance from either the top or bottom wall. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 6 Platelet trajectories showing the center of mass of all 30 platelets in the simulation for a run at 30% hematocrit. Full margination has occurred at the time when the simulation begins allowing bonds to form with the wall. The capillary number is Ca = 2. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 7 A platelet (green) forms a bond, ratchets downward from the torque, and then releases and continues flowing. Two dimensionless time elapses between each of the side-view screenshots. Additionally, a view from the bottom is shown for the beginning, middle, and end of this process. RBCs are colored red, platelets are white, and bonds are represented as thin blue lines. Hematocrit is 30%. The capillary number is Ca = 2. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 8 Inclination angle versus time shown for selected platelets at different heights of the channel at 30% hematocrit and capillary number Ca = 2. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 9 (a) Average platelet-exposed surface area per time within the bonding radius in the simulations at capillary number Ca = 2. (b) The graph normalized by the hematocrit 30% simulation, demonstrating the variability with the choice of observation time. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 10 Images of platelet adhesion to bottom wall of channel at 40% hematocrit from experimental snapshots, full data for which is presented in Table 2 and Figs. 11 and 12. The camera taking these images is positioned below the flow chamber. Images near the wall are taken every 30 s. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 11 Experimental data for adhered platelet area versus time. Each graph represents two runs from n = 5 different volunteers’ blood samples. The slopes of these lines are averaged and used for Fig. 12. These slopes, used as a measurement of adhesion rates, are presented in numerical form in Table 2. To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions

Figure 12 Comparison of simulation and experimental adhesion activity, normalized against the adhesion activity at 30% hematocrit. Experimental platelet adhesion is defined, in a graph, as a slope of standard area per time for the first 4 min, and is taken from two runs of n = 5 volunteers’ blood samples. Simulation platelet activity is defined as the average exposed platelet surface area within bonding distance per time after enabling bonding for 200 dimensionless time normalized to the area of a whole platelet. Each point was averaged over two simulations that started 15 dimensionless time apart from fully marginated equilibrium (i.e., a long enough time that we expect no autocorrelation in platelet velocity from the previous initial condition). To see this figure in color, go online. Biophysical Journal 2016 111, 577-588DOI: (10.1016/j.bpj.2016.06.024) Copyright © 2016 Biophysical Society Terms and Conditions