GSK3 and Lyn inhibition impacts capillary traversal of reticulocytes and red blood cells. (A) Percentage recovery of reticulocytes treated with the respective reagent after microsphiltration. After treatment, the cells were added to an untreated RBC suspension in a 5:95 ratio and the mixture then subjected to microsphiltration. Anti-GPA antibody was used as a positive control for negative effects on cell deformability. Bafetinib (3 µM) was used as a Lyn inhibitor and CHIR-98014 (10 nM) was used as a GSK3 inhibitor. Error bars correspond to the standard deviation (n = 3). All comparisons were performed with a paired 2-tailed Student t test. The P values for each comparison are shown below the bar graph. *P < .05, **P < 0.01. (B) Image of the microfluidic PDMS biochip used for the capillary traversal experiment. The magnified image shows a section of the microcapillary channels, through which reticulocytes and red blood cells were subjected to successive deformations. Frame sequences were obtained through the use of a high-speed camera and then subjected to image analysis for automated cell tracking with the use of TrackMate.⁵¹  Scale bars, 50 μm. (C) Percentage microcapillary traversal velocity of reticulocytes and red blood cells treated with the respective reagents. The cells were treated for a minimum period of 1 hour and subjected to passage through the microfluidic biochip. The graph shows the average cell microcapillary traversal velocity of each treated sample as a percentage of the respective DMSO control sample traversal velocity. All comparisons were performed with a 1-sample Student t test. The error bars correspond to the standard deviation (n = 3). The P values for each comparison are shown below the dot plot. *P < .05, **P < .01. A minimum of 1000 cell tracks were analyzed per sample. n.s.s., not statistically significant.