Abstract
Neutrophil migration in tissues critically regulates the human immune response and can either play a protective role in host defense or cause health problems. Microfluidic chips are increasingly applied to study neutrophil migration, attributing to their advantages of low reagent consumption, stable chemical gradients, visualized cell chemotaxis monitoring, and quantification. Most chemotaxis chips suffered from low throughput and fussy cell separation operations. We here reported a novel and simple "sample in and chemotaxis out" method for rapid neutrophils isolation from a small amount of whole blood based on a simplified magnetic method, followed by a chemotaxis assay on a microfluidic chip (SC2 chip) consisting of six cell migration units and six-cell arrangement areas. The advantages of the "sample in and chemotaxis out" method included: less reagent consumption (10 μL of blood + 1 μL of magnetic beads + 1 μL of lysis buffer); less time (5 min of cell isolation + 15 min of chemotaxis testing); no ultracentrifugation; more convenient; higher efficiency; high throughput. We have successfully validated the approach by measuring neutrophil chemotaxis to frequently-used chemoattractant (i.e., fMLP). The effects of D-glucose and mannitol on neutrophil chemotaxis were also analyzed. In addition, we demonstrated the effectiveness of this approach for testing clinical samples from diabetes mellitus type 2 (T2DM) patients. We found neutrophils' migration speed was higher in the "well-control" T2DM than in the "poor-control" group. Pearson coefficient analysis further showed that the migration speed of T2DM was negatively correlated with physiological indicators, such as HbA1c (-0.44), triglyceride (-0.36), C-reactive protein (-0.28), and total cholesterol (-0.28). We are very confident that the developed "sample in and chemotaxis out" method was hoped to be an attractive model for analyzing the chemotaxis of healthy and disease-associated neutrophils.