Quantitative dynamic footprinting microscopy reveals mechanisms of neutrophil rolling

Abstract

Adaptations to total internal reflection microscopy permit visualization of the 'footprint' of rolling cells. Applied to neutrophils rolling in whole blood at physiological levels of shear stress, this approach reveals previously unappreciated features of rolling cell biology. We introduce quantitative dynamic footprinting microscopy to resolve neutrophil rolling on P-selectin. We observed that the footprint of a rolling neutrophil was fourfold larger than previously thought, and that P-selectin–PSGL-1 bonds were relaxed at the leading edge of the rolling cell, compressed under the cell center, and stretched at the trailing edge. Each rolling neutrophil formed three to four long tethers that extended up to 16 μm behind the rolling cell.