Leukocyte migration through confined spaces contributes to immunity and may involve trailing surface contraction via actomyosin, which can push nuclei through narrow gaps (Lämmermann et al. 2008; Wolf et al. 2013), but the forces so exerted are uncertain. This study investigates intracellular pressure during such migration, by counting blebs on neutrophils. Blebs are transient surface swellings, apparently raised by intracellular hydrostatic pressure generated by actomyosin contraction in the cell cortex (Charras et al. 2008). Neutrophils (from human blood obtained by finger puncture) migrated at 22-28 oC into micropipettes containing solutions of the chemoattractant N-formyl-L-methionyl-L-leucyl-L-phenylalanine (0.1 μM) at various constant hydrostatic pressures applied using a water manometer, while viewed via a water immersion objective. Values appear as mean ± S.E.M. As neutrophils migrated into tips of radius 1.37±0.03 μm (n=144), blebs sometimes formed on their leading and/or trailing surfaces, the former implying that pressure inside the front of the cell exceeded that in the micropipette (Charras et al. 2008). Leading blebs appeared at manometer pressures between 0-0.60 kPa, but not between 0.62-1.28 kPa (Figure 1). Micropipette pressures presumably corresponded closely to these manometer readings, because other experiments showed that flows along micropipettes containing single migrating neutrophils were much lower than those along open lumens. At 0.80-0.95 kPa, single neutrophils migrated inwards along micropipettes (of radius 2.97±0.29 μm) at 0.142±0.004 μm s-1 (n=5), apparently driving single flow marker beads (of radius 1.75±0.04 μm) suspended 2-30 μm ahead at 0.134±0.008 μm s-1 (n=5; P>0.10; paired t-test). In open micropipettes at 0 kPa, beads moved inwards too quickly for speed measurement, driven by capillarity. Together, the present observations indicate that some neutrophils generated intracellular pressures of up to at least (Charras et al. 2008) 0.50-0.60 kPa, while partly impeded during chemotaxis. This resembles previous evidence from servo null micropuncture that increased intracellular pressure may correlate with different forms of protrusion in amoebae (Yanai et al. 1996) and fibroblasts (Petrie et al. 2014) and is consistent with hypotheses that such hydrostatic pressure assists some cell movements.