Microkinetics of full-length L-selectin-, LΔcyto- and L358stop-expressing 300.19 cells interacting with GlyCAM-1 100 sites/μm2 at 0.5, 1, and 1.75 dyn/cm2
. | Wall shear stressa . | . | . | . | . | . | . | ||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
. | 1.75 dyn/cm2 . | . | 1 dyn/cm2 . | . | 0.5 dyn/cmd . | . | . | ||||||
| L-selectin variant | Full-length | LΔcyto | Full-length | LΔcyto | Full-length | LΔcyto | L358stop5 | ||||||
| Rolling fraction (%) | 40 | 16 | 54 | 37 | 0 | 0 | 0 | ||||||
| Tethering fraction (%) | ND | ND | ND | ND | 32 | 26 | 7 | ||||||
| Mean pause duration (ms) | 20 ± 1 (n = 104) | 12 ± 1 (n = 88) | 31 ± 1 (n = 393) | 20 ± 1 (n = 286) | 61 ± 6 (n = 76) | 47 ± 4 (n = 65) | 23 ± 2 (n = 39) | ||||||
| koff (s−1) | 64.4 ± 5.4 | 138.7 ± 4.7 | 34.1 ± 2.3 | 61.1 ± 5.4 | 16.2 ± 1.6 | 20.9 ± 1.5 | 44.6 ± 3.8 | ||||||
| Mean step distance (μm)b | 4 ± 0.1 | 10.2 ± 0.3 | 2.5 ± 0.1 | 4.6 ± 0.1 | NAc | N.A. | N.A. | ||||||
. | Wall shear stressa . | . | . | . | . | . | . | ||||||
|---|---|---|---|---|---|---|---|---|---|---|---|---|---|
. | 1.75 dyn/cm2 . | . | 1 dyn/cm2 . | . | 0.5 dyn/cmd . | . | . | ||||||
| L-selectin variant | Full-length | LΔcyto | Full-length | LΔcyto | Full-length | LΔcyto | L358stop5 | ||||||
| Rolling fraction (%) | 40 | 16 | 54 | 37 | 0 | 0 | 0 | ||||||
| Tethering fraction (%) | ND | ND | ND | ND | 32 | 26 | 7 | ||||||
| Mean pause duration (ms) | 20 ± 1 (n = 104) | 12 ± 1 (n = 88) | 31 ± 1 (n = 393) | 20 ± 1 (n = 286) | 61 ± 6 (n = 76) | 47 ± 4 (n = 65) | 23 ± 2 (n = 39) | ||||||
| koff (s−1) | 64.4 ± 5.4 | 138.7 ± 4.7 | 34.1 ± 2.3 | 61.1 ± 5.4 | 16.2 ± 1.6 | 20.9 ± 1.5 | 44.6 ± 3.8 | ||||||
| Mean step distance (μm)b | 4 ± 0.1 | 10.2 ± 0.3 | 2.5 ± 0.1 | 4.6 ± 0.1 | NAc | N.A. | N.A. | ||||||
All measurements were performed on cell motion segments of at least 20 cells recorded at 500 frames/s.
Step distance was defined as the distance traveled by a rolling cell between successive tethers.
Not applicable, as no rolling could be detected at low shear stress.
The 358stop mutant did not support any cell tethering to GlyCAM-1 at shear stresses >1 dyn/cm2.