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Prepublished online as a Blood First Edition Paper on April 17, 2002; DOI 10.1182/blood-2001-11-0060.
HEMOSTASIS, THROMBOSIS, AND VASCULAR BIOLOGY
From the Department of Bioengineering, Rice University;
and the Division of Thrombosis Research, Department of Medicine, Baylor
College of Medicine, Houston, TX.
Ultralarge von Willebrand factor (ULVWF) multimers have
been implicated in the pathogenesis of the catastrophic
microangiopathic disorder, thrombotic thrombocytopenic purpura.
Spontaneous ULVWF binding to platelets has been ascribed to increased
avidity due to the greatly increased number of binding sites for
platelets (the A1 domain) per molecule. To address the mechanism of
enhanced ULVWF binding to platelets, we used optical tweezers
to study the unbinding forces from the glycoprotein Ib-IX (GP Ib-IX)
complex of plasma VWF, ULVWF, and isolated A1 domain. The unbinding
force was defined as the minimum force required to pull ligand-coated beads away from their attachment with GP Ib-IX-expressing cells. Beads
coated with plasma VWF did not bind to the cells spontaneously, requiring the modulators ristocetin or botrocetin. The force required to break the ristocetin- and botrocetin-induced plasma VWF-GP Ib-IX
bonds occurred in integer multiples of 6.5 pN and 8.8 pN, respectively,
depending on the number of bonds formed. In contrast, beads coated with
either ULVWF or A1 domain bound the cells in the absence of modulators,
with bond strengths in integer multiples of approximately 11.4 pN for
both. Thus, in the absence of shear stress, ULVWF multimers form
spontaneous high-strength bonds with GP Ib-IX, while plasma VWF
requires exogenous modulators. The strength of individual bonds formed
with GP Ib-IX was similar for both ULVWF and the isolated A1 domain and
greater than those of plasma VWF induced by either modulator.
Therefore, we suggest that the conformational state of ULVWF multimers
is more critical than their size for interaction with platelets.
(Blood. 2002;99:3971-3977) Platelet thrombi form in the arterial circulation
following injury and desquamation of the endothelial cells. At arterial shear stresses, platelet attachment to the vessel wall is initiated by
the binding of platelet glycoprotein (GP) Ib-IX-V complexes to von
Willebrand factor (VWF) multimers in the subendothelium. As a result,
platelets tether, roll, and then adhere firmly to the subendothelium.
This adhesion subsequently initiates a complex sequence of events that
includes platelet activation, secretion, and
cohesion.1-5
The GP Ib-IX-V complex is made up of 4 polypeptide chains: GP Ib VWF is a multimeric glycoprotein synthesized in endothelial cells and
megakaryocytes from which it is secreted constitutively or stored in
granules, the Although plasma VWF multimers are continuously in contact with
platelets, the platelet-VWF interaction cannot be detected in
circulating blood under normal conditions. In contrast, platelets will
attach to immobilized VWF multimers under the conditions of elevated
shear stress present in the normal arterial and arteriolar circulation.9,10 With pathologically elevated shear
stresses, as in regions of arterial stenosis, large plasma VWF
multimers attach to the platelets, with consequent activation and
aggregation of the platelets. ULVWF multimers are more active than the
largest plasma VWF multimers in promoting these
interactions.11 Evidence exists supporting a shear-induced
conformational alteration of both GP Ib-IX-V complexes12
and VWF multimers,13 although the precise mechanism by
which shear induces the interaction is unknown. In the absence of high
shear stress, the exogenous modulators ristocetin or botrocetin are
required for the binding of plasma VWF to GP Ib-IX.
It has been presumed that the interaction of VWF with GP Ib-IX
complexes on platelets is governed predominantly by its multimeric size.14 To address this question, we used optical tweezers
to investigate the interaction of GP Ib-IX with the different forms of
VWF: plasma VWF, ULVWF, and isolated A1 domain. We demonstrate that,
unlike plasma VWF, ULVWF requires no modulators to bind the GP Ib-IX
complex, and the minimal strength of its bond to the GP Ib-IX complex
is 1.5 times that of the plasma VWF-GP Ib-IX bond in the presence of
modulator and similar to the A1-GP Ib-IX bond. These findings suggest
that the conformational state of VWF multimers may be more critical
than their multimeric size in the VWF-GP Ib-IX interaction.
Cells expressing GP Ib-IX complexes.
Preparation of VWF and VWF-coated beads
Polystyrene beads (Polysciences, Warrington, PA) with a diameter of 2.0 µm were coated with VWF according to the following procedure. Single
drops of bead suspensions were added to tubes containing VWF at a
concentration of either approximately 10 or approximately 100 µg/mL
in citrate phosphate buffer. The beads and either purified VWF or ULVWF
multimers were then incubated for 1 hour at room temperature with
gentle mixing on an aliquot shaker. The suspension was subsequently
centrifugued for 2 minutes in an Eppendorf microcentrifuge at 15 000
rpm. The supernatant was removed, and the beads were resuspended by
vortex in Dulbecco phosphate-buffered saline (PBS) solution (pH, 7.3)
containing 10 mg/mL bovine serum albumin, 1 mg/mL sodium azide, and 5%
glycerol. The beads were then washed in 1 mL Dulbecco PBS solution,
incubated for 30 minutes, centrifuged for 2 minutes, and resuspended in 0.5 mL Dulbecco PBS solution.
Monoclonal antibody
Modulators The compounds botrocetin and ristocetin were used to induce binding between plasma VWF and GP Ib-IX. Botrocetin is a 25-kd dimeric protein purified from the venom of the South American pit viper Bothrops jararca. Botrocetin forms a complex with VWF by binding near the GP Ib-binding site.21 Ristocetin is an antibiotic glycopeptide synthesized by Nocardia lurida thought to interact with both GP Ib-IX and VWF,22 although the mechanism by which it induces the VWF-GP Ib-IX interaction remains unclear.Optical tweezers Damage to cells induced by light varies with the incident wavelength, but little or no damage has been demonstrated at 830 nm in an optical trap.23 We used a titanium-sapphire laser tuned to 830 nm (Model 3900 S; Spectra-Physics, Santa Clara, CA) in our optical tweezers setup (Figure 2). The titanium-sapphire laser was pumped by a solid-state, frequency-doubled neodynium yttrium vanadate (Nd:YVO4) laser operating at a wavelength of 532 nm (Millennia V; Spectra-Physics). The laser beam was expanded (CM cwbx-7.0-s-670/1064; Spectra-Physics) 5-fold to fill the back aperture of the microscope objective (numerical aperture 1.3). The laser light then passed through an attenuator (Model 925B; Newport Electronics, Irvine, CA) used to control the beam intensity. A dichroic mirror placed just before the bottom entrance port transmitted the laser light into the microscope (Axiovert S100TV; Carl Zeiss, Jena, Germany) and reflected the light below 650 nm toward a beam splitter, which transmitted 10% of the light to a charged-coupled device (CCD) camera (Model CCD 100; DAGE-MTI, Michigan City, IN) for imaging purposes. The solution chamber containing cells was mounted onto a piezoelectrically driven translational stage (Model P-527.3CL; Physik Instrumente, Waldbronn, Germany). The cell chamber was illuminated from the top with white light for visualizing the specimens.
Calibration procedure The optical trapping force was calibrated by moving a solution past a trapped bead at a known velocity with the use of the piezoelectric stage and calculating the force required to displace the bead from the trap using the Stokes law,
 is the solution viscosity (1 cP),
v is the solution velocity, r is the bead radius,
and h is the distance between the center of the bead and the
coverslip.24
For a given laser power, the bead will eventually escape the trap when
the drag force exerted by the fluid exceeds the trapping force. The
drag force at which the bead escapes from the trap is the escaping
force, and was determined over a range of laser powers measured past
the microscope objective lens. Figure 3
shows the escaping force as a function of laser power for a 2.0-µm
polystyrene bead placed at a height of 10 µm from the coverslip
(h = 10 µm). The system was calibrated at a height of 10 µm since the VWF-coated bead is in contact with the CHO cell
approximately 10 µm from the coverslip during the experiments. There
was a linear relationship between the escaping force and laser power
with a slope of approximately 0.9 pN/mW.
Interaction of VWF and GP Ib-IX complexes We optically trapped a 2.0-µm bead coated with VWF and moved the transfected CHO cell toward the trapped bead by using the piezoelectrically driven stage. Initial binding between plasma VWF and GP Ib-IX was induced by the addition of either 1 mg/mL ristocetin or 2 µg/mL botrocetin to the solution chamber. The CHO cell was placed in the solution chamber 20 minutes before an experiment to allow it to adhere firmly to the coverslip. The VWF-coated bead was placed into contact with the cell and allowed to remain there for 10 seconds. Adhesion times up to 90 seconds between the bead and the cell were also examined. Following adhesion, the laser power was reduced by adjusting the attenuator to the minimum level required to trap the VWF-coated bead.To determine bond strength, we attempted to detach the cell from the bead by moving the piezoelectric stage. If the cell could not be detached from the bead, the laser power was incrementally increased, and another attempt was made to detach the bead from the cell. This process was repeated until the minimum laser power required to detach the VWF-coated bead from the CHO cell was determined. The minimum optical power required to detach the bead was converted to a force value that represented the bond strength of the VWF-GP Ib-IX interaction by means of the calibration curve. To induce the formation of multiple bonds, the VWF-coated bead was forcefully pushed against the cell and subsequently detached after 10 seconds of adhesion. After measuring the forces required to detach the coated bead from the cell, we observed distinct groups of data points whose cluster means were integral multiples of putative single-bond strengths for each ligand-receptor interaction studied. Interactions of modulators with VWF Polystyrene beads (r = 2.25 µm) were coated with ristocetin or botrocetin, and smaller beads (r = 1.0 µm) were coated with plasma VWF in order to determine whether the modulators were detaching from the VWF. The modulator-coated bead was first allowed to adhere firmly to the coverslip for 20 minutes. We then trapped the VWF-coated bead and allowed it to bind to the modulator-coated bead for 10 seconds. The force to detach the VWF-coated bead from the modulator-coated bead was then determined as described above.Statistics Each result is reported as the mean ± SD. Student t test was used to evaluate differences between the mean values of the putative single-bond strengths for GP Ib-IX and its ligands.
A bond was considered to have formed if the force needed to
retract the VWF-coated bead from the CHO cell was greater than the
force needed to move the bead in solution in the absence of CHO cells.
In the absence of ristocetin or botrocetin, no adhesion was detected
between plasma VWF-coated beads and CHO
Ristocetin-induced binding of purified plasma VWF to CHO
  IX cells only in the presence of either ristocetin or
botrocetin. In the presence of ristocetin, the force necessary to break
the bonds occurred in integer multiples of the putative single-bond
strength, 6.5 ± 0.8 pN, depending upon the number of bonds
initially formed (Figure 5). With beads
coated at a higher VWF density, more bonds formed on average.
Nevertheless, the value for a single bond was the same as with beads
coated at lower density (Figure 5B).
Botrocetin-induced plasma VWF binding to CHO
Ristocetin and botrocetin adhesion to VWF To ensure that neither botrocetin nor ristocetin was being detached from VWF during the experiments, we measured the unbinding forces between plasma VWF-coated beads and beads coated with either botrocetin or ristocetin. It was not possible to detach the 2 spheres, whether the modulator was ristocetin or botrocetin, even at maximum laser power.Binding of ULVWF to CHO IX cells spontaneously, without
the need for ristocetin or botrocetin. The ULVWF-GP Ib-IX single-bond
strength was 11.4 ± 2.1 pN. Higher measured values were integral
multiples of this value (Figure 7). At
higher ULVWF-coating densities, more bonds tended to form; the strength
of the single bond was again identical with that found at lower
ULVWF-coating density.
Binding of the VWF A1 domain to CHO IX cells in the absence of modulators, with a single-bond strength of approximately 11.5 pN per bond (Figure
8), virtually identical to that observed
for the interaction of ULVWF with the CHO IX cells.
Statistics The mean single-bond strengths of the interactions between the GP Ib-IX complex and each of the different forms of VWF (plasma, ULVWF, and the A1 domain) were compared by means of the Student t test. The ULVWF-GP Ib-IX and A1-GP Ib-IX bond strengths were not significantly different (P = .7414). The remaining bond strengths were significantly different (P < .05) when compared with each other.
The interaction between the platelet GP Ib-IX-V complex and VWF is the initiating event of platelet adhesion. This interaction normally does not occur in the fluid phase, unless induced by extremely high shear stresses.3 When bound VWF is exposed on the subendothelium, however, it is able to interact with the GP Ib-IX-V complex and thus to capture platelets from the flowing blood, a process that becomes more efficient with increasing shear stress.25 Considerable evidence exists that unprocessed forms of ULVWF are more efficient at binding platelets and are capable of binding platelets spontaneously. This spontaneous binding, with subsequent aggregation of the platelets, is the presumed pathophysiology of the catastrophic arterial thrombotic disorder, thrombotic thrombocytopenic purpura (TTP), which is characterized pathologically by extensive microvascular occlusion with thrombi composed almost entirely of platelets and VWF. TTP is believed to be caused by deficiency of a recently characterized protease of the disintegrin and metalloproteinase domain, with thrombospondin type-1 modules (ADAMTS) family (ADAMTS13),26-29 which is responsible for the proteolytic processing of the ULVWF to less reactive plasma forms. Although the interaction between the GP Ib-IX-V complex and VWF has been extensively studied, the basis of the increased reactivity of ULVWF remained a mystery. It has been presumed that multimeric size was the main determinant of the reactivity of the ultralarge multimers, partially on the basis of the increased propensity for bleeding observed when the larger plasma forms are missing in types 2A and 2B von Willebrand disease and in platelet-type von Willebrand disease.25,30 This hypothesis has never been tested directly, primarily because of the difficulty of isolating single ligand-receptor bonds (by single bond, we mean here the minimum stoichiometry of ligand and receptor capable of producing a measurable bond). We were able to overcome this technical obstacle in the current studies with the use of optical tweezers. This technique allowed us to compare in detail the interaction of the GP Ib-IX complex with the different forms of VWF, and to do so at the level of single-bond units. Our results reveal a fundamental difference at the level of single
bonds between unprocessed ULVWF and VWF purified from plasma. Whereas
the plasma VWF required the modulators ristocetin or botrocetin to bind
cells expressing the GP Ib-IX complex, ULVWF bound the cells
spontaneously, without modulators. In addition, the strength of the
individual bond between VWF and the GP Ib-IX complex was more than 1.3 times greater for ULVWF than for plasma VWF in the presence of either
modulator. Thus, not only is ULVWF of much higher molecular weight on
average than plasma VWF; it is inherently stickier. Processing ULVWF to
a less adhesive form is thus part of the normal physiological role of
the ADAMTS13 protease. Of great relevance to our attempts to understand
the nature of the processing step was our finding that the strength of
the individual bonds formed between ULVWF and GP Ib-IX was virtually
identical to the strength of the bonds formed between the GP
Ib-IX complex and the isolated A1 domain. Thus, it appears that not
only does unprocessed ULVWF exist, on average, in a form of higher
molecular weight, but its GP Ib These studies also raise the possibility that one reason for the observed difference in binding between plasma VWF and VWF immobilized on the subendothelium is that the latter form may not have undergone complete processing before it binds platelets, perhaps because it was secreted abluminally to a compartment inaccessible to the plasma protease. Such a scenario would provide an elegant mechanism by which subendothelial VWF could bind platelets efficiently, while the plasma forms remain relatively unreactive. Of interest, plasma VWF can mediate the attachment and rolling of platelets when coated onto a glass surface, but the relationship of this interaction to that of platelets with subendothelial VWF in vivo is unknown. Several studies have used optical tweezers to examine
receptor-ligand relationships in biological systems. For
example, Liang et al31 examined adhesion of
Escherichia coli to mannose-presenting surfaces. In that
study, integral multiples of 1.7 pN were reported to represent the
force to detach one molecule of In addition to helping us distinguish between plasma VWF and ULVWF, the optical tweezer technique allowed us to compare the bonds induced by botrocetin and ristocetin. The 2 modulators induce the interaction by fundamentally different mechanisms, and this is borne out by our data, which demonstrate that the modulators induce bonds of different strengths between plasma VWF and the GP Ib-IX complex. These studies also preclude the possibility that we were dissociating the interaction between the modulator and VWF, as the VWF-coated beads formed much stronger bonds with beads coated with either ristocetin or botrocetin than they did with GP Ib-IX complex-expressing cells. The difference in binding strength induced between the 2 modulators and the yet stronger binding of ULVWF- or A1-coated beads also excludes the possibility that the forces being measured were those required to extract the GP Ib-IX complex from the cell membrane. Other investigators have determined that ristocetin binds to both VWF and GP Ib, thereby facilitating adhesion between the receptor and its ligand.36 The results of the ristocetin-dependent plasma VWF/GP Ib-IX-binding experiments are quite intriguing since our lowest force measurement is 6.5 pN while the subsequent value is 39 pN, exactly 6 times the initial value. The higher bond strength values all increased in integer multiples of 39 pN. This pattern of increase in the force measurement stands in stark contrast to the stepwise increase from the minimal value in the botrocetin-mediated VWF/GP Ib-IX force data. This raises the possibility that the true single-bond strength for the ristocetin-induced VWF/GP Ib-IX bond is approximately 39 pN. While the exact mechanism of ristocetin-induced binding of VWF to GP Ib is not known, some have suggested that ristocetin binds to GP Ib through 4 ristocetin monomers or through 2 ristocetin dimers.22 While 2 ristocetin dimers could indeed act as a bridge between VWF and GP Ib, it is also possible that multiple ristocetin dimers, perhaps 6, could facilitate the VWF/GP Ib interaction. Future experiments using the optical tweezers will allow us to address the nature of the ristocetin-dependent VWF/GP Ib bond. In summary, we have used optical tweezers to compare the strength of the interactions between different forms of VWF and the platelet GP Ib-IX complex. These studies confirm the notion that the modulators ristocetin and botrocetin differ in the nature of the bond they induce between the ligand and the receptor. The studies have also allowed us to examine the basis of the enhanced reactivity with platelets observed for ULVWF as compared with the usual plasma forms of VWF, revealing that ULVWF is inherently more adhesive for platelets than is plasma VWF. This finding may shed some light on the pathophysiology of the thrombotic microangiopathies.
We thank Ms Nancy Turner and Ms Letty Nolasco for preparation of the VWF and ULVWF, and Mr Zhiwei Li, Ms Kathryn Simpson, and Dr Jorge Torres for invaluable assistance with the optical tweezers.
Submitted November 19, 2001; accepted January 22, 2002.
Prepublished online as Blood First Edition Paper, April 17, 2002; DOI 10.1182/ blood-2001-11-0060.
Supported by National Institutes of Health grants 1P50 HL65967 and NS23327; the Robert A. Welch Foundation grant no. C938; a grant-in-aid from the American Heart Association-Texas Affiliate; and Special Opportunity and Development Awards from The Whitaker Foundation to Rice University.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: José A. López, Thrombosis Research Section, Department of Medicine, BCM286, N1319, Baylor College of Medicine, 1 Baylor Plaza, Houston, TX 77030; e-mail: josel{at}bcm.tmc.edu.
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 Z. M. Ruggeri and G. L. Mendolicchio Adhesion Mechanisms in Platelet Function Circ. Res., June 22, 2007; 100(12): 1673 - 1685. [Abstract] [Full Text] [PDF]
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C. K. N. K. Chion, C. J. M. Doggen, J. T. B. Crawley, D. A. Lane, and F. R. Rosendaal ADAMTS13 and von Willebrand factor and the risk of myocardial infarction in men Blood, March 1, 2007; 109(5): 1998 - 2000. [Abstract] [Full Text] [PDF]
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T. C. Nguyen, A. Liu, L. Liu, C. Ball, H. Choi, W. S. May, K. Aboulfatova, A. L. Bergeron, and J.-f. Dong Acquired ADAMTS-13 deficiency in pediatric patients with severe sepsis Haematologica, January 1, 2007; 92(1): 121 - 124. [Abstract] [Full Text] [PDF]
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Z. M. Ruggeri, J. N. Orje, R. Habermann, A. B. Federici, and A. J. Reininger Activation-independent platelet adhesion and aggregation under elevated shear stress Blood, September 15, 2006; 108(6): 1903 - 1910. [Abstract] [Full Text] [PDF]
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A. J. Reininger, H. F. G. Heijnen, H. Schumann, H. M. Specht, W. Schramm, and Z. M. Ruggeri Mechanism of platelet adhesion to von Willebrand factor and microparticle formation under high shear stress Blood, May 1, 2006; 107(9): 3537 - 3545. [Abstract] [Full Text] [PDF]
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 A. K. Chauhan, D. G. Motto, C. B. Lamb, W. Bergmeier, M. Dockal, B. Plaimauer, F. Scheiflinger, D. Ginsburg, and D. D. Wagner Systemic antithrombotic effects of ADAMTS13 J. Exp. Med., March 20, 2006; 203(3): 767 - 776. [Abstract] [Full Text] [PDF]
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R. Donadelli, J. N. Orje, C. Capoferri, G. Remuzzi, and Z. M. Ruggeri Size regulation of von Willebrand factor-mediated platelet thrombi by ADAMTS13 in flowing blood Blood, March 1, 2006; 107(5): 1943 - 1950. [Abstract] [Full Text] [PDF]
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B. Plaimauer, J. Fuhrmann, G. Mohr, W. Wernhart, K. Bruno, S. Ferrari, C. Konetschny, G. Antoine, M. Rieger, and F. Scheiflinger Modulation of ADAMTS13 secretion and specific activity by a combination of common amino acid polymorphisms and a missense mutation Blood, January 1, 2006; 107(1): 118 - 125. [Abstract] [Full Text] [PDF]
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Z. Tao, Y. Peng, L. Nolasco, S. Cal, C. Lopez-Otin, R. Li, J. L. Moake, J. A. Lopez, and J.-f. Dong Recombinant CUB-1 domain polypeptide inhibits the cleavage of ULVWF strings by ADAMTS13 under flow conditions Blood, December 15, 2005; 106(13): 4139 - 4145. [Abstract] [Full Text] [PDF]
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J. J. J. Hulstein, P. G. de Groot, K. Silence, A. Veyradier, R. Fijnheer, and P. J. Lenting A novel nanobody that detects the gain-of-function phenotype of von Willebrand factor in ADAMTS13 deficiency and von Willebrand disease type 2B Blood, November 1, 2005; 106(9): 3035 - 3042. [Abstract] [Full Text] [PDF]
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 S. M. Kawut, E. M. Horn, K. K. Berekashvili, A. C. Widlitz, E. B. Rosenzweig, and R. J. Barst von Willebrand Factor Independently Predicts Long-term Survival in Patients With Pulmonary Arterial Hypertension Chest, October 1, 2005; 128(4): 2355 - 2362. [Abstract] [Full Text] [PDF]
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 K. Nishio, P. J. Anderson, X. L. Zheng, and J. E. Sadler Binding of platelet glycoprotein Ib{alpha} to von Willebrand factor domain A1 stimulates the cleavage of the adjacent domain A2 by ADAMTS13 PNAS, July 20, 2004; 101(29): 10578 - 10583. [Abstract] [Full Text] [PDF]
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A. Bernardo, C. Ball, L. Nolasco, J. F. Moake, and J.-f. Dong Effects of inflammatory cytokines on the release and cleavage of the endothelial cell-derived ultralarge von Willebrand factor multimers under flow Blood, July 1, 2004; 104(1): 100 - 106. [Abstract] [Full Text] [PDF]
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C. Klaus, B. Plaimauer, J.-D. Studt, F. Dorner, B. Lammle, P. M. Mannucci, and F. Scheiflinger Epitope mapping of ADAMTS13 autoantibodies in acquired thrombotic thrombocytopenic purpura Blood, June 15, 2004; 103(12): 4514 - 4519. [Abstract] [Full Text] [PDF]
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A. Padilla, J. L. Moake, A. Bernardo, C. Ball, Y. Wang, M. Arya, L. Nolasco, N. Turner, M. C. Berndt, B. Anvari, et al. P-selectin anchors newly released ultralarge von Willebrand factor multimers to the endothelial cell surface Blood, March 15, 2004; 103(6): 2150 - 2156. [Abstract] [Full Text] [PDF]
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 J. E. Sadler, J. L. Moake, T. Miyata, and J. N. George Recent Advances in Thrombotic Thrombocytopenic Purpura Hematology, January 1, 2004; 2004(1): 407 - 423. [Abstract] [Full Text] [PDF]
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J.-f. Dong, J. L. Moake, A. Bernardo, K. Fujikawa, C. Ball, L. Nolasco, J. A. Lopez, and M. A. Cruz ADAMTS-13 Metalloprotease Interacts with the Endothelial Cell-derived Ultra-large von Willebrand Factor J. Biol. Chem., August 8, 2003; 278(32): 29633 - 29639. [Abstract] [Full Text] [PDF]
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 K. H. Simpson, G. Bowden, M. Hook, and B. Anvari Measurement of Adhesive Forces between Individual Staphylococcus aureus MSCRAMMs and Protein-Coated Surfaces by Use of Optical Tweezers J. Bacteriol., March 15, 2003; 185(6): 2031 - 2035. [Abstract] [Full Text] [PDF]
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J.-f. Dong, J. L. Moake, L. Nolasco, A. Bernardo, W. Arceneaux, C. N. Shrimpton, A. J. Schade, L. V. McIntire, K. Fujikawa, and J. A. Lopez ADAMTS-13 rapidly cleaves newly secreted ultralarge von Willebrand factor multimers on the endothelial surface under flowing conditions Blood, December 1, 2002; 100(12): 4033 - 4039. [Abstract] [Full Text] [PDF]
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J. E. Sadler A new name in thrombosis, ADAMTS13 PNAS, September 3, 2002; 99(18): 11552 - 11554. [Full Text] [PDF]
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 J. L. Moake Thrombotic Microangiopathies N. Engl. J. Med., August 22, 2002; 347(8): 589 - 600. [Full Text] [PDF]
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Top
Abstract
Introduction
= 830 nm) is reflected by a
dichroic mirror and is focused onto the objective lens of the inverted
microscope to trap the bead. A charged-coupled device (CCD) camera
collects 10% of the light for image visualization. The CHO cell and
VWF-coated bead are in a solution chamber mounted onto a
piezoelectrically driven translational stage.
