Biomedical Engineering Reference
In-Depth Information
thrombin generation as well as the total amount of thrombin formation. Thrombin generation can
be determined in several ways. The easiest way is to determine thrombin generation under static
conditions. For this, preferably full blood is incubated with the blood-contacting surface and the
concentration of thrombin is determined in samples taken over the period of the experiment.
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The
only problem is that when a clot has formed, the experiment has to be stopped since no homogenous
samples can be drawn anymore. Since most thrombin generation occurs inside the clot, after clot
formation, the time for thrombin generation is the only parameter that can be determined using
this assay.
109,111
However, not only the lag time in thrombin generation is a parameter for blood
compatibility, but also the so-called thrombin potential is important, which is an indicator for the
total amount of thrombin formed.
109,112,113
This thrombin potential can be determined in PRP using
a fl uorescently labeled thrombin substrate. The blood-contacting material is incubated with PRP
and fl uorescent substrate, and fl uorescence is constantly monitored. When the fl uorescence is plot-
ted against time, the slope of the curve is a direct measure for the concentration of active thrombin
(Figure 17.14). This assay has recently been optimized and can now be seen as the golden standard
to study hemostatic function of the blood. Since the method is very sensitive to a variety of param-
eters, it may be well suited to determine biomaterial-induced thrombin generation.
Dynamic incubations with blood mimic the
in vivo
situation better, although the choice of the
fl ow rate, and consequently shear rates, are of critical importance. There are several methods for
dynamic blood contact
in vitro
.
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The fi rst is to pump recalcifi ed blood along the material or through
a tube on which the surface coating of the device is deposited (Figure 17.15).
This blood can be analyzed for a number of coagulation parameters at the end of the conduit.
The concentration of thrombin, thrombin-AT (TAT) complexes, fi brinogen degradation products,
activation of the complement system, the consumption of platelets as a measure of platelet adhe-
sion, can all be determined in the collected blood samples.
106
A second, regularly used, set-up
is the closed-loop model, in which the inside of a tube is representative for the synthetic surface
of an implant.
114
The blood can be left in the tube for any desired period of time, and samples
can be taken at desired intervals although sampling means the system has to be stopped and
opened, which is not an ideal condition in practice. Both these methods use only parts of the blood-
contacting device and often also in another confi guration, implying that these devices are not as
used in the fi nal device.
PTFE
PTFE + heparin
50
40
30
20
10
0
30
50
60
10
20
40
Time (min)
FIGURE 17.14
Thrombin generation using fl uorescent substrate. The fl uorescence over time is measured
and the differential is plotted. The area under the curve represents the thrombin potential.
