Thrombus Formation on Ultrasonically Vibrating Surface
Shigehiro Hashimoto, Tomoaki Nakamura, Osamu Hasegawa, Michitake Yasaka
Department of Electronics, Information and Communication Engineering, Osaka Institute of Technology
5-16-1, Ohmiya, Asahi-ku, Osaka, 535-8585, Japan
hasimoto@elc.oit.ac.jp

ABSTRACTThe thrombus formation on ultrasonically vibrating
surface was experimentally studied in vitro. A blood
sample was contained on the glass plate attached to thepiezoelectric oscillator, and erythrocytes aggregation wasobserved during clotting process with a phase contrast
microscope. A drop of blood sample of 0.01 ml wasspotted on the vibrating rock crystal, and the resonancefrequency was continuously measured with an electric
circuit. The results show that clotting time is shortened on
the vibrating surface, and that the resonance frequency at
oscillating crystal shifts when the contained blood starts
forming clot.

was initiated by adding calcium chloride (0.1:1 calcium
chloride/ blood; 0.25 M aqueous solution) before the test.
To observe erythrocytes aggregation during clotting
process on ultrasonic vibrating surface, a piezoelectricoscillator (from 1 Hz to 1 MHz) was attached to the glassplate (Fig. 1). The blood sample was contained on thevibrating glass plate, and observed for ten minutes with aphase contrast microscope, a CCD (charge-coupleddevice) camera, a video tape recorder, a monitor, and acomputer (Fig. 2).

Keywords: Bio-Measurement, Thrombus, UltrasonicVibration, Piezoelectric Oscillator, Crystal Oscillator,
Resonance

1. INTRODUCTION
Many factors control the dynamic balance between theformation and removal of clots. This control repairs theruptured vessel wall, and maintains the blood flow path.
These factors include platelets, coagulants andanti-coagulants. Many studies have been performed inthis research field from the biochemical point of view,which includes the interaction between the blood and thesurface materials [1]. On the other hand, there is ahydrodynamic effect on clot growth in shear flow. The
clotting time decreases in pulsatile flow [2]. The stirring
effect of blood coagulants may accelerate the reaction forclot formation. Many instruments with ultrasonic wave
are used in medical treatment. To design the applicationof ultrasonic oscillation to medical devise in contact withblood, the effect of ultrasonic vibration on thrombus
formation was investigated in vitro.

Fig. 1. Piezoelectric oscillator.

2. METHODS
Erythrocyte aggregation
Human blood was collected into sodium citrate aqueous
solution (1:6.6 sodium citrate/ blood; 9.8 mM final
concentration in blood). The blood coagulation process

Fig. 2. Experimental system.

Resonance frequency shift
A rock crystal (quartz) oscillator (Fig. 3) of 18 MHz
resonance frequency was applied to detect blood clotting
time. After removing the cover of the crystal oscillator, adrop of blood sample of 0.010 ml was spotted on thevibrating crystal, and the resonance frequency wascontinuously measured with an electric circuit (Figs. 4, 5).

Fig. 5. Measurement system. Left, Frequency
counter. Right, Oscilloscope.

Fig. 3. Crystal oscillator. Left, with cover. Right,Oscillator disk (8.0 mm diameter) after removing
the cover.

Fig. 6a. Erythrocytes before aggregation (0 min).

Fig. 4. Electric circuit

Fig. 6b. Erythrocytes aggregation during clotting
process (2 min).

Time (t), sec
0100

200300400

Fig. 6c. Erythrocytes aggregation during clotting
process (4 min).

Frequency shift (Fs), kHz

0-0.2
-0.4
-0.6
-0.8
-1
-1.2
calcium
anticoagulant

Fig. 7. Relation between frequency and time.

forming clot. After forming clot (“calcium” in Fig. 7), thefrequency shift was different from that of blood with“anticoagulant”.

4. DISCUSSION
Fig. 6d. Erythrocytes aggregation during clotting
process (6 min).

3. RESULTS
Fig. 6 shows erythrocytes aggregation during clotting
process at 1 MHz. Density of erythrocytes was uniform,
before formation of clot (Fig. 6a). After a few minutes,
erythrocytes started to aggregate and form some colonies(Figs. 6b-6d). This aggregating time was three minutes
shorter on the vibrating glass surface than on the static
glass surface.
Fig. 7 exemplifies the relation between resonancefrequency and time. The resonance frequency decreased
by 0.1 % (Fo) after a drop of blood spotted on thevibrating crystal. Frequency shift (Fs) was calculated byFs = Ft – Fo(1)where Ft is the resonance frequency at each time (t), andwhere Fo is resonance frequency at fifteen seconds after
spotting the blood. The frequency shifted by step in a few
minutes (3.5 minutes in Fig. 7), when the blood started

Many instruments with ultrasonic wave are used inmedical treatment. To design the application of ultrasonicoscillation to medical devise in contact with blood, theeffect of ultrasonic vibration on thrombus formation wasinvestigated in vitro.
The previous study shows that the clotting time decreases
in the pulsatile flow by the stirring effect of blood
coagulants, while clot does not grow in the pulsatile flowby the wash out effect [2]. The present study shows that
there is an analogy between clot formation on theultrasonically vibrating surface and that in the pulsatile
flow. The ultrasonically vibrating surface shortens theclotting time and may prevent clot growth.
The crystal oscillator has stable resonance frequency. A
shift of resonance frequency occurs with molecularadsorption, and it has been applied to the gas molecule
sensor [3]. In the present study, the crystal oscillator wasused to apply ultrasonic vibration to the blood, and todetect clot formation. The present study shows that theshift of resonance frequency is able to be applied tomeasurement of clot formation.

5. CONCLUSION
The present study shows that the clotting time decreaseson the ultrasonically vibrating surface. The shift ofresonance frequency is able to be applied to measurementof clotting time.

6. ACKNOWLEDGMENT
This work was supported in part by a Grant-in-Aid for
Scientific Research from the Japanese Ministry ofEducation, Science, Sports and Culture.

7. REFERENCES[1] E. W. Salzman, Interaction of the Blood with Naturaland Artificial Surfaces, New York: Marcel Dekker,
1981.
[2] S. Hashimoto, “Clot Growth under PeriodicallyFluctuating Shear Rate”, Biorheology, Vol. 31, 1994,
pp. 521-532.
[3] M. Ohnishi, T. Ishibashi, M. Aoki, C. Ishimoto,“Gassensitivity of Modified Surfaces of a Surface
Acoustic Wave Device by Chemical Adsorption
Technique and Langmuir-Blodgett Technique”, Jpn. J.
Appl. Phys. Vol. 33, 1994, pp. 5981-5986.