Yoshihito Hayashi

Dielectric coagulometry: a new approach to estimate venous thrombosis risk.

We present dielectric coagulometry as a new technique to estimate the risk of venous thrombosis by measuring the permittivity change associated with the blood coagulation process. The method was first tested for a simple system of animal erythrocytes suspended in fibrinogen solution, where the coagulation rate was controlled by changing the amount of thrombin added to the suspension. Second, the method was applied to a more realistic system of human whole blood, and the inherent coagulation process was monitored without artificial acceleration by a coagulation initiator. The time dependence of the permittivity at a frequency around 1 MHz showed a distinct peak at a time that corresponds to the clotting time. Our theoretical modeling revealed that the evolution of heterogeneity and the sedimentation in the system cause the peak of the permittivity.

Dielectric inspection of erythrocyte morphology

We performed a systematic study of the sensitivity of dielectric spectroscopy to erythrocyte morphology. Namely, rabbit erythrocytes of four different shapes were prepared by precisely controlling the pH of the suspending medium, and their complex permittivities over the frequency range from 0.1 to 110 MHz were measured and analyzed. Their quantitative analysis shows that the characteristic frequency and the broadening parameter of the dielectric relaxation of interfacial polarization are highly specific to the erythrocyte shape, while they are insensitive to the cell volume fraction. Therefore, these two dielectric parameters can be used to differentiate erythrocytes of different shapes, if dielectric spectroscopy is applied to flow-cytometric inspection of single blood cells. In addition, we revealed the applicability and limitations of the analytical theory of interfacial polarization to explain the experimental permittivities of non-spherical erythrocytes.

Temporal variation of dielectric properties of preserved blood

Rabbit blood was preserved at 277 K in Alsevers solution for 37 days, and its dielectric permittivity was monitored in a frequency range from 0.05 to 110 MHz throughout the period. The relaxation time and Cole-Cole parameter of the interfacial polarization process for erythrocytes remained nearly constant during the first 20 days and then started to increase and decrease, respectively. On the other hand, the relaxation strength and the cell volume fraction continued to decrease for 37 days, but the decrease rates of both changed discontinuously on about the 20th day. Microscope observation showed that approximately 90% of the erythrocytes were spinous echinocytes at the beginning of preservation and started to be transformed into microspherocytes around the 20th day. Therefore, dielectric spectroscopy is a sensitive tool to monitor the deterioration of preserved blood accompanied by morphological transition of erythrocytes through the temporal variation of their dielectric properties.

Dielectric Cytometry with Three-Dimensional Cellular Modeling

We have developed what we believe is an efficient method to determine the electric parameters (the specific membrane capacitance C(m) and the cytoplasm conductivity kappa(i)) of cells from their dielectric dispersion. First, a limited number of dispersion curves are numerically calculated for a three-dimensional cell model by changing C(m) and kappa(i), and their amplitudes Deltaepsilon and relaxation times tau are determined by assuming a Cole-Cole function. Second, regression formulas are obtained from the values of Deltaepsilon and tau and then used for the determination of C(m) and kappa(i) from the experimental Deltaepsilon and tau. This method was applied to the dielectric dispersion measured for rabbit erythrocytes (discocytes and echinocytes) and human erythrocytes (normocytes), and provided reasonable C(m) and kappa(i) of the erythrocytes and excellent agreement between the theoretical and experimental dispersion curves.

The effects of erythrocyte deformability upon hematocrit assessed by the conductance method

A comparative study of centrifugation and conductance methods for the estimation of cell volume fraction (phi) was performed to examine whether the strong forces exerted upon erythrocytes during centrifugation affect their volume, and the results are discussed in terms of erythrocyte deformability. Rabbit erythrocytes of four shapes (spherocytes, echinocytes, stomatocyte-like enlarged erythrocytes and discocytes) were prepared by controlling the pH of the suspending media. The packed cell volumes of the suspensions were measured by standard hematocrit determination methods using centrifugation in capillary tubes. Simultaneously, the same suspensions and their supernatants were used in dielectric spectroscopy measurements, and the low-frequency limits of their conductivities were used for the numerical estimation of phi. The hematocrit values of spherocytes and echinocytes were markedly less than the volume fractions obtained by the conductance method. Namely, the centrifugation reduced the cell volume. For enlarged erythrocytes and discocytes, however, the reduction of cell volume was not observed. These findings showed that phi obtained by the centrifugation method can be greatly affected by the deformability of the cells, but the level of the effect depends on the cell types. Consequently, phi obtained by the centrifugation method should be carefully interpreted.