WO2008097598A2

WO2008097598A2 - Blood volume analyzer

Info

Publication number: WO2008097598A2
Application number: PCT/US2008/001605
Authority: WO
Inventors: Kenneth R. Kensey; Daniel J. Cho
Original assignee: Kensey Kenneth R; Cho Daniel J
Priority date: 2007-02-06
Filing date: 2008-02-07
Publication date: 2008-08-14
Also published as: WO2008097663A3; WO2008097663A2; WO2008097598A3

Abstract

A method for estimating blood content of whole blood that includes measuring an electrical property of whole blood, diluting the whole blood with a fluid of known electrical property, measuring the electrical property of the diluted whole blood, and estimating the content of blood based on the changes in the electrical property of the whole blood before and after dilution.

Description

BLOOD VOLUME ANALYZER

RELATED APPLICATION

[0001] The application is based on and claims priority to United States Provisional

Application Serial No. 60/888,398, filed February 6, 2007, entitled Blood Volume

Analyzer, to which a claim of priority is hereby made and the disclosure of which is incorporated by reference.

FIELD OF THE INVENTION

[0002] The present invention relates generally to blood volume analysis and more particularly to measuring the volume fraction of red blood cells (hematocrit) in whole blood.

BACKGROUND

[0003] Hematocrit is a volume percent of red blood cells in whole blood. The normal range of the hematocrit is 37 - 47% for females, and 45 - 52% for males. Small office labs and stat labs often measure hematocrit simply by spinning down a whole blood sample in a capillary tube and measuring the length of the column of red blood cells relative to the length of the column of the whole blood specimen.

[0004] Larger labs use automated methods employing, for example, a Coulter counting machine, Bayer ADVIA 120 analyzer, Sysmex analyzer, and ABX analyzer. Most automated methods measure the electrical impulse generated by the cell when it passes through an impedance-based analyzer or an optical-based analyzer. The volume of individual erythrocytes can be electronically determined by the measurement of electrical impedance or light-scattering properties thereof. For example, in a Coulter counting machine, the blood sample is diluted in an electrically charged solution and is moved slowly through an aperture across which a specific current is applied. As each cell passes through, the impedance varies and accordingly the voltage changes, creating a pulse. The voltage magnitude varies also with cell size, which allows the cells to be counted, and sized. Particles greater than a certain threshold value are then counted as red blood cells. [0005] Due to the plasma trapping in the corner of red blood cells, the hematocrit readings obtained from the automated methods often give less than those obtained from the manual spinning method using a micro-centrifuge.

[0006] The membrane of red blood cells is made of a phospholipid layer, which has a dielectric property. On the other hand, plasma is essentially made of water, which conducts electrons and ions very well. Thus, the electric conductivity or conversely electrical resistance of whole blood varies significantly as a function of content of red blood cells. In whole blood with hematocrit of 40-45%, the electrical resistivity can be approximately 1.4 - 1.7 [Q m] . For example, the values of the electrical resistance of blood are 1.465 and 1.90 for Hct of 40% and 50% respectively. On the other hand, the electrical resistivity of the plasma is only 0.7 [Ω ■ m] because of the presence of a large number of ions such as sodium. [0007] Maxwell-Frick equation provides the following relationship for resistivity of

, , .₊ . ._ 1 + 0.0125Hct hematocπt: p = 0.586

1 - 0.0 lHct

[0008] Geddes and Sadler [1973] have provided the following relationship for resistivity of hematocrit: p[Ωm] = 0.537 _e ^0025Hct

[0009] The foregoing relationships can be used to estimate blood content from whole blood conductivity/resistivity.

SUMMARY OF THE INVENTION

[0010] In a method according to the present invention, whole blood is diluted by a quantity of a fluid having a known electrical resistivity (e.g. saline solution) and an electrical property of the whole blood such as the electrical conductivity or electrical resistivity thereof is measured before and after the addition of the fluid. Since the amount of the added saline solution is known accurately, one can estimate the blood volume in a body from the conductivity data obtained before and after the addition of the saline solution. [0011] In a first embodiment of the present invention, a quantity of whole blood is extracted from a patient's body before and after diluting the whole blood with a quantity of fluid having a known electrical property and the electrical property of the undiluted and diluted blood are measured in-vitro using a standard resistance measurement device, or a potentiostat. During the measurement, preferably the whole blood is continuously mixed using an agitator to avoid the sedimentation of red blood cells in the whole blood. [0012] In a second embodiment of the present invention, the electrical property of whole blood is measured in-vivo before and after diluting the whole blood with a quantity of fluid having a known electrical property. For example, microelectrodes may be used to measure changes in an electrical property of whole blood due to addition of a fluid such as a saline solution, and the changes in the electrical property of the whole blood due to the addition of the quantity of the fluid can be used to determine the volume of blood in the whole body.

[0013] In a third embodiment of the present invention, saline or the like fluid may be continuously added to whole blood, preferably in-vivo, and the electrical property of the blood is preferably measured periodically. Thereafter, a slope indicating the volume of added saline solution over time and a slope indicating the rate of change of electrical property of the whole blood can be constructed and used in estimating the volume of blood in the whole body.

[0014] Other features and advantages of the present invention will become apparent from the following description of the invention which refers to the accompanying drawings.

BRIEF DESCRIPTION OF THE FIGURES

[0015] Fig. 1 illustrates steps in a method according to the first embodiment of the present invention.

[0016] Fig. 2 illustrates a preferred setup for the practice of a method according to the present invention.

[0017] Fig. 3 diagrammatically illustrates a setup for the measurement of an electrical property of whole blood in a method according to the present invention.

[0018] Fig. 4 schematically illustrates a setup for the measurement of an electrical property of whole blood according to the preferred embodiment of the present invention.

DETAILED DESCRIPTION OF THE EMBODIMENTS

[0019] Referring to Fig. 1, in a method according to a first embodiment of the present invention, first a quantity of whole blood is obtained from a patient SlO. Thereafter, an electrical property of the whole blood is measured S 12. Thus, for example, as the electrical property, in the preferred embodiment, the resistivity of the whole blood, or the conductivity of the whole blood (which is the inverse of the resistivity value) is measured. Next, in order to dilute the whole blood in the body, a known quantity of a fluid having a known electrical property S 14 is injected into the body. For example, in the preferred embodiment, a known quantity of a saline solution with a known electrical resistivity is injected into the body. Thus, the percentage of hematocrit will decrease with the dilution and the electrical resistivity of the diluted blood will decrease accordingly. Then, diluted blood is again obtained and the electrical property of whole blood measured in S12 is again measured S 16. For example, the electrical resistivity or the electrical conductivity of the diluted whole blood is measured. Thereafter, since the exact amount of the saline volume added to the circulation system is known, the total volume of the original whole blood prior to the addition of the saline solution can be estimated Sl 8 based on the values obtained from the measurement S 12 prior to dilution of the whole blood and the measurement S 16 after the dilution of the whole blood. [0020] Considering, for example, that the initial volume of whole blood in the body is X mL and the corresponding hematocrit is y_\ . Then, if a known volume of saline solution, for example 400 mL, and the hematocrit of the diluted whole blood is measured to be y₂, since the total accumulative volume of the cells in the whole blood does not change before and after the dilution, the following mass balance equation for the cell volume is

X y, = (X+400) y₂ where the hematocrits yj and y₂ are known values from measurements. Thus, determine the volume of whole blood in the body X can be determined. For example, if the initial hematocrit is measured to be 50%, when a Dextran solution of 400 mL is injected into the circulation system, and the hematocrit of the diluted blood is measured again to be 47%, then the following mass balance equation can be solved for the blood volume:

X(0.5) = (X+400) (0.47)

Thus, the initial blood volume X is determined to be 6,266 mL.

[0021] According to an aspect of the present invention, instead of actually measuring the two hematocrits before and after dilution of whole blood, in a method according to the present invention the hematocrit measurements are replaced with the measurements of electrical resistivity.

[0022] Referring to Fig. 2, a set up for practicing the first embodiment of the present invention includes a container 10 to retain the whole blood 12 obtained from a patient. The set up further includes a first electrode 14 and second electrode 16 each extending from the exterior to the interior of container 10 through a wall thereof. Electrodes 14 and 16 are electrically coupled to a resistance measurement device 18. Resistance measurement device 18 is provided to determine the resistivity of whole blood 12 according to a preferred embodiment of the present invention. [0023] Preferably, the set up further includes an agitator 20. Container 10 is placed atop agitator 20. Agitator 20 then agitates whole blood 12 in order to continuously mix the red blood cells therein in order to prevent the sedimentation of the red blood cells. [0024] The electrical conductivity/resistivity of a whole blood sample can be measured using an electrochemical method. In one preferred embodiment, a constant voltage potentiostat may be employed in the measurement of the electrical conductivity/resistivity of the whole blood before and after the dilution thereof.

[0025] A typical potentiostat includes a working electrode WE and a counter electrode CE. As is known, a constant voltage potentiostat keeps the voltage between the working electrode and the counter electrode constant by varying the supply of current to the same. Thus, a simple application of Ohm's law may be used to determine the conductivity/resistivity of an electrically conductive fluid, e.g. whole blood, which is penetrated by the electrodes of a potentiostat. Furthermore, because a potentiostat is used in a preferred method according to the present invention, the following relationship can be used to determine the content of blood in whole blood: h = K^e

[0026] where i_t and i_o represent the currents at time t and the initial condition. The exponent k is a proportionality constant which depends on diffusion coefficient of ions in blood, reduction-oxidation reactions, the material and surface characteristics of the electrodes. Note that the relationship set forth above indicates that the current decreases exponentially with time when a potentiostat is used to determine electrical conductivity/resistivity.

[0027] Referring, for example, to Figs. 3 and 4, in a typical application of the preferred embodiment of the present invention, counter electrode CE and working electrode WE form a circuit in which the whole blood (before and after dilution) forms a resistor 22 therebetween. During the operation of potentiostat 24, the initial peak current strength between electrodes CE and WE is reflective of the conductivity of the whole blood prior to the dilution.

[0028] In a preferred embodiment, short pulses (in the order of several microseconds) of current are supplied to the blood sample by the working electrode and the counter electrode of the potentiostat and the attenuation of current flow is monitored over time. The application of short pulses advantageously reduces the possibility of false readings in that a long lasting DC signal may lead to reactions which may adversely affect the impedance of the blood sample due to reduction or oxidation of electrodes CE and WE. [0029] In the preferred embodiment, a signal can be supplied to blood sample 22 periodically, and then the peak current value obtained after each period is compared to the current value obtained from the previous period. Thus, for example, a signal is supplied to blood sample 22 every 10 microseconds and the current value measured for each ten microsecond period is compared to the current value from the previous 10 microsecond period. That is, the value of the current (μA) at time t is compared to the value of current at t- Δt where t represents the last period and t-Δt represents the period immediately preceding the last period.

[0030] In the preferred embodiment, the working electrode can be made of an inert electrode such as platinum, gold or carbon rod, whereas the counter electrode can be made of normal hydrogen electrodes such as silver/silver chloride (Ag/ AgCl) or calomel wire. Preferably, the working electrode can have a diameter in the range of 0.5-5mm, which is a standard size for potentiostats. A much smaller diameter wire of 10 μm can also be used for the working electrode. For a counter electrode, a platinum wire can be used. [0031] In a second embodiment of the present invention, the electrical conductivity/resistivity of whole blood is measured in- vivo thus eliminating the need for obtaining blood from a patient and measuring the electrical conductivity/resitivity thereof before and after dilution thereof with saline in- vitro. Specifically, the electrical conductivity/resistivity of whole blood is measured in- vivo, a saline solution is added to the circulation system of the patient by any desired method including injection, and the electrical conductivity/resistivity of whole blood is measured again in- vivo after the addition of saline, all without withdrawing blood from the patient. With the addition of saline, the percentage of red blood cells should decrease while the plasma portion should increase within several minutes of the addition of saline. Hence, given that the volume of the added saline can be determined precisely, if the electrical conductivity/resistivity before and after the addition of the saline solution is measured, the blood volume in the circulation system can be estimated in-vivo. Thus, in a method according to the second embodiment of the present invention, the volume of added saline is monitored and correlated to the changes in the electrical conductivity/resistivity of whole blood in order to estimate the volume of blood in the whole blood. [0032] In a method according to the third embodiment of the present invention, a gradiometer concept can be employed to estimate the volume of blood in whole blood, hi this embodiment, a saline solution is continuously added to whole blood preferably in-vivo. Preferably, saline solution is linearly added over time. While saline is being added, the electric conductivity/resistivity of blood is measured periodically over a period of time. For example, electrical conductivity/resistivity of whole blood is measured every one minute over, for example, a ten minute period. Thereafter, two slopes can be constructed based on the data obtained. The first slope would indicate the change in the content of saline solution with time (i.e. the volume of saline added to the blood over time), and the second slope would indicate the rate of decrease in the electric conductivity (or rate of increase in the resistivity) of whole blood; i.e. the change in the electrical property of whole blood (e.g. electrical conductivity/resistivity) over time. From these two slopes, one can accurately estimate the blood volume in a body. Preferably, in the third embodiment, saline is added in-vivo, and the electrical conductivity/ resistivity is measured in-vivo. However, this method may be applied to blood that is extracted from a patient and retained in a container (i.e. in-vitro) without deviating from the scope and the spirit of the present invention. Moreover, the electrical property of whole blood may be measured continuously, rather than periodically. [0033] As an alternative to using a potentiostat, digital signal processing techniques can be used to determine hematocrit content in whole blood. Specifically, microelectrodes can be used to perform electrical conductivity/resistivity measurements on the blood (before and after dilution with saline or the like fluid). One of the advantages of using microelectrodes is that the measurement of the conductivity/resistivity of whole blood can be completed in a fraction of a second. Thus, blood content can be measured in-vivo without the extraction of blood from a patient. Furthermore, since the measurement is completed in a fraction of a second, the measurement error due to the separation of red blood cells from plasma can be practically eliminated. Thus, the blood volume can be measured in-vivo continuously over an extended period of time.

[0034] Although the present invention has been described in relation to particular embodiments thereof, many other variations and modifications and other uses will become apparent to those skilled in the art. It is preferred, therefore, that the present invention be limited not by the specific disclosure herein, but only by the appended claims.

Claims

WHAT IS CLAIMED IS:

1. A method for determining blood volume of whole blood, comprising: measuring an electrical property of said whole blood; diluting said whole blood with a quantity of a fluid having a known electrical property to obtain diluted whole blood; measuring said electrical property of said diluted whole blood; and estimating volume of blood in said whole blood based on said measured property of said whole blood before and after said diluting step.

2. The method of claim 1 , wherein said measured electrical property of said whole blood before and after said diluting step includes electrical resistivity of said whole blood.

3. The method of claim 1, wherein said measured electrical property of said whole blood before and after said diluting step includes electrical conductivity of said whole blood.

4. The method of claim 1, wherein said fluid comprises saline.

5. The method of claim 1, further comprising agitating said whole blood to prevent sedimentation of red blood cells.

6. The method of claim 1, wherein at least one of said measuring steps includes applying a DC current through said whole blood.

7. The method of claim 1, wherein said electrical property of said whole blood is measured using a potentiostat.

8. The method of claim 1, wherein at least one of said measuring steps comprises a hematocrit measurement using digital signal processing.

9. The method of claim 1, wherein at least one of said measuring steps is carried out using a constant voltage potentiostat having a working electrode and a common electrode.

10. The method of claim 9, further comprising periodically sending current pulses through said whole blood using said working electrode and said common electrode, and monitoring the initial peak value and the attenuation of current flow through said whole blood as a function of time.

11. The method of claim 10, wherein said current is measured every ten microseconds.

12. The method of claim 10, wherein said monitoring includes comparing values of the initial peak current through said whole blood with each current pulse.

13. The method of claim 9, wherein said working electrode is comprised of an inert material.

14. The method of claim 13, wherein said inert material is comprised of one of gold, platinum, and carbon.

15. The method of claim 9, wherein said common electrode is comprised of a hydrogen electrode.

16. The method of claim 15, wherein said hydrogen electrode is comprised of one of silver/silverchloride and calomel wire.

17. The method of claim 9, wherein said working electrode includes a wire having a diameter in the range 0.5-5 mm.

18. The method of claim 9, wherein said working electrode is comprised of a wire having a diameter of 10 microns.

19. The method of claim 9, wherein said common electrode is comprised of a platinum wire.

20. The method of claim 1, wherein said measuring steps and said diluting steps are performed in-vivo.

21. The method of claim 1 , further comprising measuring a quantity of said fluid, and correlating changes in electrical property of said whole blood to said quantity of said fluid.

22. The method of claim 1, wherein said whole blood is diluted by adding said fluid continuously to said whole, and further comprising monitoring a quantity of said added fluid, and periodically measuring said electrical property of said whole blood over time, wherein said volume of said blood is estimated based on said quantity of added fluid and a rate of change of said electrical property of said whole blood.