JPH04318463A - Method for measuring blood coagulation time - Google Patents

Abstract

PURPOSE:To enable a measurement error to be reduced by excluding an unneeded optical output which is obtained by an optical method in a method for measuring a blood-coagulation time using the optical method. CONSTITUTION:In a method for measuring a blood-coagulation time using an optical method, after a blood-coagulation inducing substance is added to a plasma. aging of an optical output within a certain time (T0) and a total amount of change ( Y) in optical output of that period are measured and then a time T where a specified amount of change, namely 20-50%. range of the total amount of change, is reached is obtained. A constant time width ( T) before and after the time T is used as a data processing range and optical output in this range is processed.

Description

[Detailed description of the invention]

0001

BACKGROUND OF THE INVENTION 1. Field of the Invention The present invention relates to a method for measuring blood coagulation ability by mixing plasma and a coagulation reagent to precipitate fibrin and measuring blood coagulation time using optical means.

0002

BACKGROUND OF THE INVENTION A blood coagulation test is an important test for determining whether blood coagulation factors exist normally in blood and whether they work effectively.

[0003] Blood maintains its fluidity inside blood vessels and flows therein, but once bleeding occurs, the bleeding is stopped by the action of the blood vessels, platelets, and coagulation factors in the blood. This is because various coagulation factors present in plasma and platelets are activated in a chain manner, and fibrinogen in plasma is converted to fibrin and precipitated.

[0004] Therefore, when coagulation factors are absent or insufficient in the blood, problems arise in that blood coagulation is insufficient and bleeding cannot be stopped or hemostasis is delayed. Therefore, it is important to measure blood coagulation time in advance, especially during surgical operations.

Measurement items related to blood coagulation time include prothrombin time (PT), activated partial thromboplastin time (APTT), and fibrinogen level (Fb).
g) and Ca re-addition time. These items are
It depends on which component in the blood is reacted with the reagent added. For example, in the measurement of prothrombin time (PT), when sufficient amounts of tissue thromboplastin and calcium are added to plasma in blood components after centrifugation,
Measure the time it takes for white reticular fibrin to precipitate,
As another example, the activated partial thromboplastin time (A
In measuring PTT, an activated partial thromboplastin reagent is added to plasma collected after centrifugation, and the time until fibrin is precipitated is measured.

[0006] The above measurement times differ only in the reagents that induce coagulation, and are all measured by observing the phenomenon in which fibrinogen in plasma is converted to fibrin by the action of coagulation factors. Typically, these times are defined as the time from the time the reagent is mixed with the plasma until fibrin begins to precipitate.

As mentioned above, although the method for measuring blood coagulation time has basically been established, it is quite difficult to grasp the fibrin precipitation initiation state with good reproducibility during actual measurement. To this end, various methods for measuring blood coagulation time using optical techniques have been proposed.

For example, Japanese Patent Publication No. 61-10777 discloses a method of measuring the scattered light intensity of a mixture of plasma and a coagulation reagent, and measuring the blood coagulation time based on the time when the rate of change of the scattered light intensity reaches its maximum. Are listed. In this method, the scattered light intensity is differentiated in real time, but when measuring blood coagulation time, the scattered light signal is disturbed by the presence of precipitates or air bubbles that are unrelated to the original coagulation reaction. This often results in errors in measured values.

Furthermore, Japanese Patent Application Laid-Open No. 59-203959 discloses that based on the time when the amount of change in the scattered light intensity exceeds a certain amount, or the amount of change that exceeds a certain percentage of the total amount of change up to the solidification end point, A method for measuring blood clotting time is described. However, in reality, the fibrinogen concentration in the specimen varies, and because of these differences, the amount of change is not uniform.Also, changes in the scattered light intensity occur due to shrinkage of the fibrin clot that occurs following fibrin precipitation. Since the amount of change at the end point differs significantly, there is a problem in that the measured values include errors even in this method.

FIG. 1 shows an ideal curve when measuring scattered light intensity, which is an example of optical output, in measuring blood coagulation time. This curve represents the behavior in which the amount of scattered light in plasma changes over time due to the coagulation reaction. In "Figure 1", part a indicates the state where the reaction is progressing in the mixture of test plasma and reagent, but fibrin precipitation has not yet been reached, and point b indicates the start of fibrin precipitation. It is understood that point c indicates a state in which fibrin precipitation is most active, and point d indicates a state in which most of the fibrinogen in the plasma has been converted to fibrin. Further, "FIG. 1" also shows a curve obtained by differentiating the scattered light intensity with respect to time.

[0011] However, the curve shown in "Figure 1" is an idealized version of the curve actually measured, and the curve actually obtained is one that can clearly distinguish the state of plasma as described above. Instead, it is often extremely difficult to judge the state of plasma as a result of abnormal reactions occurring or noise being included.

FIG. 2 shows an example of changes in the intensity of scattered light when measuring blood coagulation time. In this case, for example, noise may occur before the plasma and coagulation-inducing reagent are mixed at time 0, fibrin is precipitated by the reaction, and the scattered light intensity increases (portion C where the scattered light intensity finally rises). There are parts (peak A and peak B) where the scattered light intensity temporarily increases.

[0013] "Figure 3" is the result of differentiating the scattered light intensity of "Figure 2" with respect to time, and peak A', peak B', peak C
There are three types such as ' (for simplicity, the negative peak is ignored). Since it is known from experience that fibrin does not precipitate for a while after adding a coagulation-inducing agent, data should not be measured during this period and peaks A or A'
can be excluded from the measurement target.

[0014] Furthermore, it is also possible to set an appropriate threshold value and perform measurement only when the scattered light intensity increases to a certain level or more. However, as mentioned above, there are individual differences in the fibrinogen concentration in the specimen, and as a result, the size of the peak and the amount of change in scattered light intensity (or change in scattered light intensity) differ, so if the threshold value is set too large, the sensitivity will deteriorate.
Even desired peaks (or changes in scattered light intensity) may be eliminated. Conversely, if the threshold value is made small, extra peaks such as noise will also be measured. In order to solve this problem and avoid missing the originally required peak due to fibrin precipitation, there is a method of setting a threshold value that decreases over time until a certain period of time, as shown by the broken line in Figure 3. Conceivable.

However, as shown in the figure, noise such as peak B or B' increases the original scattered light intensity (C).
It is impossible to eliminate such peaks using conventional methods such as those described above. For example, in the method described in Japanese Patent Publication No. 61-10777, it may be determined that the amount of change is the largest at peak B', and the blood coagulation time may be calculated based on this.

[0016] Furthermore, based on the total amount of change in the final scattered light intensity, the time when the scattered light intensity changes by a certain percentage is determined to be the time when fibrin has precipitated. Even if the method described above is adopted, as can be clearly seen from "Figure 2", the amount of change in peak A and peak B is quite large, even if temporarily, so these In some cases, it may be determined that a blood coagulation time exists around the time when the peak occurs.

[0017]

[Problem to be Solved by the Invention] As mentioned above, when measuring blood coagulation time using an optical method, it is necessary to judge the validity of each data (peak or state of change) generated by the optical method. Otherwise, the reliability of the measurements obtained cannot be improved. Therefore, the problem to be solved by the present invention is to provide a method for measuring blood coagulation time that makes it possible to judge the validity of data obtained by optical methods.

[0018]

[Means for Solving the Problems] The above-mentioned problem is solved by a method for measuring blood coagulation time using an optical method, which is the first aspect of the present invention. ) and the total amount of change (ΔY) in the optical output during that time, and determine the time T when a predetermined amount of change within the range of 20 to 50% of the total amount of change is reached. It has been found that the problem can be solved by a blood coagulation time measuring method characterized in that a constant time width (ΔT) before and after time T is set as a data processing range and optical output in this range is processed.

The optical method used in the method of the present invention is to detect the precipitation of fibrin in the plasma by irradiating the plasma with light and measuring the amount of transmitted light or scattered light or changes in the intensity thereof. This is a detection method, and is described in, for example, Japanese Patent Publication No. 10777/1983. Specifically, it is a method in which one of the above-mentioned coagulation-inducing substances is added to plasma, irradiated with light, and the turbidity of the plasma is measured over time by the amount of scattered light or transmitted light.

As described in the above-mentioned publication, when measuring using scattered light and measuring using transmitted light, the output data is qualitatively symmetrical, and the absolute They just have different values. Therefore, the method of the present invention when using scattered light can be similarly applied when using transmitted light. The present invention will be described below using an example of using scattered light.

[0021] The fixed time (To) for measuring the change in optical output over time is determined depending on the type of coagulation time to be measured, the conditions of the coagulation reaction, the processing method of the obtained optical output, etc. It is. In the present invention, basically, it is possible to adopt the empirically known time at which the coagulation reaction of most specimens is considered to be almost completely completed, and does not necessarily have to be completely completed. For example, prothrombin time measurement is about 60 seconds, activated partial thromboplastin time is about 200 seconds, and fibrinogen content is about 5 seconds.
It is sufficient to set it to 0 seconds.

Therefore, the coagulation reaction is almost completed within this time, and the difference between the scattered light intensity before the start of coagulation and the scattered light intensity after the completion of coagulation, that is, the total amount of change in the scattered light intensity (ΔY,
(see “Figure 2”) is calculated. It is empirically known that the time when the fibrin production rate reaches its maximum exists around the time when the amount of change reaches 20 to 50% of the total amount of change. Therefore, the time (T) at which a predetermined amount of change within the range of 20 to 50% of the total amount of change is reached is found retroactively from the end point side of the reaction, and a predetermined time before and after that time is cut out as the data processing range. By processing the optical output data over a range, eg, differentiating the optical output with respect to time, the time at which the fibrin deposition rate is maximum can be estimated.

The predetermined amount of change of 20 to 50% of the total amount of change can be appropriately selected depending on the measurement item, but generally about 30% is sufficient, and basically data processing The selection is made so that the maximum value of the time differential value of the scattered light intensity is included within the range.

Therefore, an appropriate time width (ΔT) extending before and after time T (that is, from time T−ΔT1 to time T+ΔT2
Therefore, fibrin actually precipitates within the time (ΔT = ΔT1 + ΔT2) (the rising part at the right end in "Figure 2", C), and the time differential value of the measured optical output (data) a peak associated with fibrin precipitation (peak C in “Figure 3”)
') is presumed to exist. Therefore, in the method of the present invention, peaks with a small proportion (20% or less) of the total variation are eliminated, and peaks with a large proportion (20% or more) of the total variation are eliminated. Also, by tracing back from the end point of the reaction, unnecessary peaks (peaks that appear before the necessary peaks that are not caused by fibrin precipitation) are detected.
is excluded. Therefore, peak A' and peak B' in "FIG. 2" are excluded from the measurement target. Also, depending on the target item to be measured, etc., the time width Δ extending before and after time T
T can be varied as appropriate, and ΔT1 and ΔT2 can be the same or different.

Further, ΔT can be empirically defined as a temporary function of T, and ΔT1 and ΔT2 are each expressed by the following equations. ΔT1=a1T+b1 ΔT2=a2T+b2 a1=0.1~0.3 b1=2.0~6.0 a2=0.1~0.3 b2=2.0~6.0

By appropriately setting the predetermined amount of change and ΔT as described above, it becomes possible to focus on the time period in which fibrin is actually deposited, and even when fibrin is not actually deposited, First, it is possible to eliminate signals caused by noise, etc., which may cause a false recognition as if precipitation is occurring. Therefore, it is sufficient to set the data processing range from time T-ΔT1 to time T+ΔT2, and process optical signals during this period.

For example, when applying the method described in Japanese Patent Publication No. 61-10777, the time (T) at which fibrin is actually deposited is specified by the method of the present invention, and the time data in the vicinity of that time is used. The scattered light intensity is differentiated with respect to time as the processing range (ΔT), and the point where the differential value is maximum is determined to obtain the time (Tm) at which the fibrin precipitation rate is the highest.

As mentioned above, 1/m(
m is a predetermined number greater than 1), and there are times before time T within the data processing range (that is, there are two times before and after time T at which the differential value is 1/m). The end point of the blood coagulation time is defined as the time (from the time of addition of the blood coagulation inducing reagent) until the time when the blood coagulation inducing reagent is added. The value of m is generally about 2 to 4. The end point determined in this way is closer to the starting point of fibrin precipitation than conventional visual methods. Moreover, this method has very good reproducibility, and the value has a certain relationship with the value determined by visual inspection by an expert, so the concept of clotting time, which has traditionally been used for diagnostic purposes, is can be easily applied to

Even after determining the data processing range using the method of the present invention, there may be unreasonable peaks within the processing range due to noise or the like. It is desirable to be able to determine appropriate peaks even in such cases. Therefore, the present invention also provides a method for determining the validity of a peak after specifying the data processing range as described above in the second aspect.

The second gist of the present invention, which involves determining a data processing range by the method of the present invention and then determining a more appropriate peak within that range, will be described below with reference to a specific example.

In the data processing range extracted by the method of the present invention, noise and peaks due to fibrin deposition may be close to each other. Examples are shown in "FIG. 4" and "FIG. 5.""FIG.4" shows the scattered light intensity, and "FIG. 5" shows its time differential value. In this case, the conventional method of determining based on the maximum differential value (Japanese Patent Publication No. 10777/1983) ends up determining that the peak F' is a valid peak. However, as explained above, it is empirically known that the maximum value of the time differential value of the scattered light intensity due to fibrin precipitation occurs at a portion of about 20 to 50% of the total change in the scattered light intensity. Therefore, the amount of change is about 40
It can be seen that the peak G', which is about %, is a reasonable peak.

[0032] Compared to the conventional method, which processes data over a fairly wide range without any particular limitations, the method of the present invention reduces the number of unnecessary peaks by specifying the data processing range. This makes it possible to reduce the number of peaks whose validity should be determined. Furthermore, since the validity of the remaining peaks can also be determined, the number of peaks that are actually subject to data processing is further reduced, allowing for more reliable measurements.

As mentioned above, select the data processing range,
Even if the validity of the peaks included in the range is determined and a valid peak is selected, the target peak may partially overlap with other unnecessary peaks, resulting in a normal peak shape. You may not be able to get it. In this case, if the method described in Japanese Patent Publication No. 61-10777 is applied, the obtained results may contain errors.

For example, measurement results such as those shown in FIG. 6 showing the differential value of the scattered light intensity may be obtained. In "Figure 6", the amount of change in the scattered light intensity at peak I' is about 10% of the total amount of change, about 30% for peak J', and about 30% for peak K.
' is about 70%. Therefore, as described above, it can be determined that the peak J' is an appropriate peak that should originally be measured from the viewpoint of the amount of change relative to the total amount of change.

However, in the case of "FIG. 6", even if a reasonable peak is determined to be J', if the method of the prior art is adopted, blood will be detected at some time during the rising portion of peak I'. It will be determined that a coagulation time exists (for example, Tc in "FIG. 6"). Original peak J'
If only, the curve should be as shown by the broken line in "FIG. 6", and the blood coagulation time should be longer than Tc, for example, Te in "FIG. 6".

[0036] In order to solve such problems, a standard peak pattern (shape) (that is, no unnecessary peaks are superimposed) and an actual peak pattern (shape) are used.
By comparing the peaks with 6) can be estimated.

The original pattern in case of blood coagulation, namely:
A standard pattern to be measured in the absence of noise etc. is determined in advance according to the item to be tested, coagulation time, fibrinogen concentration, etc. For example, various patterns as shown in "FIG. 7" are obtained. After that, if the actually measured data has a pattern as shown in "Figure 6", the rising part of peak J' is estimated from the selected standard peak (for example, the leftmost standard peak in "Figure 7") as shown by the broken line. do. Based on this estimated pattern, the blood coagulation time Te is determined by a conventional method.

This standard peak pattern is estimated based on the ratio of blood coagulation time (Tc) obtained by various methods to the time when the blood coagulation rate reaches its maximum (Tm) and the shape near the top of the peak. . That is, the measured T
If c/Tm is significantly different from the Tc/Tm value of the standard pattern, it is determined that the peak shape is abnormal for some reason, and then the shape near the top of the peak is measured. Select standard peaks with similar shapes. The Tc corresponding to this standard peak Tm is determined and taken as the blood coagulation time Te.

Selection of the data processing range of the present invention as described above, subsequent data processing, and determination of peak validity;
Estimation of blood coagulation time based on estimation of peak shape can be easily programmed as long as the method described above is provided. Accordingly, the present invention also provides a blood coagulation time measuring device that includes a data processing device incorporating a program capable of implementing the method described above.

An apparatus for carrying out the method of the present invention is schematically shown in FIG. "FIG. 8" is a block diagram of the blood coagulation time measuring device according to the present invention, in which a pipette 4 containing a reagent 3 is placed above a cell body 2 containing a mixture 1 of test plasma and a blood coagulation-inducing reagent. A light source lamp 5 and a condensing lens 6 are arranged below the cell body 2 as means for irradiating the mixed liquid 1 in the cell body 2 with a constant amount of light beam, and on the side of the cell body 2, the mixed liquid 1 is placed. A photodetector 7 that measures the amount of scattered light and converts it into an electrical signal is arranged, and a signal amplifier 8 that amplifies the scattered light output signal from the photodetector 7 converts the amplified scattered light output signal into a digital signal. The microcomputer 10 is an arithmetic device that inputs and processes signals from the A-D converter 9, and a printer 11 is an output device that displays measurement and arithmetic results.

First, the light emitted from the light source lamp 5 is made into a luminous flux of a constant amount by the condensing lens 6, passes through the bottom of the cell body 2, and irradiates the mixed liquid 1 in the cell body 2, so that a part of the light is mixed. The light is scattered by the liquid 1 and enters the photodetector 7. In this case, the light source lamp 5 may be a tungsten lamp or the like, and any light source system that emits a stable light beam with a constant amount of light may be used without using the condenser lens 6, and the photodetector 7 may include a phototube, Any type of photovoltaic cell or other device that converts light into an electrical signal can be used. In addition, cell body 2
A test tube or other cylindrical body with a bottom made of a transparent material is used, and in order to measure scattered light, it is preferable to use a well-shaped one without scratches. In the illustrated example, the photodetector 7
is attached in a direction of 90 degrees with respect to the optical axis of the light emitted by the light source 5, but there is no need to particularly limit the attachment angle since it is sufficient to receive scattered light.

Next, the A-D converter 9 receives the scattered light output signal M obtained by amplifying the electric signal continuously emitted by the photodetector 7 by the signal amplifier 8, samples it at regular time intervals, and converts it into a digital signal. A signal N is input to the microcomputer 10 at the same periodic intervals.

The microcomputer 10 measures the blood coagulation time using the programmed method of the present invention described above, and transmits the result to the printer 11 to print it out. In addition, "Figure 8" shows the measurement start switch 1.
2 is also illustrated.

In the illustrated embodiment, the microcomputer 10 is used as the arithmetic unit, but it is also possible to design a dedicated arithmetic control circuit using an IC-based CPU. Furthermore, arithmetic device measurement start switch 1
2, a pipette with a switch that is linked to the addition operation of the pipette that adds the reagent 3 to the cell body 2 can be used, or in the case of an automatic measuring device, a switch that is linked to the reagent extrusion operation in the nozzle can be used.

Furthermore, the method of the present invention described above is disclosed in Japanese Patent Application Laid-Open No.
It can also be applied to the method for measuring blood coagulation time described in Japanese Patent No. -203959. In this method, since the final total amount of change is used as a reference, the constant time To in the first aspect of the present invention is
must be the time required for the coagulation reaction to complete. Furthermore, the blood coagulation time may be defined as a time that includes the time T at which a predetermined amount of change in the range of 20 to 50% of the total amount of change is reached and shows a predetermined amount of change in the curve of the scattered light intensity before and after that time.

[Brief explanation of the drawing]

[FIG. 1] "FIG. 1" is a schematic diagram showing an idealized measurement result of scattered light intensity in blood coagulation time measurement.

[FIG. 2] "FIG. 2" is a diagram schematically showing the measurement results of scattered light intensity.

[FIG. 3] "FIG. 3" shows the result of differentiating the scattered light intensity of "FIG. 1" with respect to time, and "'" is attached to the alphabet to indicate that the differential relationship corresponds.

FIG. 4 is a diagram schematically showing the scattered light intensity measured when noise is included within the data processing range selected by the method of the present invention.

[Figure 5] "Figure 5" shows the result of differentiating the scattered light intensity of "Figure 4" with respect to time, and "'" indicates "Figure 3".
It is similar to

[FIG. 6] "FIG. 6" is a schematic diagram when an unnecessary peak is superimposed on the peak to be measured when the scattered light intensity is differentiated with respect to time.

FIG. 7 is a diagram showing an example of a standard peak for estimating the original peak shape.

FIG. 8 is a block diagram of a blood coagulation time measurement device that implements the blood coagulation time measurement method of the present invention.

[Explanation of symbols]

1... Mixed liquid, 2... Cell body, 3... Blood coagulation reagent, 4... Pipette, 5... Light source lamp, 6... Condensing lens, 7... Photodetector, 8... Signal amplifier, 9... A-D converter, 10 ...Microcomputer, 11...Printer, 12...Switch for starting measurement.

Claims (3)

[Claims] Claim 1: In a method for measuring blood coagulation time using an optical method, a temporal change in optical output within a certain time (To) after addition of a blood coagulation-inducing substance to plasma, and a total amount of change in optical output during that period. (ΔY) and 2 of the total amount of change.
The method is characterized in that the time T at which a predetermined amount of change within the range of 0 to 50% is reached is determined, and the optical output in this range is processed using a constant time width (ΔT) before and after time T as a data processing range. A method for measuring blood clotting time. [Claim 2] The processing of the optical output is carried out at a time of 1/m (m is a predetermined value of 1 2. The method according to claim 1, wherein the blood coagulation time is defined as a time before time T within the data processing range, which shows a differential value of . 3. The method for measuring blood coagulation time according to claim 2, wherein when at least two maximum values exist, one maximum value is selected based on the ratio of optical output in the maximum values. .