JPS6259772B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a method for processing differential images of serum obtained using an electrophoresis device.

The basic electrophoretic image pattern obtained by electrophoresing human serum using an electrophoresis device using a cellulose acetate membrane as a support and then measuring the light using a densitometer (generally, this is the pattern for healthy people) is a concentration distribution as shown in FIG. Such an electrophoretic image pattern usually consists of five peaks, A, B, C, and N, located at positions corresponding to a ₀ , a ₁ , a ₂ , a ₃ , and a ₄ on the x-axis shown in Figure 1. , 5 fractions of e, namely albumin, α ₁ globulin, α
₂ globulin, β globulin, γ globulin 5
Fractionate into fractions. Then, a determination of normality and abnormality is made from such an analog diagram and the percentage value for each fraction. However, in the electrophoresis pattern obtained by photometry of the partial images obtained by actually performing electrophoresis using the specimen to be measured, peaks may occur due to various causes in addition to the peaks shown in FIG. 1. For example, the one shown in FIG. 2 has a peak at a portion _a5 in addition to the five peaks. This occurs when serum is cloudy, and is the result of substances that are not migrated after application and are fractionated as traces of application. Also, the one shown in Figure 3 has the _fourth
The one shown in the figure has a peak at position _a7 . These occur depending on the freshness of the serum, and are caused by the fractionation of certain components in the serum; Fig. 3 shows the result of β lipoprotein, and Fig. 4 shows the result of β ₁ c globulin.

If a peak appears in a portion other than the peak of the basic electrophoresis pattern as described above, it will be inconvenient when performing colorimetric photometry and automatically processing the data on a computer.

An example of a densitometer and photometric device currently in use is shown in Figure 5.
Light from a light source 3 passes through a lens 4, a filter 5, and a slit 6, and is applied to a support 1, which will be described later, and is further detected by a light receiving element 7. Separate images 2, 2', 2'', etc. of serum are formed on the support 1 as shown in FIG. Photometry is performed by scanning in a direction perpendicular to the direction of movement of the body.In other words, the light from the light source that passes through the specimen on the support is received by the light receiving element 7, and the output from the light receiving element according to the concentration of the specimen is sent to the preamplifier 8. The amplification and logarithm conversion unit 9 converts the value into a logarithm value, and based on this value, an analog pattern as shown in Fig. 1 is obtained.The output from the logarithm conversion unit 9 is also converted to an A/D.
input to the converting section 10 and the computer section 12
The conversion command signal generating section 11 is operated according to the photometry command 11a from the photometry command 11a, and digital conversion is performed at regular time intervals. Based on the digital data obtained here, the percentage value of each fraction is determined.

In the above operation, in the case of an electrophoretic image pattern as shown in FIG. 1, it is sufficient to find each minimum value point as a separation point and find the % value between them. However, Figures 2 to 4
In the case of an electrophoresis pattern like the one shown in the figure, it is not possible to obtain values for 5 fractions. Therefore, if there are 6 or more fractions, the person in charge of the test will look at the analog pattern and the electrophoresis image and select albumin, _α1 globulin, _α2 globulin, β globulin, or γ globulin to make the 5th fraction. I had to recalculate by including it. Alternatively, if the fractionation abnormality is due to a disease, it may be reported as a fractionation abnormality without treatment.

This was invented based on the above circumstances, and even in the case of electrophoretic image patterns that are fractionated into 6 or more fractions, a computer automatically processes to include peak values other than the above-mentioned peak values in the 5 fractions. A patent application was filed in 1984 as a treatment method that could be carried out on a regular basis.
There is a method described in No. 64814 (see Japanese Patent Publication No. 61-16019).

The method (hereinafter referred to as the first conventional method) is briefly explained as follows.

First, a standard serum such as a commercially available control serum is run, and then photometry is performed using a densitometer to obtain a standard pattern, which is a migration pattern of 5 fractions. The peak position and fractionation point of the electrophoresis pattern are approximately determined by the type of support and the electrophoresis conditions. Therefore, if the type of support and the electrophoresis conditions are the same, the development length of the electrophoresis pattern of the serum to be tested is almost the same as that of the standard serum.

The standard length of this standard serum (the length from the starting point etc. to the peak value or fractionation point) is determined as follows. When the electrophoretic image is photometered, an analog pattern as shown in FIG. 7 is obtained. This is sampled at regular time intervals, A/D conversion is performed, and the density at the sampling point is stored.

Concentration (digital value) at each sampling point
is as follows when considered in correspondence with the analog pattern shown in FIG. x as shown in Figure 7
Considering the axes and the y-axis, sampling at the above-mentioned intervals starts from the origin _a0 on the x-axis, and each point at each interval is a respective sampling point. Therefore, each stored density corresponds to the y value at each sample point in FIG. Detection of fractionation points will be explained based on the concentration at each of these sample points. Since the dividing point is a point on the x-axis that corresponds to the valley of the analog pattern, a sampling point x _{b -1} where an arbitrary sampling point on the x-axis is x b and the value of y at that sampling point is y _b The y value at sampling point x _{b+1 is y b-1} , and the y value at sampling point x _b+1 is y _b+1
Then, a sampling point x _b having a value of y _b that satisfies the following relationship is a dividing point.

y _b < y _b-1 , y _b < y _b+1 Next, the peak position is from the starting point x ₀ to the fractionation point
a ₀ is between b ₁ , the dividing point b ₁ , a ₁ is between b ₂ , the dividing point
There is a ₂ between b ₂ and b ₃ , a ₃ between dividing points b ₃ and b ₄ , and a ₄ from dividing point b ₄ to the end point x _o , and these points are divided into This is the x _a point that has a value of y _a that satisfies the following relationship in the same way as the points were found.

y _a > y _a-1 , y _a > y _a+1 The values of b ₁ , b ₂ ..., a ₀ , a ₁ ..., etc. obtained in this way are the length from the starting point x ₀ to each point ( (length on the x-axis), with a one-to-one correspondence. Therefore, these coordinates (coordinates in units of fixed time intervals) may be used instead of the length from the starting point.

In this manner, the fractionation of the standard serum is completed.

Next, the fractionation point is determined using the method described above for the pattern obtained by electrophoresing the sample to be measured. Next, a point a ₀ on the x-axis corresponding to the peak point of the standard serum,
Using a ₁ , a ₂ , and a ₃ as reference points, the same values are positioned on the x-axis of the pattern of the object to be measured as shown in FIG. In this sample pattern, a ₀ a ₁
Count the number of dividing points (points on the x-axis corresponding to the minimum values that become valleys) in each interval between a ₁ a ₂ , a ₂ a ₃ , and a ₃ a ₄ . If the number is 1, that fractionation point is set as the fractionation point, and if the number is 2 or more, the one with the lowest concentration is determined as the fractionation point among the fractionation points, and the others are eliminated. For example, in Figure 8, the space between a ₀ a ₁ and a ₂ a ₃ are both one fractional point (b ₁ and b ₃ ), so these are defined as the first and third fractional points, respectively. , between a ₁ a ₂ is b ₅ , b ₂ ,
Since there are 3 pieces of b ₆ , the smallest of them, b _2, is taken as the dividing point, and between a ₃ a ₄ , there are 2 pieces of dividing points, b ₇ and b ₄ , so b ₄ is taken as the fourth dividing point. Furthermore, all fractional points after a ₄ (b ₈ in FIG. 8) are eliminated.

In other words, fractionation points due to β-lipoprotein, β ₁ c protein, and foreign matter at the coating point always take higher values than the normal 5-fractionation points. Therefore, according to the above processing method, it is possible to perform 5-fraction processing.

The above explanation uses the starting point as the origin, but the first
The origin may be a ₀ corresponding to the peak of .

However, this first conventional method requires a standard serum that always exhibits a normal fraction. Some commercially available standard sera do not necessarily provide normal fractionation, making selection of standard sera troublesome. In addition, the specimen is affected by the conditions of the place where it is stored, and if it is contaminated with bacteria, it may not be fractionated into the correct five fractions. Therefore, storage of standard serum is a very complicated problem.

A fractionation processing method in electrophoresis was proposed that utilizes the first conventional method mentioned above and also allows determining the correct fractionation position without using standard serum. (see Publication No.).

Next, this method (hereinafter referred to as the second conventional method) will be explained. FIG. 9 is a block diagram of a mechanism for performing fractionation processing using the first conventional method, in which a standard serum is measured by an electrophoretic measuring mechanism 21, and the measured value is sent to a fraction determining mechanism 22 using the method described above. The x-coordinate position of the peak (a ₀ , a ₁ , a ₂ , a ₃ , a ₄ ) and the fraction position (b ₁ , b ₂ ,
b ₃ , b ₄ ) are sent to the reference position storage mechanism 23 and stored therein. Next, the sample to be measured is measured by the electrophoretic image measurement mechanism 21, and the x-coordinate values of the maximum value, minimum value, etc. are determined from the measured values by the fractionation position determination mechanism 22.
The obtained value is compared with the value of standard serum stored in advance by the 5-fraction processing mechanism 24, the correct fraction position is determined by the method described above, and the fraction data is calculated based on this fraction data output. is obtained.

The second conventional method does not use standard serum as in the first conventional method, but instead determines the reference position using the data of a standard pattern that has been successfully divided into five fractions of the sample to be measured. This is a 5-fraction process. For example, it is known that the integrated value of the concentration of each fraction of serum from a normal person falls within a certain range. The second conventional method focuses on this point and determines whether the sample to be measured is in 5 fractions and whether the concentration integral value or the ratio between one fraction and other fractions falls within a certain range. It is determined whether the sample is a fraction, a reference position is determined based on what is determined to be a normal fraction, and the fraction position of the sample to be measured is determined by referring to this.

The second conventional method will be explained based on the block diagram of FIG.
The fact that the coordinate values are determined by the fractional position determination mechanism 22 is the same as in the case of FIG. Subsequently, a normal fraction determining mechanism 25 determines whether the value obtained by the fraction position determining mechanism 22 is a normal 5 fraction. It is 5 fractions, and each valley position (b ₁ , b ₂ , b ₃ ,
b ₄ ) A method in which the integrated value of the concentration of each fraction classified according to 4) is determined, and whether or not it is a normal fraction is determined based on whether or not these integrated values fall within a preset range. According to etc. Only the data of local maximum values and local minimum values determined to be normal by this normal fraction determination mechanism 25 is sent to the next reference position calculation mechanism 26, and an appropriate reference position is calculated by appropriate statistical processing. . The reference position calculated here is sent to the next reference position storage mechanism 27 and stored therein. The reference position stored here is the first conventional method (Figure 9)
This corresponds to the reference position of the standard serum stored in the reference position storage mechanism 23 of . Next, in the same way as the method shown in FIG. 9, the data from the fraction position determining mechanism for the sample to be measured is sent to the 5-fraction processing mechanism 24 through the switching mechanism 28. The 5-fraction processing mechanism 24 outputs data that has been fractionated with the correct fraction value, and calculates the integrated value of the concentration of each fraction. As described above, the reference position is determined using the test specimen itself without using standard serum, and this is used for the fractionation process.

The purpose of the present invention is to process the analog pattern of a sample that is not normally fractionated into the correct 5 fractions. The present invention provides a fractionation processing method in electrophoresis.

In the fractionation processing method of the present invention, a sampling point corresponding to a peak concentration value (a point on the x-axis) or a sampling point corresponding to a valley concentration value in standard serum is used as a reference position, and the distance between this reference position and the analyte to be measured is The correct demarcation point is determined by comparing the sampling point with the minimum value or peak value in the analog pattern.

First, the fractionation processing method of the present invention will be specifically explained for a specimen having an analog pattern as shown in FIG. In other words, a specimen having more fraction points than regular fraction points will be explained.

For a specimen such as that shown in FIG. 11, a sampling point having a minimum value is determined by the method already explained. In this figure, the point _a0 on the x-axis, which corresponds to the peak value of the albumin fraction, is selected as the origin. And the sampling point corresponding to the minimum value (point on the x-axis)
are respectively b′ ₁ , b′ ₂ , b′ ₃ , and b′ ₄ . Furthermore, in the case of this specimen, there is a minimum value due to the abnormal fraction, so these points on the x-axis are designated b' ₅ and b' ₆ . On this analog pattern of the sample to be measured, each standard position b ₁ , b ₂ , b ₃ , b ₄ is positioned with the point a ₀ of the standard sample aligned with the origin. Each reference position b ₁ , b ₂ , on the x-axis in FIG.
Compare b ₃ , b ₄ with the positions b′ ₁ , b′ ₂ , b′ ₃ , b′ ₄ , b′ ₅ , b′ ₆ on the x-axis corresponding to the x-axis corresponding to the minimum value and find the standard. The position closest to the position is determined as the dividing point.
In Figure 11, the positions near each reference position are
b′ ₁ , b′ ₂ , b′ ₃ , b′ ₄ , these are the dividing points, and b′ ₅
and b′ ₆ are far from any reference position, so these points are deleted. As a result, abnormal fraction points are deleted, and processing can be performed using the correct five fractions.

As described above, for samples with five fractions or more, fraction points other than the normal fraction points are deleted because they are located further away from the reference position. Therefore, samples with patterns as shown in FIGS. 2 to 4 can also be correctly fractionated.

Next, a specimen with four fractions or less as shown in FIG. 12 will be explained. In FIG. 12, the points b'' ₁ , b'' ₃ , b'' ₄ on the x-axis corresponding to the minimum values are determined as described above. Also, the reference positions b ₁ , b ₂ , b ₃ and b ₄ are set on the x-axis of the pattern shown in Figure 12.By this, the position corresponding to the minimum value near these reference positions is found and this is set as the dividing point.In this example, In this case, there is no position close to the reference position b _2. In this way, if there is no fractionation point, the reference value positioned on the pattern of the sample to be measured is added as a fractionation point.This allows for 5 fractions. processing becomes possible.

In this case, there is a problem as to whether the newly added fractionation point is actually the correct fractionation point. However, as a feature of electrophoresis, the development length of each fraction is approximately constant if the electrophoresis time and other electrophoresis conditions are kept constant. Therefore, if a pattern obtained by forming fractional images under the same electrophoresis conditions as the specimen to be measured is used as a standard specimen pattern, almost correct fractionation positions will be obtained.

The fractionation processing method of the present invention explained above can be automatically processed by a computer. In other words, the x-coordinate values of the points on the x-axis corresponding to the minimum values of the standard sample and the sample to be measured can be determined by employing the first conventional method. In addition, the method of finding the reference position on the pattern of the sample to be measured is to set the origin of the standard sample and the sample to be measured at the same point (for example, the point on the x-axis corresponding to the first peak), then the reference position of the standard sample can be found. Since the x-coordinate values correspond to the same x-coordinate values of the specimen to be measured, finding the x-coordinate of the reference position is the same as positioning it on the pattern of the specimen to be measured. Furthermore, the method of selecting a normal demarcation point among the positions on the x-axis corresponding to the minimum value is to find the difference between these values and the value of the reference position, and then find the smallest one among them. Good. In this case, in the case of a sample with 6 or more fractions, the incorrect fractionation point is always farther away from the reference position compared to the correct fractionation point, so the difference is greater than the difference between the correct fractionation point and the reference position. Therefore, it will not be a dividing point. In addition, in the case of 4 fractions or less, the difference from the reference position corresponding to the undetected fraction point is larger than the difference between other reference positions and the fraction points close to it, so the fraction corresponding to that reference position is It is immediately apparent that no pixel exists. Then, as described above, the reference position itself is added as a dividing point.

In the above explanation, the points on the x-axis corresponding to the first peak value of the standard sample (peak value of albumin) and the point on the x-axis corresponding to the first peak value of the sample to be measured are selected as the origin, respectively. Fractionation was performed after matching the two. In other words, each first peak value is y
After selecting the axes and matching them, fractionation processing was performed.

In addition to the above method, the start position of the analog pattern of the specimen may be selected as the origin. In other words, select c ₀ in Fig. 7 as the origin and then set the values on the x-axis c ₁ , c ₂ ,
Set c ₃ , c ₄ , etc. to values corresponding to the reference position or minimum value, and use the same method as above to calculate the difference between the values of the standard specimen c ₁ , c ₂ , c ₃ , c ₄ and the value of the measured specimen. Fractionation treatment may also be performed.

In addition, in the above method, the reference position must be determined using a standard sample every time each sample is fractionated. However, this method is, so to speak, the first conventional method (Japanese Patent Application No. 1983-
64814) in a completely different method, the present invention is applied to the sample fractionation processing means in the first stage of the second conventional method (invention of Japanese Patent Application No. 146,836/1983). By using the method described above, it is also possible to adopt the second conventional method and perform fractionation processing on the analyte to be measured without constantly performing photometry and fractionation processing on the standard specimen.

As explained above, according to the fractionation processing method of the present invention, accurate fractionation processing can be automatically performed by a computer. Furthermore, fractionation processing is also possible for specimens with four fractions or less.

[Brief explanation of the drawing]

Figure 1 is an analog pattern of a normal specimen, Figure 2
Figures 4 to 4 are analog patterns containing abnormal fractions, Figure 5 is a diagram showing the configuration of a commonly used densitometer, Figure 6 is a diagram showing the position of the specimen on the support, and Figure 4 is a diagram showing the configuration of a commonly used densitometer. 7 and 8 are explanatory diagrams of the first conventional method, FIG. 9 is a block diagram of the first conventional method, and FIG. 10 is a block diagram of the second conventional method.
FIG. 11 and FIG. 12 are explanatory diagrams of the processing method of the present invention, respectively.

Claims (1)

[Scope of Claims] 1. A step of determining a minimum value of a reference pattern and storing it as a reference position, a step of associating the reference position with a pattern obtained by electrophoresing a specimen to be measured, and a step of determining the minimum value of a reference pattern. The minimum value of the obtained pattern is determined and compared with the reference position, and the closest minimum value is treated as a dividing point and the others are not treated as dividing points. A method for fractionation processing in electrophoresis, which comprises the step of treating a reference position as a fractionation point when it does not exist. 2. The fractionation processing method in electrophoresis according to claim 1, wherein the reference pattern is obtained by electrophoresing a standard serum such as a control serum. 3. The method of fractionation processing in electrophoresis according to claim 1, wherein the reference pattern is obtained from a sample that shows a normal fraction after electrophoresis of the analyte to be measured.