JPH0259955B2 - - Google Patents

Description

DETAILED DESCRIPTION OF THE INVENTION The present invention relates to a signal processing method in autoradiography. More specifically, the present invention relates to a method for determining a scanning direction in digital signal processing for obtaining position information of a radiolabeled substance as symbols and/or numerical values in autoradiography.

Autoradiography is already known as a method for obtaining positional information of radiolabeled substances that are distributed in at least one dimension to form a distribution array on a support medium.

For example, after attaching a radioactive label to a biologically derived polymeric substance such as a protein or a nucleic acid, the radioactively labeled polymeric substance, its derivative, etc.
Alternatively, the decomposition products (hereinafter also referred to as radiolabeled substances) can be separated and developed by performing separation operations such as electrophoresis on a gel-like support medium.
A separated and developed array of radiolabeled substances is formed on the support medium, and then this separated and developed array is transferred to a radiation film and visualized to obtain an autoradiograph as a visible image. location information is obtained. Also,
Based on the obtained location information of the radiolabeled substance,
Methods for separating and identifying the polymeric substances, or evaluating the molecular weight and characteristics of the polymeric substances have already been developed and are in actual use. Autoradiography as described above has been effectively used, particularly in recent years, to determine the base sequence of nucleic acids such as DNA.

As mentioned above, in autoradiography that uses conventional radiography, it is essential to visualize an autoradiograph containing this position information on radiographic film in order to obtain position information of radiolabeled substances. It's summery.

Therefore, researchers determine the distribution of radiolabeled substances on the support medium by visually observing the visualized autoradiograph.
Furthermore, various analyzes are performed based on the visually obtained positional information of the radiolabeled substance to evaluate the characteristics, functions, etc. of the radiolabeled substance.

However, in conventional autoradiography, the analysis work relies on the human eye as mentioned above, so the position information of the radiolabeled substance obtained by analyzing the autoradiograph, which is a visible image, is There are problems such as differences between researchers, and there are limits to the accuracy of the information that can be obtained. In particular, if the autoradiograph visualized on the radiographic film does not have good image quality (sharpness, contrast), it is difficult to obtain satisfactory information, and its accuracy tends to decrease. Conventionally, in order to improve the accuracy of the positional information to be obtained, a method has been used in which, for example, the visualized autoradiograph is measured using a measuring instrument such as a scanning densitometer. However, there is a limit to the improvement of accuracy in methods that simply use such measuring instruments.

For example, during an exposure operation in which the radiation film is brought into close contact with the support medium on which the separation development rows are formed, a shift may occur in the overlay, and in this case, a visible image may be obtained on the radiation film. The separation development column (for example, the electrophoresis column) is not parallel to the length direction of the film,
As a result, errors tend to occur when the positional information of the radiolabeled substance is visually cut in half, and its accuracy tends to decrease. Furthermore, depending on the support medium and the separation and development conditions, the obtained separation and development rows often are not parallel to the length direction of the support medium or are distorted.

Furthermore, when using gel as a support medium, this gel does not have self-supporting properties, so it is usually separated and developed with both sides sandwiched between glass or the like. There may be unevenness, and the radiolabeled substance is not necessarily separated and developed uniformly on the support medium. Similar non-uniform separation and development also occurs when the gel contains air bubbles or when the composition of the gel is non-uniform. For this reason, a so-called smiling effect often appears, in which, for example, the moving distance of the separation and development rows at both ends is relatively short compared to the movement distance of the separation and development rows near the center of the support medium. Alternatively, when performing separation and development by electrophoresis, the voltage may not be applied uniformly to the support medium, and even in such cases, the separation and development conditions will vary locally on the support medium, so the resulting separation and development column Distortion tends to occur.

In the above cases, it becomes particularly difficult to analyze the positional information of the radiolabeled substance, and even if the above-mentioned measuring instruments are used, it is difficult to obtain the positional information of the separated and expanded radiolabeled substance with sufficient accuracy. It is difficult.

The present invention utilizes a radiation image conversion method using a stimulable phosphor sheet instead of the radiography method using a radiation film used in conventional autoradiography, thereby obtaining positional information of a radiolabeled substance. A method is used in which the positional information of an autoradiograph is obtained as a digital signal without specifically converting it into an image, and the obtained digital signal is subjected to specific signal processing to determine the scanning direction for sampling point detection. The inventors have discovered that by doing so, it is possible to obtain positional information of the separated developed product on the support medium with high accuracy, and have arrived at the present invention.

The present invention provides a stimulable phosphor sheet that absorbs radiation energy emitted from a radiolabeled substance that is distributed in at least one dimension to form a distribution array on a support medium. After accumulating and recording an autoradiograph having positional information of the radiolabeled substance on a phosphor sheet, the stimulable phosphor sheet is scanned with electromagnetic waves to emit the autoradiograph as photostimulated light. Scanning different positions on the digital image data at least twice across the one-dimensional distribution direction of the radiolabeled substance with respect to the digital signal corresponding to the autoradiograph obtained by photoelectrically reading out the light, By obtaining the relationship between the position in the scanning direction and the signal level for each scan,
Detect the distribution points of the radiolabeled substance on each scan, then obtain a straight line or curve connecting the distribution points of the radiolabeled substance on each scan in order, and use the obtained straight line or curve for sampling point detection. The present invention provides a signal processing method in autoradiography, characterized in that the scanning direction is set to .

That is, in the present invention, the radiation energy emitted from the sample is absorbed by the stimulable phosphor sheet by overlapping the sample and the stimulable phosphor sheet, and then the stimulable phosphor sheet is exposed to visible light and By scanning with electromagnetic waves (excitation light) such as infrared rays, the radiation energy stored in the stimulable phosphor sheet is released as fluorescence (stimulated luminescence), and this fluorescence is read photoelectrically to obtain an electrical signal. A radioactive image conversion method is used in which this electrical signal is A/D converted to obtain a digital signal.

The radiation image conversion method described above is described in, for example, Japanese Patent Laid-Open No. 12145/1983.

The stimulable phosphor sheet used in the present invention contains, for example, a stimulable phosphor such as a divalent europium-activated alkaline earth metal fluorohalide phosphor. This stimulable phosphor is
After being irradiated with radiation such as rays, alpha rays, beta rays, gamma rays, and ultraviolet rays and accumulating a portion of that radiation energy, when irradiated with electromagnetic waves (excitation light) such as visible light and infrared rays, the accumulated energy is It has the property of exhibiting stimulated luminescence depending on the

According to the present invention, the positional information of a radiolabeled substance is directly obtained as a digital signal without going through any imaging process, using the radiation image conversion method using the above-mentioned stimulable phosphor sheet.

In the present invention, "position information" refers to various types of information centered on the position of a radiolabeled substance or an aggregate thereof in a sample, such as the location and shape of an aggregate of radioactive substances present in a support medium. , refers to various types of information obtained as one or any combination of information such as the concentration and distribution of radioactive substances at that location.

According to the present invention, the positional distortion during separation and development of the radiolabeled substance on the support medium as described above, or the autoradiograph of the radiolabeled substance distributed in a one-dimensional direction to form a distribution row. Even if the entire autoradiograph transferred and accumulated on the stimulable phosphor sheet is distorted or misaligned due to positional misalignment during the operation of transferring it to the stimulable phosphor sheet, the one-dimensional The distribution (separation development) direction can be automatically found and used as the scanning direction for sampling point detection, and position information of the distribution row of radioactive labeled substances can be obtained with high precision along this scanning direction. This is what makes it possible. Furthermore, in the case where the autoradiograph consists of radiolabeled substances distributed in one-dimensional direction with multiple distribution rows,
Even for distortions in individual columns, the one-dimensional distribution direction can be accurately found and set as the scanning direction.

In the present invention, a distribution column refers to a column in which radiolabeled substances are scattered in one direction in the form of bands or spots, such as a migration column obtained by electrophoresis. Moreover, digital image data means a collection of digital signals corresponding to an autoradiograph of a radiolabeled substance.

Examples of samples used in the present invention include:
A support medium in which a radiolabeled substance is separated and developed in a one-dimensional direction can be mentioned. Examples of radioactively labeled substances include biopolymer substances to which radioactive labels have been added, derivatives thereof, and decomposition products thereof.

For example, in the case where the biopolymer substances to which a radioactive label has been given are macromolecular substances such as proteins, nucleic acids, derivatives thereof, and decomposition products thereof, the present invention provides for separation of these biopolymer substances. , useful for identification, etc. Furthermore, the present invention can be effectively used to analyze the entire or partial molecular weight of these biopolymer substances, their molecular structure, or their basic unit configuration. In particular, it is very effective in determining the base sequence of nucleic acids such as DNA.

In addition, methods for separating and developing radiolabeled substances using support media include, for example, gel-like support media (any shape such as layered or columnar), polymer moldings such as acetate, or various types of supports such as filter paper. Typical methods include electrophoresis using a medium and thin layer chromatography using any support medium such as silica gel.
The separation and development method is not limited to these methods.

However, the sample that can be used in the present invention is not limited to the above sample, but is a support medium containing a radiolabeled substance distributed in at least one dimension, and a stimulable phosphor sheet. Any device may be used as long as it is capable of storing and recording an autoradiograph having positional information of the radiolabeled substance.

The basic structure of the stimulable phosphor sheet used in the present invention is a support, a phosphor layer, and a transparent protective film. The phosphor layer is made of a binder containing and supporting the stimulable phosphor in a dispersed state,
For example, it can be obtained by dispersing divalent europium-activated barium fluoride bromide (BaFBr: Eu ²⁺ ) phosphor particles in a mixture of nitrocellulose and linear polyester. A stimulable phosphor sheet is, for example, one in which a sheet of polyethylene terephthalate or the like is used as a support, the above-mentioned phosphor layer is provided on this sheet, and a polyethylene terephthalate sheet or the like is further provided as a protective film on the phosphor layer. .

In the present invention, the operation (exposure operation) of transferring and accumulating radiation energy emitted from a support medium containing a radiolabeled substance onto a stimulable phosphor sheet is performed as follows:
By overlapping the support medium and the stimulable phosphor sheet for a certain period of time, at least a portion of the radiation emitted from the radiolabeled substance on the support medium is absorbed by the stimulable phosphor sheet. This exposure operation can be carried out by placing the support medium and the stimulable phosphor sheet in close proximity to each other, for example, by keeping them in this state for at least several seconds at room temperature or low temperature.

For details of the stimulable phosphor sheet and the exposure operation, please refer to the patent application No. 57-193418 filed by the present applicant.
No. 59-83057).

Next, in the present invention, a method for reading out one-dimensional positional information of a radiolabeled substance on a support medium transferred and accumulated on a stimulable phosphor sheet and converting it into a digital signal will be described in FIG. 1 of the attached drawings. This will be briefly described with reference to an example of the reading device (or reading device) shown in FIG.

Figure 1 shows a pre-reading system for temporarily reading out one-dimensional positional information of a radiolabeled substance accumulated and recorded on a stimulable phosphor sheet (hereinafter sometimes abbreviated as phosphor sheet) 1. Part 2
1 shows a schematic diagram of an example of a reading device comprising a main reading reading section 3 having a function of reading out an autoradiograph stored and recorded on a phosphor sheet 1 in order to output positional information of a radiolabeled substance. There is.

In the prefetch reading unit 2, the following prefetch operation is performed.

The laser light 5 generated from the laser light source 4 passes through the filter 6, whereby a portion of the wavelength range corresponding to the wavelength range of stimulated luminescence generated from the phosphor sheet 1 in response to excitation by the laser light 5 is cut out. Ru. Next, the laser beam is deflected by an optical deflector 7 such as a galvano mirror, reflected by a flat reflecting mirror 8, and then reflected by a phosphor sheet 1.
It is deflected one-dimensionally and incident on the top. The laser light source 4 used here is selected so that the wavelength range of its laser light 5 does not overlap with the main wavelength range of stimulated luminescence emitted from the phosphor sheet 1.

The phosphor sheet 1 is transported in the direction of the arrow 9 under irradiation with the above-mentioned polarized laser light. Therefore, the entire surface of the phosphor sheet 1 is irradiated with the polarized laser light. Note that regarding the output of the laser light source 4, the beam diameter of the laser light 5, the scanning speed of the laser light 5, and the transport speed of the phosphor sheet 1, the energy of the laser light 5 for the pre-reading operation is smaller than the energy used for the main reading operation. It will be adjusted so that

When the phosphor sheet 1 is irradiated with the laser light as described above, it exhibits stimulated luminescence with an amount of light proportional to the accumulated and recorded radiation energy, and this light enters the pre-reading light-guiding sheet 10 . The light guide sheet 10 has a linear incident surface and is placed close to the scanning line on the phosphor sheet 1 so as to face it, and its exit surface forms an annular ring to allow light such as a photoprint to pass through. It communicates with the light receiving surface of the detector 11. The light guide sheet 10 is made by processing a transparent thermoplastic resin sheet such as acrylic synthetic resin, and allows light incident from the incident surface to be transmitted to the exit surface while being totally reflected inside. It is configured to The stimulated luminescence from the phosphor sheet 1 is guided through the light guide sheet 10 and reaches the exit surface, is emitted from the exit surface, and is received by the photodetector 11.

A filter is attached to the light-receiving surface of the photodetector 11, which transmits only light in the stimulated emission wavelength range and cuts out light in the excitation light (laser light) wavelength range.
It is designed to detect only stimulated luminescence. Stimulated luminescence detected by the photodetector 11 is converted into an electrical signal, which is further amplified by the amplifier 12 and output. The accumulated recording information output from the amplifier 12 is input to the control circuit 13 of the main reading reading section 3. The control circuit 13 outputs an amplification factor setting value a and a recording scale factor b so that a signal at an appropriate level can be obtained according to the obtained accumulated recording information.

The phosphor sheet 1 whose pre-reading operation has been completed as described above is transferred to the main reading reading section 3.

In the main reading reading section 3, the following main reading operation is performed.

The laser beam 15 emitted from the main reading laser light source 14 passes through a filter 16 having the same function as the filter 6 described above, and then the beam
The beam diameter is precisely adjusted by the expander 17. Next, the laser beam is deflected by a light deflector 18 such as a galvano mirror, reflected by a plane reflecting mirror 19, and then one-dimensionally deflected and incident on the phosphor sheet 1. Note that an fθ lens 20 is provided between the optical deflector 18 and the plane reflecting mirror 19.
etc. are arranged so that when the polarized laser beam scans the phosphor sheet 1, a uniform beam speed is always maintained.

The phosphor sheet 1 is transported in the direction of the arrow 21 under irradiation with the above-mentioned polarized laser light. Therefore, the entire surface of the phosphor sheet 1 is irradiated with the polarized laser light, as in the pre-reading operation.

When the phosphor sheet 1 is irradiated with laser light as described above, it emits stimulated luminescence with an amount of light proportional to the accumulated and recorded radiation energy, as in the pre-reading operation, and this light is used as a guide for main reading. The light enters the optical sheet 22. This light-guiding sheet 22 for main reading has the same material and structure as the light-guiding sheet 10 for pre-reading, and the stimulated luminescence guided through the interior of the light-guiding sheet 22 for main reading through repeated total reflections. The light is emitted from the exit surface and is received by the photodetector 23. Note that a filter that selectively transmits only the wavelength region of stimulated luminescence is attached to the light receiving surface of the photodetector 23, so that the photodetector 23 detects only stimulated luminescence.

The stimulated luminescence detected by the photodetector 23 is converted into an electrical signal, which is amplified to an appropriate level electrical signal in the amplifier 24 whose sensitivity is set according to the amplification factor setting value a, and then A/D converted. The signal is input to the device 25. The A/D converter 25 converts the signal into a digital signal using a scale factor suitable for the signal fluctuation width according to the recording scale factor setting value b.

Regarding the method for reading the positional information of the radiolabeled substance on the support medium that has been transferred and accumulated on the stimulable phosphor sheet in the present invention, the readout operation consisting of the pre-reading operation and the main reading operation has been described above. The read operations that can be utilized in the present invention are not limited to the above examples. For example, if the amount of radiolabeled substance on the support medium and the exposure time of the stimulable phosphor sheet for the support medium are known in advance, the look-ahead operation can be omitted in the above example.

Furthermore, as a method for reading the positional information of the radiolabeled substance on the support medium that has been transferred and accumulated on the stimulable phosphor sheet in the present invention, it is of course possible to use any method other than those exemplified above. be.

The digital signal corresponding to the sample autoradiograph thus obtained is then input to the signal processing circuit 26. The signal processing circuit 26 determines the scanning direction for sampling point detection.

In the following, the digital signal processing of the present invention will be explained using as an example an autoradiograph obtained by separating and developing a mixture of radiolabeled substances on a support medium by electrophoresis or the like.

Figure 2 shows the transfer and accumulation of multiple types of radiolabeled substances on a stimulable phosphor sheet of a sample, which forms two separate and expanded distribution rows in the length direction on a support medium. An example of an autoradiograph is shown. The autoradiograph on the stimulable phosphor sheet is distorted as shown in Figure 2, due to the fact that the sample and the stimulable phosphor sheet are misaligned and overlapped during the storage transfer. .

By applying the radiation image conversion method to this sample as described above, the signal processing circuit 2
The input digital signal of No. 6 is an address (x,
y) and the signal level (z) at that address, and the signal level corresponds to the amount of photostimulated light. That is, the digital signal corresponds to the autoradiograph shown in FIG. Therefore, digital image data having positional information of the radiolabeled substance is input to the signal processing circuit 26.

In FIG. 2, assuming that the horizontal direction is the x-axis direction and the vertical direction is the y-axis direction with respect to the stimulable phosphor sheet, the scanning direction is determined, for example, through the following steps. can be determined.

First, the digital image data is numerically scanned in the x-axis direction so as to cross the one-dimensional distribution direction of the radiolabeled substance, that is, the direction of the distribution column. Obtain the relationship between the signal level (z) at the position.
For example, when considering a graph in which the horizontal axis represents the position (x) on the scanning area and the vertical axis represents the signal level (z), a graph as shown in FIG. 3A is obtained.

Figure 3A is a graph when the radiolabeled substance is separated and developed over a certain width, and the digital image data corresponding to the autoradiograph in Figure 2 is scanned as described above. The graph obtained by is shown below.

In the graph A of FIG. 3, each midpoint (x _an ) of the region where the signal level is maximum is defined as the distribution point of the radiolabeled substance in each column. Here, a
is a positive integer and represents the scan number (i.e., how many times the scan is performed), and m is a positive integer and represents the distribution column number (in this case, a=1,
or 2). Therefore, x _an above means the distribution point of the radiolabeled substance on the m-th separation development column detected in the a-th scan.

In the signal processing method of the present invention, the digital signal obtained by reading out the stimulable phosphor sheet is temporarily stored in a memory in the signal processing circuit 26 (or stored in a buffer memory or a non-volatile memory such as a magnetic disk). ). In signal processing, scanning digital image data means selectively retrieving only the digital signal at this scanning location from memory.

By scanning at least twice at different positions on the digital image data, that is, at different y-coordinates, a graph as shown in Fig. 3A is obtained for each scanned area in a virtual manner, and from these graphs, the above-mentioned In this way, two or more sets of distribution points of radiolabeled substances, {(x _a1 , x _a2 ), a=1, 2, . . . } are obtained.

Next, a straight line (or a broken line) passing through the distribution points of the radiolabeled substance in order is obtained for each column. The obtained straight line (or polygonal line) is taken as the scanning direction for sampling point detection. Of course, a least squares straight line or curve may be determined for each point.

That is, for example, by scanning two coordinates (y ₁ and y ₂ ) on the y-axis twice in parallel to the x-axis direction, a set of distribution points of the radiolabeled substance, {(x ₁₁ , x ₁₂ ), ( x ₂₁ , x ₂₂ )}, the scanning direction for sampling point detection for the first column is the distribution point (x ₁₁ ,
Let it be a straight line passing through the two points y ₁ ) and (x ₂₁ , y ₂ ).
Similarly for the second column, the distribution point (x ₁₂ , y ₁ )
The scanning direction is the straight line passing through the two points of and (x ₂₂ , y ₂ ).

This scanning in the x-axis direction can be performed at any position in the y-axis direction. However, when determining the scanning direction for sampling point detection by scanning twice, that is, using two distributed points, the scanning direction between each scanning position should be It is preferable that the intervals between the two are as far apart as possible, and the scanning positions are at the top (or near the top) of the separation and development row of radiolabeled substances.
and the lower end (or its vicinity). Furthermore, although the above-mentioned scanning does not necessarily have to be performed in parallel, it is preferable to perform the scanning in parallel to each other.

Furthermore, the above scanning needs to be performed with a width that covers at least one separation and development site of the radiolabeled substance for each column. That is, the digital image data is scanned in the y-axis direction with a constant width centered at the scanning position. If the scanning width is too narrow, not only may the area where the radiolabeled substance is separated and developed (the area where the separated and developed radiolabeled substance is present) not be covered, but even if it is However, if the distribution of the radiolabeled substance in that region is uneven, errors may occur in the detected distribution points. On the other hand, if the scanning width is too wide, an error will occur in the detected distribution point of the radiolabeled substance.
Therefore, it is desirable to set the scanning width depending on the sample.

The graph shown in FIG. 3A is obtained, for example, by repeatedly extracting digital signals within a certain scanning width in the y-axis direction and adding the levels of the signals for each x-coordinate. Furthermore, it is also possible to perform threshold processing on this addition data. Alternatively, it can be obtained by repeatedly extracting digital signals within this scanning width in the x-axis direction and adding digital signals subjected to threshold processing for each y-coordinate in order to reduce noise. At this time, the representative value of the y-coordinate is a point corresponding to the center of the scanning width.

Here, threshold processing is to set the signal level to 1 for digital signals whose signal level is above a certain level value (i.e., threshold value), and to set the signal level to 0 for digital signals whose signal level is less than the threshold value. This refers to a binarization process in which the level of all digital signals is displayed as 1 or 0.

Further, this scanning can also be performed as follows. That is, after repeatedly extracting digital signals within the above scanning width in the x-axis direction and finding the x-axis coordinate x _ai at which the signal level is maximum for each y-coordinate, the local average coordinate of each x-coordinate is calculated. _{The distribution point} ⁽ _{x a1 , _ y1} )
shall be.

The above scanning position and scanning width may be independently input to the signal processing circuit 26 for each sample to be measured before the digital signal thereof is processed. By setting the scanning position and scanning width for each sample used in this way, even if the one-dimensional distribution of radiolabeled substances varies depending on the type of sample, separation and development conditions, etc., it is possible to Distribution points can be detected accurately.

The more the number of scans is than two, the more distribution points of the radiolabeled substance can be detected.
Straight line (broken line) obtained by connecting those distribution points
If this is the scanning direction for sampling point detection, it will follow the distribution line of the radiolabeled substance even more. Furthermore, by approximating the obtained polygonal line with an appropriate curve, it becomes possible to determine the scanning direction for sampling point detection even more accurately. However, as a result of increasing the number of scans,
Since there is a problem that signal processing becomes complicated and processing time becomes long, it is preferable that the number of scans is determined depending on the condition of the sample and the accuracy of wear in autoradiographic measurement.

For example, in autoradiography of a sample in which radioactively labeled nucleic acids, their derivatives, or their decomposition products are separated and developed on a support medium using electrophoresis or the like,
It is possible to determine the scanning direction for sampling point detection with high accuracy by performing two scans. At this time, it is preferable that the scanning width is a width that spans two to three radiolabeled substance distribution (separation and development) sites for each row.

By determining the scanning direction for sampling point detection as described above, it is possible to narrow the width of each distribution site of the radiolabeled substance to about 3 mm. Therefore, according to the method of the present invention, it is possible to reduce the amount of radiolabeled substance per separation and development column, and therefore, it is possible to increase the number of separation and development columns per support medium. That is, with a single autoradiograph measurement, it is possible to obtain more positional information than was obtained through conventional operations.

In addition, in the signal processing of the present invention, the distribution points of radiolabeled substances on each scanning region can be determined by obtaining a graph obtained by differentiating the graph A in FIG.
Its detection becomes easier. In other words, by performing a differential operation on the above graph, the edges of the separation development column can be emphasized, and therefore both ends of the separation development column in the width direction can be easily detected on the digital image data, and the radiolabeled substance can be easily detected. This is because it can be easily detected by setting the distribution point of, for example, the midpoint of both edges.

A in FIG. 4 shows a graph obtained by differentiating the graph A in FIG. 3. From the graph A in Figure 4,
The edges of each separation expansion column can be easily detected. That is, the distribution point of the radiolabeled substance in each column can be set to each midpoint (x _an ') when the differential level value changes from positive to negative.

On the other hand, if the radiolabeled substance is separated and developed in the form of spots, if there is a significant shift or distortion in the separation and development row, and/or due to the above scanning conditions (scanning position and scanning width), the horizontal axis indicates the position in the scanning direction. A graph in which (x) is plotted and the signal level (z) is plotted on the vertical axis is as shown in FIG.

In the graph (b) of FIG. 3, each point ( _xbo ) at which the signal level is maximum can be taken as the distribution point of the radiolabeled substance in each column. here,
b is a positive integer and represents the scan number, and n
is a positive integer and represents the column number.

4 shows a graph obtained by differentiating the graph B of FIG. 3. In the graph (b) of FIG. 4, the distribution points of the radiolabeled substance in each column can be set to the respective midpoints ( _xbo ') when the differential level value changes from positive to negative. In each of the above cases, the scanning direction for sampling point detection can be determined in the same manner as the method described above.

In addition, in the above-mentioned example shown in FIGS. 2 to 4, the case where there are two separation and expansion columns has been explained, but the signal processing method of the present invention is not limited to these two columns. The present invention can be applied even if the radiolabeled substance distribution array is one row, such as a separation and development row, or multiple rows of three or more rows.

Along the scanning direction obtained by the signal processing method of the present invention, it is possible to determine sampling points for detecting the distribution site of the radiolabeled substance. If there are multiple distribution rows in a one-dimensional direction, the one-dimensional positional information of the radiolabeled substance can be expressed as a symbol by comparing and identifying the determined sampling points between corresponding positions in the scanning direction. and/or can be obtained as a numerical value.

The signal processing method in autoradiography of the present invention is a very useful method, for example, in a DNA base sequencing method using autoradiography such as the Maxam-Gilbert method.

In other words, the above Maxam-Gilbert method is
Automated DNA is obtained by specifically cleaving radioactively labeled DNA into its constituent units, each of the four types of bases, and then separating and developing the mixture of the base-specific cleavage products using electrophoresis. This is a method for determining DNA base sequences from radiographs, and according to the signal processing method of the present invention, no matter what combination of base-specific cleavage products there are, separation and expansion can be performed for each of the separation and expansion columns. The direction can be found and taken as the scanning direction.

[Brief explanation of the drawing]

FIG. 1 shows an example of a reading device (or reading device) for reading positional information of a radiolabeled substance in a sample that has been transferred and accumulated on a stimulable phosphor sheet in the present invention. 1: stimulable phosphor sheet, 2: readout section for pre-reading, 3: readout section for main reading, 4: laser light source,
5: Laser light, 6: Filter, 7: Optical deflector, 8: Planar reflector, 9: Transfer direction, 10: Light guide sheet for prereading, 11: Photodetector, 12: Amplifier, 13: Control circuit, 14: Laser light source, 1
5: laser beam, 16: filter, 17: beam expander, 18: optical deflector, 19: plane reflecting mirror, 20: fθ lens, 21: transport direction, 2
2: Light guide sheet for main reading, 23: Photodetector, 2
4: Amplifier, 25: A/D converter, 26: Signal processing circuit. FIG. 2 is an autoradiograph in which a radiolabeled substance is transferred and accumulated on a stimulable phosphor sheet of a sample separated and developed on a support medium. 3A and 3B are graphs representing the relationship between the scanning position and the level of the digital signal, respectively. Fourth
A and B in the figure are graphs obtained by differentiating the graphs A and B of FIG. 3, respectively.

Claims (1)

[Claims] 1. By causing a stimulable phosphor sheet to absorb radiation energy emitted from a radiolabeled substance that is distributed in at least one dimension to form a distribution row on a support medium, After accumulating and recording an autoradiograph having positional information of the radiolabeled substance on the stimulable phosphor sheet, the stimulable phosphor sheet is scanned with electromagnetic waves to emit the autoradiograph as photostimulated light, and Scanning different positions on the digital image data at least twice so as to traverse the distribution row of the radiolabeled substance with respect to the digital signal corresponding to the autoradiograph obtained by photoelectrically reading out this photostimulated light, By obtaining the relationship between the position in the scanning direction and the signal level for each scan, each distribution point of the radiolabeled substance is detected on each scan, and then the distribution point of the radiolabeled substance on each scan is sequentially detected. A signal processing method in autoradiography, characterized in that a continuous line consisting of a straight line, a broken line, or a curved line is set by connecting the lines, and the set continuous line is used as a scanning direction for sampling point detection. 2. The autoradiograph according to claim 1, wherein scanning of different positions on the digital image data is performed at least twice in substantially parallel to each other so as to traverse the distribution row of the radiolabeled substance. Signal processing method in E. 3. The automatic system according to claim 1 or 2, wherein the scanning position and scanning width on the digital image data are set before signal processing according to the state of the distribution column to be measured. Signal processing methods in radiography. 4 Scan the digital image data twice to detect the distribution points of the radiolabeled substance at two locations for one distribution column, and use the straight line obtained by connecting these two distribution points to scan for sampling point detection. A signal processing method in autoradiography according to any one of claims 1 to 3, characterized in that the signal processing method is performed in a direction. 5. The radiolabeled substance forming a distribution array on the support medium is a biopolymer substance, a derivative thereof, or a decomposition product thereof, to which a radiolabel is attached and which is separated and developed in a one-dimensional direction on the support medium. The first claim characterized in that
A signal processing method in autoradiography according to any one of items 1 to 4. 6. The signal processing method in autoradiography according to claim 5, wherein the biopolymer substance is a nucleic acid, a derivative thereof, or a decomposition product thereof.