JPS59126529A - Signal processing method in autoradiography - Google Patents

Abstract

PURPOSE:To enable the detection of highly accurate positional information on a medium of a radioactive marking material by obtaining the positional information on said medium as a digital signal and subjecting the signal to special processing and utilizing a method for determining the scanning direction for the purpose of detection of sampling point. CONSTITUTION:The accelerated luminous light from a phosphor sheet is conducted to the inside of a photoconductive sheet 10 and arrives at an exit face from which said light is received in a detector 11. The accelerated luminous light detected by said detector is converted to an electrical signal which is inputted via an amplifier 12 to a control circuit 13. A signal of an adequate level according to the obtained and accumulated recording information is obtd. in the circuit 13. The graph differentiating the graph shown in the figure is obtd., by which the detection thereof is made easy. More specifically, the edge in the separating and developing row can be stressed by subjecting the signal to a differential calculation and therefore both ends in the transverse direction of the separating and developing row can be detected easily on the digital image data. If the distribution point of the radioactive marking material is made, for example, midpoint of both edges, said material is easily detected.

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 positional 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 a radiolabeled substance that is distributed in at least one dimension to form a distribution array on a support medium.

For example, after attaching a radioactive label to a macromolecular substance derived from a biological body such as a protein or nucleic acid, the radiolabeled macromolecular substance, its derivative, or its decomposition product (hereinafter also referred to as the radiolabeled substance) is used. ) is subjected to a separation operation such as electrophoresis on a gel-like support medium and subjected to fractional development to form a separated and developed column of the radiolabeled substance on the support medium, and then this fractional development column is transferred to a radiographic film. By transferring and visualizing the radiograph, the radiograph is obtained as a visible image, and position information of the radiolabeled substance is obtained from this visible image. Also, based on the location information of the radiolabeled substance obtained in 1).

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. Therefore, autoradiography as described above has been particularly effectively used in recent years for determining the base sequence of nucleic acids such as DNA.

- In autoradiography using conventional radiography as described above, in order to obtain positional information of a radioactively labeled substance, it is necessary to use an autoradiography system that has this positional information. - Visualization of the radiograph on radiographic film is an essential requirement.

Therefore, researchers determine the distribution of radiolabeled substances in the support medium by visually observing the visualized autoradiograph.

Furthermore, the characteristics, functions, etc. of radiolabeled substances are evaluated by performing various analyzes based on visually obtained positional information of the radiolabeled substances.

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 used for research. There are problems such as discrepancies depending on the person, and there is a limit to the accuracy of the printed information. 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 desired positional information, a method has been used in which the visualized autoradiograph is measured using an Al11 constant instrument such as a scanning densitometer. however,
There is a limit to the accuracy of such a method that uses a full-length device.

For example, during an exposure operation in which the support medium on which the separation and development rows are formed and the radiation film are brought into close contact with each other, there may be a misalignment in the overlapping. (-) square development rows (for example, electrophoresis rows) are not parallel to the length direction of the film, but are shifted, so errors are likely to occur when visually determining the positional information of the radiolabeled substance. Therefore, the accuracy tends to decrease. Furthermore, depending on the support medium and the unfolding conditions, the resulting separated and unfolded rows may not be parallel to the length direction of the support medium or may be distorted.

Furthermore, when using gel as a support medium, this gel does not have self-supporting properties, so it is usually spread out with both sides sandwiched between glasses, etc., but the thickness of the gel may be uneven due to deformation of the covering. The radiolabeled substance may not be distributed evenly on the support medium.
It doesn't necessarily mean it will be expanded. 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, in the case of electrophoretic separation and development, the voltage may not be applied uniformly to the support medium, and in such cases, the minute gI
Because the unfolding conditions vary locally on the support medium, distortions tend to occur in the resulting separated unfolded array.

In such cases, it becomes particularly difficult to analyze the positional information of the radiolabeled substance, and even with the use of measuring instruments such as those mentioned above, it is not possible to obtain the positional information of the separated and deployed radiolabeled substance with sufficient accuracy. It is difficult to obtain.

The present inventor has succeeded in obtaining positional information of radiolabeled substances by using a radiographic image conversion method using a stimulable phosphor sheet instead of the radiographic method using a radiographic film used in conventional autoradiography. A method for determining the scanning direction for sampling point detection by converting the position information of an autoradiograph into a digital signal without specifically converting it into an image, and applying specific signal processing to the obtained digital signal. The present inventors have discovered that by utilizing this method, it is possible to obtain positional information of a developed product on a support medium with high accuracy, and have arrived at the present invention.

The present invention enables 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 a sheet, the stimulable phosphor sheet is scanned with electromagnetic waves to emit the autoradiograph as photostimulated light, and this photostimulated light is emitted. scanning different positions of the digital image data at least twice across the one-dimensional distribution direction of the radiolabeled substance for the digital signal corresponding to the autoradiograph obtained by photoelectrically reading it out; 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. The present invention provides a signal processing power method in autoradiography, which is characterized in that a uniform line or curve is obtained by connecting the lines, and the obtained straight line or curve is used as a scanning direction for sampling point detection.

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, infrared rays, etc. By scanning with electromagnetic waves (excitation light), the radiation energy stored in the stimulable phosphor sheet is converted into fluorescence (stimulated luminescence).
This method utilizes a radiation image conversion method that consists of emitting this light as 11 photoelectrically to obtain an electrical signal, and A/D converting this electrical signal to output it as a digital signal. .

Regarding the radiation image conversion method described above, for example, U.S. Pat. J55
-12145, etc.

The stimulable phosphor sheet used in the present invention is, for example,
It is used to stimulate stimulable phosphors such as divalent europium-activated alkaline earth metal fluorohalide phosphors. This stimulable phosphor is irradiated with radiation such as X-rays, α-rays, β-rays, γ-rays, and ultraviolet rays and accumulates a portion of the radiation energy. When irradiated with light (light), it exhibits stimulated luminescence depending on the accumulated energy.

According to the present invention, the positional information of a radiolabeled substance is directly obtained as a digital signal having a certain level without going through any particular 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. , concentration of radioactive material at that location, minute 711, etc., or any combination of information obtained.

According to the present invention, the positional distortion when the radiolabeled substance is spread on the support medium as described above, or the autoradiation of the radiolabeled substance that is distributed in the -dimensional direction and forms a distribution row. If distortion or misalignment occurs throughout the Oi Radiograph transferred and stored on the stimulable phosphor sheet due to misalignment during the operation of transferring the graph onto the stimulable phosphor sheet, the The direction of dimensional overlap IO (minute Nil open) can be automatically found and used as the scanning direction for sampling point detection, and along this scanning direction, the minute /
[This makes it possible to obtain position information in five columns with high precision. In addition, when an autoradiograph consists of a radiolabeled substance distributed in multiple distribution rows in the dimensional direction, the direction of the dimensional distribution can be accurately determined even when the individual rows are distorted. The scanning direction can be set as the scanning direction.

In the present invention, a distribution array refers to, for example, a migration array obtained by electrophoresis, in which a radiolabeled substance is
Refers to rows scattered in one direction in the form of 12 or sponts. Moreover, digital image data means a collection of digital signals corresponding to an autoradiograph of a radiolabeled substance.

A. An example of a sample used in the invention is a support medium in which a radiolabeled substance is separated and developed in one dimension. Examples of radiolabeled substances include biopolymer substances to which radioactive labels have been added, 1″A conductors thereof, and decomposition products thereof.

For example, in the case where the biopolymer substance to which the radioactive label has been labeled (+'i') is a macromolecular substance such as a protein, a nucleic acid, a derivative thereof, or a decomposition product thereof, It is useful for the separation and identification of polymeric substances.Furthermore, the present invention can be used to analyze the entire or partial molecular weight of these biological polymeric substances, their molecular structure, or their basic unit configuration. It can be used effectively, and is particularly effective in determining the base sequence of nucleic acids such as DNA.

In addition, radiolabeled substances can be separated using a support medium.
Examples of methods for performing it include electrophoresis using various support media such as gel-like support media (any shape such as layered or columnar), polymer moldings such as acetate, or cancer paper, and supports such as silica gel. Although thin layer chromatography using a medium is a typical method, 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 sample listed in item 1, but is a support medium containing a radiolabeled substance weighing at least 1 g in one dimension, and that has cumulative fluorescence. Any device may be used as long as it is capable of storing and recording an autoradiograph having positional information of the radiolabeled substance in the body.

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, divalent 1 europium, activated fluorofluorobromide/1lium (BaFB r :Eu
”) is obtained by dispersing phosphor particles in a mixture of nitrocellulose and linear polyester.A stimulable phosphor sheet can be obtained by, for example, using a sheet of polyethylene terephthalate as a support. The following phosphor layer was provided on the sheet, and a polyethylene terephthalate sheet was further provided as a protective film on the phosphor layer.

In the present invention, a stimulable phosphor C.I.
The transfer/accumulation operation (exposure operation) to the stimulable phosphor sheet involves overlapping the support medium and the stimulable phosphor sheet for a certain period of time to transfer at least a portion of the radiation emitted from the radiolabeled substance on the support medium to the stimulable phosphor sheet. It is carried out by absorbing it into a 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.

The details of the M-resistant phosphor sheet and the exposure operation are described in Japanese Patent Application No. 193418/1983, filed by Mizu Applicant.

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 apparatus drawing. 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 radioactive label stored in a stimulable phosphor sheet (hereinafter sometimes abbreviated as phosphor sheet). 2 and a main reading reading section 3 having a function of reading out the autoradiograph stored in the phosphor sheet 1 in order to output positional information of the radiolabeled substance.

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, so that the part of the wavelength range corresponding to the wavelength range of stimulated luminescence generated from the phosphor sheet 1-1 in response to excitation by the laser light 5 is be cut. Next, the laser beam is deflected by the optical deflector 7 of the galvano mirror '5, reflected by the plane reflecting mirror 8, and then incident on the phosphor sheet 11 after being deflected in a -dimensional manner. The laser light source 4 used here is selected so that the wavelength range of the laser light 5 thereof does not overlap with the main wavelength range of stimulated luminescence emitted from the light body 7j' 1-1.

The phosphor sea 1・l is 11(χ
Under fire, it is transported in the direction of arrow 9. Therefore, the polarized laser beam is irradiated onto the entire surface of the light body 1.l. In addition, the amount of laser light (output of 4, beam diameter of laser light 5, scanning speed of laser light 5,
The transport speed of the phosphor sheet 1-1 is adjusted so that the energy of the laser beam 5 for the pre-reading operation is smaller than the energy used for the main reading operation.

When the phosphor sheet 1 is irradiated with the laser beam 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 groove optical sheet 10. . This light guiding sheet 1O has a linear incident surface, and the phosphor sheet 1! It is arranged close to one side so as to face one scanning line, and its exit surface forms a ring and communicates with the light-receiving surface of the formal photodetector 11. This light-guiding sheet 1.10 is made by modifying a transparent thermoplastic resin sheet 1., for example, an acrylic synthetic resin, and the light □ incident from the use surface is completely absorbed inside it. It is configured to be transmitted to the exit surface while being reflected. The stimulated luminescence of the phosphor C i-1 is due to this light-guiding she! - The light is guided through the lo 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 1l, which transmits only light in the wavelength region of stimulated luminescence and cuts out light in the wavelength region of excitation light (laser light), so that only stimulated luminescence is detected. It is made detectable. Stimulated luminescence detected by the photodetector 11 is converted into an electrical signal, which is further amplified by the amplifier 12 and output. The stored and recorded information output from the amplifier 12 is sent to the control circuit l3 of the reading section 3 for main reading. The control circuit 13 outputs an amplification factor setting value a and a recording scale factor b in accordance with the obtained accumulated recording information so that a signal at an appropriate level can be obtained.

Phosphor sheet 1 whose pre-reading operation has been completed as described above
is transferred to the tree reading reading unit 3.

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

Laser light for reading books! The laser beam 15 emitted from the laser beam 714 passes through a filter 16 having the same function as the filter 6 described above, and then the beam diameter is precisely adjusted by a beam expander 17. Next, the laser beam is deflected by a light deflector 18 such as a galvanometer mirror, reflected by a plane reflecting mirror 19, and then one-dimensionally deflected and incident on the phosphor sheet l. Note that an fO lens 20 or the like is arranged between the optical deflector 18 and the plane reflecting mirror 19, so that when the deflected laser beam scans the phosphor sheet 1, a uniform beam speed is always maintained. has been done.

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

When the phosphor sheet 1 is irradiated with laser light in a bipedal manner, 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 in the main reading. The optical groove 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. The emitted light is emitted from the emission surface and is received by the photodetector 23.A filter is attached to the light receiving surface of the photodetector 23 that selectively transmits only the wavelength range of stimulated luminescence. The detector 23 is designed to detect 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 sent to the A/D converter 25. is input. A/D converter 2
5 is converted into a digital signal with a scale factor suitable for the signal fluctuation range according to the recording scale factor setting value.

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 tree-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 by taking 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 on a stimulable phosphor sheet of a sample in which multiple types of radiolabeled substances are spread out in two rows in the length direction on a support medium. An example of an autoradiograph is shown. Autoradio using stimulable phosphor sheets.

The graph is distorted as shown in FIG. 2 due to the fact that the sample and the stimulable phosphor sheet were misaligned and overlapped during the storage transfer.

By applying the radiation image conversion method to this sample as described above, the digital signal input to the signal processing circuit 26 is converted to an address expressed in the coordinate system fixed to the stimulable phosphor sheet. (X.

y) and the signal level (2) 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 row, to determine the position (X) on the scanning area and its location. The signal level at the position (2
) to get a relationship.

For example, if we consider it as a graph where the horizontal axis is the order 1δ(X) on the scanning area and the vertical axis is the signal level (2),
A graph as shown in FIG. 3(a) is obtained.

Figure 3 (a) is a graph when the radiolabeled substance is separated and developed with 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 of FIG. 3(a), each midpoint (Xam) 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 (that is, how many times the scan is performed), and m is a positive integer and represents the number of the distribution column (in this case, a-1 , or 2). Therefore, the above Xim 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 (8. In a nonvolatile memory such as a far memory or a magnetic disk). be remembered). In signal processing, scanning digital image data means extracting 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 like the one shown in FIG. - Two or more sets of distribution points of radiolabeled substances, 1 (Xat, Xaz), as shown in F.
a= 1, 2, --) is 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 minimum perpendicular line or curve may be found for each point. Of course, a least squares straight line or curve may be determined for each point.

That is, for example, two coordinates on the y-axis (yt and yz
) twice in parallel to the X-axis direction, the set of distribution points of the radiolabeled substance, f(X+t,Xtz)+(
When two sets of distribution points (X211X22)) are obtained, the scanning direction for sampling point detection for the first column is
Let it be a straight line passing through two points: X's t + yx) and ゛(xz buy, yz). Do the same for the second column,
Distribution points (X1' 21 yt') and (X221 V
Let the straight line passing through the two points z) be the scanning direction.

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 from two scanning points, that is, from two distributed points, the scanning direction between each scanning position must be adjusted so that the obtained scanning direction matches the actual separation development row as much as possible. It is preferable that the intervals between the two are as far apart as possible, and the scanning positions are
It is desirable to select the upper end (or its vicinity) and the lower end (or its vicinity) of the radiolabeled substance development column.

Yes. 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 two-legged scan must be performed with a width that covers at least one development site of the radiolabeled substance for each row. In other words, the digital image data is scanned in the y-axis direction with a constant width centered at the scanning position.
Ru. If the scan width is too narrow, the radiolabeled substance should be
Not only is there a possibility that it will not reach the deployment site (the site where the deployed radiolabeled substance is present), but even if it does reach the separated deployment site, the distribution of the radiolabeled substance at that site is uneven. , there is a possibility that 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 distribution point of the detected 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 signal levels 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 X-coordinate in order to reduce noise. At this time, the representative value of the X coordinate is a point corresponding to the center of the scanning width.

Here, threshold processing means that for digital signals whose signal level is above a certain level value (i.e., dark value), the signal level is set to 1, and for digital signals below the threshold value -5, the signal level is set to 1. By setting it to 0, the signal level for all digital signals can be set to 1 or O.
This refers to the binarization process that is displayed in .

Further, this scanning can also be performed as follows. In other words, after repeatedly extracting the digital signal within the above scanning width in the X-axis direction and finding the X-coordinate X at which the signal level is maximum for each X-coordinate, the average coordinate of each X-coordinate is calculated. The calculated X coordinate (X o = winter
').

The above-mentioned 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 the radiolabeled substance varies depending on the type of sample, separation and development conditions, etc., it is possible to Points can be detected accurately.

The more the number of scans than two, the more distribution points of the radiolabeled substance can be detected, and the straight line (broken line) obtained by connecting these distribution points was used as the scanning direction for sampling point detection. In some cases, the distribution line will be more along the distribution line of the radiolabeled substance. Furthermore, by approximating the obtained polygonal line with an appropriate curve, it becomes possible to more accurately determine the scanning direction for sampling point detection. However, as a result of increasing the number of scans, there is a problem that signal processing becomes complicated and processing time becomes longer, so the number of scans should be determined depending on the condition of the sample and the accuracy of wear in autoradiographic measurements. preferable.

For example, in autoradiography of a sample in which radioactively labeled nucleic acid derivatives or their decomposition products are separated and developed on a support medium using electrophoresis or the like, the number of scans is two. It is possible to determine the scanning direction for sampling point detection with high accuracy. At this time, it is preferable that the scanning width is a width that spans ij (separation and development) sites of 2 to 3 radiolabeled substances 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 minute development column, thereby increasing the minute development column per support medium. With multiple autoradiographs, it is possible to obtain more positional information than could be obtained through conventional operations.

Furthermore, in the signal processing of the present invention, the distribution points of the radiolabeled substance in each scanning region can be easily detected by obtaining a graph obtained by differentiating the graph of (a) in FIG. In other words, by performing a differential operation on the above graph, the edges of the 1 minute # 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 9111 point of , for example, as the midpoint between both edges.

FIG. 4(A) shows a graph obtained by differentiating the graph of FIG. 3(A). From the graph (a) in Figure 4,
The edges of each separation expansion column can be easily detected.

That is, the distribution points of the radiolabeled substances in each column are the midpoints (Xam
') can be.

, if the deviation or distortion of the development row is significant, and/or according to the scanning conditions (scanning position and scanning width), the position (X) in the scanning direction is set on the horizontal axis, A graph in which the signal level (Z) is plotted on the vertical axis is as shown in (b) of FIG.

In the graph of FIG. 3 (b), each point (Xbn) where the signal level is maximum can be taken as the distribution point of the radioactive label or 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.

Moreover, (b) of FIG. 4 shows a graph obtained by differentiating the graph of (b) of FIG. 3. In the graph in (b) of Figure 4, the distribution points of the radiolabeled substances in each column are the midpoints (Xb,
1'). 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 minute # expansion columns has been explained, but the signal processing method of the present invention is not limited to these two columns. It can be applied even if there is only one radiolabeled substance distribution column, such as a separation/development column, or multiple columns of three or more columns.

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 the −-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 can be applied, for example, to Maxam Gilbert (Maxam-G.
This is a very useful method in DNA base sequencing methods using autoradiography, such as the 1999 method.

That is, the above-mentioned Maxam φ Gilbert method specifically cleaves radioactively labeled DNA for each of the four types of bases that are its constituent units, and then generates a mixture of the base-specific cleavage products. DN from an autoradiograph obtained by electrophoretic development.
According to the signal processing method of the present invention, no matter what combination of base-specific cleavage and decomposition products are used, it is possible to determine the base sequence of A.
The opening 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. Eco-storage 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: plane reflecting mirror, 9: transport direction, 10: light guide sheet for pre-reading, 11: photodetector, 12-amplifier, 13: control circuit, 14: laser light source, 15 : Laser light, 16: Filter, 17: Beam φ expander 118: Optical deflector, 19: Plane reflecting mirror, 20: fO lens, 21: Transfer direction, 22 Light guiding sheet for double reading), 23: Photo detection instrument, 24: amplifier, 25:
A/D converter, 26: Signal processing circuit FIG.・This is the autoradiograph transcribed and accumulated above. (A) and (B) in FIG. 3 are graphs showing the relationship between the scanning position and the level of the digital signal, respectively. (A) and (B) in Figure 4 correspond to (A) and (B) in Figure 3.
Graphs obtained by differentiating the graphs of (b) Patent applicant Fuji Photo Film Co., Ltd. Agent Patent attorney Yasuo Yanagawa Figure 3 (/1) - Teito Mitsugu? JE Book January 25, 1980 Commissioner of the Japan Patent Office Kazuo Wakasugi 2゜Name of the invention Signal processing method in autoradiography 3゜Relationship with the case of the person making the correction Patent applicant address (520) Fuji Photo Film Co., Ltd. Name The number of inventions increases due to the amendment by Representative Yu Nishi 4 degrees and attorney 6 degrees None Procedural amendment document O month, Showa A//day Commissioner of the Japan Patent Office Kazuo Wakasugi 1, Indication of case 1988 Patent application no. 1328 No. 2, Title of the invention: Signal processing method in autoradiography 3
Relationship with the case of the person making the amendment Patent Applicant 4, Agent 8, Contents of the Amendment As shown in the attached sheet, the "Detailed Description of the Invention" column of the specification will be amended as follows. Subscript and (2) Page 11, line 7 Has a certain level → Excluding 1 (3) Page 16, line 7 Memory → 4 (4
) if (page 10th line memory - record (5
) Page 25, line 19 Of course, each point → Delete (8
) Page 25, line 20 (- line deleted) (7)
2B page 1st line → Portrait (
8) Page 28, line 18 Sunahachi → womuh
b(9) 2B page 20th level is maximum → k
5ru is extremely thick λminato (10) page 28, line 20 X coordinate X + each coordinate X1yu (+2) page 29, line 3 Xll”Σxi/
N -” xa1 = each xat/N (13) 29 pages 4
Line Xll → Xal's (+4)2
Page 9, line 4
) Page 29, line 5, y coordinate Vo → Average position of L coordinate (18) Page 2B, line 5 (X n + V
n) →
A corrected drawing of Figure 4 is attached. that's all

Claims (1)

[Claims] ■. By causing the stimulable phosphor sheet to absorb radiation energy emitted from the radiolabeled substance distributed in at least one dimension and forming 41 rows on the support body, the stimulable phosphor sheet After accumulating and recording an autoradiograph having positional information of the radiolabeled substance, the stimulable phosphor sheet is scanned with electromagnetic waves to emit the autoradiograph as photostimulated light. to the digital signal corresponding to the autoradiograph obtained by optically reading out the
Accordingly, by scanning different positions on the digital image data at least twice across the distribution row of the radiolabeled substance and obtaining the relationship between the position in the scanning direction and the signal level for each scan, each Detect each distribution point of the radiolabeled substance on the scan, and then connect the distribution points of the radiolabeled substance on each scan to H+1/+ to set a continuous line consisting of a straight line, broken line, or curved line. A signal processing method in autoradiography whose key feature is to use a line as the scanning direction for sampling point detection. 2. The autoradio 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 methods in graphics. 3. The scanning position and scanning width on the digital image data are determined based on the shape of the distribution row of the measurement target.6. Claim 1 or 2 is characterized in that the settings are made before signal processing, depending on the
Signal processing method in autoradiography as described in section. 4゜By scanning the digital image data twice, the distribution points of the radiolabeled substance are detected at three locations for one distribution column, and the straight line obtained by connecting these two distribution points is used 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 scanning direction. 5゜ A biopolymer material to which a radioactive label is attached, in which a radiolabeled substance forming a linear array on a support medium is spread out in a one-dimensional direction on the support medium,
The signal processing method in autoradiography according to any one of claims 1 to 4, characterized in that the signal processing method is a derivative thereof or a decomposition product thereof. 6. Claim 5, wherein the biopolymer substance is a nucleic acid, a derivative thereof, or a decomposition product thereof.
Signal processing method in autoradiography as described in section.