JPS61219862A - Autoradiograph analysis - Google Patents

Abstract

PURPOSE:To make it possible to perform autoradiograph analysis with high sensitivity and good efficiency, by performing proper signal processing by imparting necessary information relating to various strains of the digital signal corresponding to an autoradiograph. CONSTITUTION:A mixture consisting of four kinds of base specific DNA segments having radioactive labels applied thereto is separated and developed on a gel support medium by electrophoresis and positional informations of said segments are obtained as digital signals. When a smiling phenomenon is generated in a migration pattern, the peaks of each band are displayed on a picture as spots in a superposed state and a cursor point is coincided with the migration start point of the lane having generated smiling to confirm said position and coincided with the arbitrary position on said lane to confirm said position before being moved downwardly along the lane direction to be fixed at a proper position.

Description

DETAILED DESCRIPTION OF THE INVENTION Field of the Invention The present invention relates to an autoradiographic analysis method.

[Background of the Invention] 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, a radiolabel is added to a biologically derived polymeric substance such as a protein or a nucleic acid, and the radiolabeled polymeric substance, its derivative, its decomposed product, or its composite is separated using gel electrophoresis or other methods. By overlapping the gel-like support medium and a high-sensitivity X-ray film for a certain period of time, the film is exposed to light, and the radiolabeled substance on the support medium obtained from the exposed area is released. Based on the location information, the polymer substance can be separated and identified, or the molecular weight of the polymer substance,
Methods for evaluating characteristics have also been developed and are in actual use.

Particularly in recent years, autoradiography has been
A, it is effectively used for determining the base sequence of nucleic acids such as RNA. It is also an indispensable tool for gene screening using halipridization methods such as Southern blotting, Northern blotting, and Western blotting.

The applicant has already filed a patent application for a method of utilizing a radiation image conversion method using a stimulable phosphor sheet in place of the conventional radiography method using the above-mentioned radiation film in autoradiography (Japanese Unexamined Patent Publication No. 59/1999). -830
No. 57, JP 60-10174, Patent Application 1982-17
3393), a stimulable phosphor sheet is made of a stimulable phosphor, and after radiation energy is absorbed by the stimulable phosphor of the phosphor sheet, electromagnetic waves in the visible to infrared region (excitation light ) can emit radiation energy as fluorescence. According to this method, exposure time can be significantly shortened and chemical fog, which has been a problem in the past, does not occur.Furthermore, autoradiographs of radioactively labeled substances can be produced by Since the light is stored in a phosphor sheet and then read out in time series as photostimulated light, it is possible to display and record it in any form such as symbols and numbers in addition to images.

Traditionally, those who wish to analyze autoradiographs have been able to detect the distribution of radiolabeled substances on the support medium by visually judging the visualized autoradiographs and detecting the distribution of radiolabeled substances on the support medium. We have obtained knowledge of location information (and various information based on it, such as identification of polymeric substances, evaluation of their molecular weights and properties, etc.) for specific substances. For example, the DNA base sequence can be determined by visually determining the separation position for each base-specific DNA fragment or mixture thereof that has been given a radioactive label, and by mutually comparing the separation and development positions of the base-specific DNA fragments. It is determined by comparing the Therefore, a great deal of time and effort is spent on analyzing autoradiographs.

Furthermore, since it relies on the human eye, there are limits to the accuracy of the information obtained, such as the positional information obtained by analyzing an autoradiograph differing depending on the analyst.

Therefore, the present applicant obtained an autoradiograph of a radiolabeled substance distributed in at least one dimension on a support medium as a digital signal, and then subjected this digital signal to appropriate signal processing to obtain a radiolabeled substance. We have already filed a patent application for a method for automatically obtaining positional information of substances as desired symbols and/or numerical values (Japanese Patent Application Laid-Open Nos. 59-126527, 1982-126278, etc.), which is compatible with autoradiographs. When a conventional radiographic film is used, the digital signal is obtained by first visualizing the autoradiograph on the film and then reading it photoelectrically using reflected or transmitted light. When a stimulable phosphor sheet is used, an autoradiograph can be obtained by directly reading out the phosphor sheet on which the stimulable phosphor sheet has been stored.

As a result of various studies on autoradiographic measurements used for microanalysis of proteins, base sequencing of nucleic acids, etc., the present inventor discovered that radiolabeled substances to be detected separated and developed on a support medium It has been found that autoradiographs usually contain noise due to various distortions or impurities. Physically, distortion of the entire separation development pattern occurs due to insufficient alignment between the photosensitive material and the support medium during exposure, the separation development conditions are not constant, the support medium is not uniform, etc. These include the occurrence of meandering and smiling phenomena in the separation/deployment column, offset distortion that occurs depending on the attachment of the sample to the support medium, and the occurrence of noise due to radiation emitted from radioactive impurities or natural radiation.

[Summary of the Invention] The present inventor has proposed a method for analyzing an autoradiograph having two-dimensional positional information of a radiolabeled substance separated and developed on a support medium, in which a smiling phenomenon and an offset distortion occur. Even if it is a pattern, the signal processing of the digital signal corresponding to the autoradiograph is not completely automated, but the information necessary for analysis is input as appropriate, and each stage of signal processing based on this input information is By displaying images, it was possible to obtain desired position information with high accuracy.

That is, the present invention performs signal processing on a digital signal corresponding to an autoradiograph having two-dimensional positional information of a radiolabeled substance separated and developed on a support medium, thereby converting the positional information of a radiolabeled substance into a symbol. and/or obtain numerical values, the autoradiograph analysis method comprises: 1) electrically displaying the autoradiograph as an image based on the digital signal; 2) radiolabeled substances determined from the displayed image; 3) a step of extracting a signal in a certain range along the separation and expansion sequence based on input information regarding the separation and expansion sequence; 3) determining the separation and expansion region by performing one-dimensional signal processing on the extracted signal; 4) A step of superimposing and displaying the signal corresponding to the separated deployment site after correcting the separation deployment distance based on input information regarding the position of one separated deployment site determined from the display image. and 5) correcting the start position of separation development based on input information regarding the position of one separation development site determined from the display image, for the signal whose separation development distance has been corrected. The present invention provides an autoradiograph analysis method characterized by the following.

The present invention also provides a method for analyzing an autoradiograph, which comprises obtaining positional information of a radiolabeled substance as a symbol and/or numerical value by performing signal processing on a digital signal corresponding to the autoradiograph. 1) electrically displaying an image of the autoradiograph based on a digital signal; 2) a certain range along the separation line of the radiolabeled substance based on input information regarding the separation line of the radiolabeled substance determined from the displayed image; The process of extracting the signal.

3) Performing one-dimensional signal processing on the extracted signals to determine the separation development distance, and then displaying the separation in a superimposed manner; 4) One separation determined from the display image for the signal corresponding to the separation development region. 5) correcting the starting position of separation deployment based on the input information regarding the position of the deployment site, and then displaying it in a superimposed manner; The present invention also provides an autoradiograph analysis method comprising: correcting a one-separation deployment distance based on input information regarding the position of one separation deployment site.

In the present invention, "position information" refers to various types of information centering on the position of radiolabeled substances or their aggregates in a sample, such as the location and concentration of radioactive substance aggregates on a support medium. 1 distribution, identification of radioactive substances, identification of samples, etc. based on these, and various information obtained as one or any combination of information.

[Effects of the Invention] The method of the present invention semi-automatically obtains position information of a radiolabeled substance by passing a digital signal corresponding to an autoradiograph of the radiolabeled substance through an appropriate signal processing circuit having a signal processing function. It is obtained as a desired symbol and/or numerical value. The signal processing circuit then semi-automatically determines desired information, such as the base sequence of DNA, by further applying appropriate arithmetic processing and other related information to the positional information represented by symbols and/or numerical values. It is something that can be done.

In general, when analyzing autoradiographs with the various types of distortions or noises mentioned above, it is necessary to automatically recognize and correct distortions or noises on the corresponding digital image data using a high-speed, large-capacity computer. It requires an automatic electronic computer, and it is difficult to guarantee that the analysis results will always have high accuracy.

According to the method of the present invention, rather than completely automatically processing and analyzing digital signals corresponding to autoradiographs obtained by reading out photosensitive materials such as radiation films and stimulable phosphor sheets, By providing the necessary information about the various distortions mentioned above and performing appropriate and efficient signal processing based on this information, the time and labor required in the conventional case manually by analysts can be significantly reduced. At the same time, it makes it possible to perform analysis with higher accuracy and efficiency than if the analysis were completely automated. In other words, by semi-automating the analysis operation, desired position information can be easily obtained with high reliability.

That is, when providing necessary information prior to processing at each stage of signal processing, an autoradiograph based on a digital signal is displayed as an image on a CRT or the like.
The analyst can judge and provide necessary information from this displayed image. In particular, the separation and development array of radiolabeled substances on the support medium often meander due to separation and development conditions such as electrophoresis, individual differences in the support medium, exposure conditions, etc. Also, the size of each separation and development site, The shape, center position, etc. are also different. Therefore, by inputting information about the two-separation development sequence prior to digital signal processing, subsequent signal processing such as determination of the separation development region can be performed with high accuracy.

In addition, the digital image data that has been signal-processed at each stage of signal processing can be displayed overlaid on the autoradiograph before signal processing, allowing the analyst to judge from this displayed image what further signal processing is necessary. You can order the execution. For example, when a separation and development method such as electrophoresis is used, the separation and development distance of the rows at both ends of the support medium is generally longer than the separation and development distance of the rows at the center of the support medium due to heat dissipation during the separation and development process (so-called edge effect). According to the method of the present invention, by inputting information about the position of any one separation development site for each separation development row,
Correction for this smiling phenomenon can be performed easily and with high precision.

Furthermore, the shapes (sizes of recesses) of the numerous slots (sample injection ports) provided at the upper end of support media such as gel media are not completely the same, but may differ from each other, and when attaching the sample to the support medium, The adhesion positions of the samples may be shifted from each other, or the rates at which the sample penetrates the support medium may differ due to insufficient washing of urea from the support medium immediately before sample injection. As a result, since the starting position or start time of the separation and development of the sample is different in each row, mutual positional deviation between the rows (so-called offset distortion) tends to occur. By inputting information about the position, correction for this offset distortion can be performed easily and with high precision.

According to the present invention, desired position information can be obtained with high precision regardless of which of the above-mentioned smile phenomenon correction and offset distortion correction is performed first.

In addition, the digital image data processed at each stage of signal processing can be displayed on the screen, making it possible to check at each stage of signal processing whether or not the analysis is being performed appropriately. It is possible. In particular, the analyst can compare and confirm the positional information of the radiolabeled substance obtained through signal processing and the autoradiograph displayed as an image. You will be given an opportunity to make corrections.

Therefore, in the present invention, the digital signal processing for autoradiographic analysis is semi-automated, and the processing function is suitably controlled. Since Section A can be performed, autoradiographic analysis can be performed with high reliability.

[Structure of the Invention] Samples to be separated and developed in the present invention include:
Examples include macromolecular substances derived from living organisms such as proteins, nucleic acids, derivatives thereof, decomposition products thereof, and synthetic products having radioactive labels. Radioactive labels are imparted by allowing these substances to retain radioactive isotopes such as 32p, lAC, Atsushi, CoH, Al, etc. in an appropriate manner. When the sample is one of these biopolymer substances, the method of the present invention can be effectively used as a means for separating the substance and identifying its single molecular structure. However, the substances to be subjected to the autoradiographic analysis of the present invention are not limited to these polymer substances.

The radiolabeled substance as a sample is separated on the support medium by various separation and development methods such as electrophoresis, thin layer chromatography, column chromatography, and paper chromatography using various known support media such as gel support media. Be expanded.

Next, the support medium on which the radiolabeled substance has been separated and developed is subjected to radiography using a conventional photographic material.
Alternatively, an autoradiograph thereof is obtained by a radiation image conversion method using a stimulable phosphor sheet, and then a digital signal corresponding to the autoradiograph of the radiolabeled substance is obtained via an appropriate readout system.

When using the former radiographic method, first the support medium and the photographic material such as X-ray film are heated at a low temperature (-90 to -
The radiographic film is exposed to light by overlapping at 70℃ for a long time (several tens of hours), and then developed to create a visible image of the autoradiograph of the radiolabeled substance on the radiographic film.
The autoradiograph visualized on the radiographic film is then read using an image reading device. For example, an autoradiograph is obtained as an electrical signal by irradiating a radiation film with a light beam and photoelectrically detecting the transmitted or reflected light. Furthermore, by A/D converting this electrical signal, a digital signal corresponding to an autoradiograph can be obtained.

When using the latter radiation image conversion method, first, the support medium and stimulable phosphor sheet are heated at room temperature for a short period of time (several seconds to
The autoradiograph is recorded as a kind of latent image on the phosphor sheet by overlapping the phosphor sheets (for several tens of minutes) and storing the radiation energy emitted from the radiolabeled substance in the phosphor sheet. Here, the stimulable phosphor sheet includes a support made of, for example, a plastic film, divalent europium activated barium fluoride bromide (BaFBr:Eu2°
) and a transparent protective film are laminated in this order. The stimulable phosphor contained in the stimulable phosphor sheet absorbs and accumulates radiation energy when it is irradiated with radiation such as X-rays, and then accumulates when excited with light in the visible to infrared region. It has the property of emitting radiation energy as photostimulated light.

Next, the autoradiograph stored and recorded on the stimulable phosphor sheet is read out using a reading device, specifically,
For example, by scanning a phosphor sheet with a laser beam to emit radiation energy as photostimulated light, and then detecting this photostimulated light photoelectrically, an autoradiograph of a radiolabeled substance can be directly obtained without creating a visible image. obtained as an electrical signal. Furthermore, by A/D converting this electrical signal, a digital signal corresponding to an autoradiograph can be obtained.

Note that in any of the above methods, it is not necessary to read (or read out) the autoradiograph over the entire surface of the radiation film (or stimulable phosphor sheet), and it is of course possible to perform it only on the image area. .

The obtained digital signal nxy consists of the coordinates (x, y) expressed in the coordinate system fixed to the radiation film (or phosphor sheet) and the signal level (Z) at those coordinates.
The level of the signal represents the image density at that coordinate, that is, the amount of radioactive label. Therefore, the series of digital signals (ie, digital image data) has two-dimensional positional information of the radiolabeled substance.

For details of the above-mentioned autoradiograph measurement operation and method of obtaining a digital signal corresponding to the autoradiograph, see the above-mentioned JP-A-59-83057 and JP-A-59-1.
It is described in various publications such as No. 26527 and Japanese Unexamined Patent Publication No. 59-126278.

In the above, a method using conventional radiography and radiographic image conversion methods was described as a method for obtaining a digital signal corresponding to an autoradiograph of a radiolabeled substance separated and developed on a support medium. The analysis method of the present invention is not limited to these methods, and the analysis method of the present invention can be applied to digital signals obtained by any other method as long as there is a correspondence with the autoradiograph of the radiolabeled substance. It is possible.

The digital signal corresponding to the autoradiograph of the radiolabeled substance separated and developed on the support medium thus obtained is subjected to signal processing by the method of the present invention as described below, and the autoradiograph is An analysis of the graph is performed.

An embodiment of autoradiographic analysis using the method of the present invention will be described using DNA base sequencing as an example.

Base-specific DNA labeled with the following four types of radioactive labels
Regarding a sample in which a mixture of A fragments is separated and developed on a gel support medium by electrophoresis.

By using a radiation image conversion method using a stimulable phosphor sheet,
Alternatively, the autoradiograph is obtained as a digital signal by radiography using a photographic material.

1) Guanine (G) specific DNA fragment 2) Adenine (
A) Specific DNA fragment 3) Thymine (T) specific DNA
Fragment 4) Cytosine (C)-specific DNA fragment Here, each base-specific DNA fragment is a DNA fragment of various lengths that has been cleaved, degraded, or synthesized in a base-specific manner, that is, has the same terminal base. consists of things.

The digital signal is stored in a -H memory (buffer memory or nonvolatile memory such as a magnetic disk) in the signal processing circuit.

First, based on this digital signal, a display device such as a CRT connected to a signal processing circuit displays an image of the autoradiograph to be analyzed as shown in FIG. The digital signal may be subjected to signal processing (image processing) in advance to obtain a visible image with appropriate density and contrast and excellent observation and interpretation performance. Examples of image processing include spatial frequency processing, Examples include adjustment processing, averaging processing, reduction processing, and enlargement processing.

FIG. 1 shows an autoradiograph of the electrophoresis pattern obtained by electrophoresing the above-mentioned four types of base-specific DNA fragments into four slots.

Next, each electrophoresis row (lane) is determined, and the digital image data is made one-dimensional.

If the entire electrophoretic pattern displayed as an image at this time is tilted due to misalignment during the exposure operation (registration irregularity), as shown in Figure 1, rotate the pattern in advance. By doing so, an erect image (see FIG. 2) can be obtained.

FIG. 2 shows an example of an electrophoretic pattern that has been subjected to a pattern rotation process.As will be described later, offset distortion and a smiling phenomenon have occurred in the electrophoretic pattern.

To rotate a pattern, for example, the operator inputs information about the required amount of rotation (angle, direction) to a signal processing circuit based on the display screen, and then performs signal processing for pattern rotation based on this input information. It is done by doing. Information can be input by keyboard operation, joystick operation, or operation using an input member such as a light pen on the screen. Specifically, the digital signal DXF is subjected to coordinate rotation processing (X →
X', y+y') to create a digital signal I)'x
Convert to y. Digital signal D after pattern rotation
'8. The electrophoresis pattern (erect image) based on the image is displayed as an original pattern.

A cursor line is displayed on the screen for this erect image, and information about the lane of each slot is input using this cursor line. First, the operator moves the displayed cursor lines so that they overlap on one lane. To move the cursor line, use the keyboard, for example.
This can be done by joystick operation or touch pen operation on the screen. At this time, it is preferable to make the cursor line blink or display it in a different color from the image so that it can be distinguished from the lane. (area close to ). When the cursor line passes through approximately the center of each electrophoresis region (band) on the lane, information indicating that an electrophoresis column exists at the fixed position of the cursor line is input to the processing circuit by key operation.

As shown in Fig. 3(a), if the cursor line 2 does not match the lane completely because the lane l is not straight but meandering, the cursor line is corrected on the screen and the lane information is displayed. input.

Note that FIGS. 3(a) and 3(b) each show an example of the lane displayed as an image.

For example, when performing this by keyboard operation, first recognize the lowest position 3 where cursor line 2 matches lane 1, then fix the cursor line segment up to this position. Move the cursor on the line in the X direction, then move the cursor point in the X direction without changing the y coordinate, and then fix the cursor point.

By this input operation, as shown in FIG. 3(b).

A new cursor line segment connecting part M3 and this fixed cursor point position with a straight line is displayed on the screen. If this operation is repeated with sections of appropriate length and each cursor line segment passes through approximately the center point of the band over the entire lane, it is determined that an electrophoresis column exists at the position of the final cursor line. information is input. Alternatively, the cursor line can also be corrected by touching the center point of each band in a mismatched area on one screen with a touch pen.

Based on this lane information, a certain range of digital signals along each lane is extracted. Specifically, a digital signal that matches the cursor line or a digital signal that is within a certain width area from the cursor line is extracted. That is, only the digital signal D''XF having certain (x', y') coordinates based on the lane information is extracted.

This extracted digital signal D″xy is the slot number (n
) and the migration coordinate (y”). It can be expressed as y. In other words, once the slot is determined, the migration coordinates (
The digital image data, which could be represented only by ``y'') and had two-dimensional information consisting of (x', y'), has been converted to one-dimensional information for each lane. Therefore, by this signal processing, the electrophoresis A representative profile of the pattern is obtained.

Note that the profile can be confirmed by displaying an image of the representative profile obtained here superimposed on the original pattern.

Next, one-dimensional signal processing is performed on the extracted digital signal D".y for each slot to detect all bands B3a on the electrophoresis pattern. Band detection is performed, for example, as follows.
This can be done by creating a one-dimensional waveform (a graph in which the horizontal axis represents the position on the lane and the vertical axis represents the signal level) for the digital signal of each lane, and then searching for its peak. Here, the detected band But is.

It has information consisting of migration coordinate y'' and band intensity 2'' in the n-th slot. Peak B of each band
nF is superimposed and displayed on the screen as a point or a simulated band (a band perpendicular to the electrophoresis direction).

Thereafter, as shown in FIG. 2, the migration pattern in which offset distortion and smiling phenomena have occurred is corrected, for example, in the following manner.

Note that the presence or absence of offset distortion can be determined from the fact that the entire lane is vertically shifted uniformly in the displayed original pattern. In addition, the presence or absence of the smiling phenomenon can be determined by: 1) The length of the lanes at both ends of the displayed original pattern is shorter than the length of the lane at the center; This can be determined from the fact that the large lower band) is not perpendicular (that is, horizontal) to the migration direction (see Figure 2).

To correct offset distortion, move the cursor point to any band position on the lane where the offset distortion has occurred to recognize that position, and then move the cursor point upward or upward along the lane direction for the lane that is &2. Move it downward and lock it in place. Through this operation, correction information is input so that the band position becomes the position where the cursor is fixed (that is, the position where the base-specific DNA fragment would have originally migrated if no offset distortion occurred).

Specifically, the migration coordinate y' of the band can be approximated by equation (I).

y”= a t + b (I)(
However, a represents the migration speed of each band, t represents the migration time, and b represents the coordinates of the migration start point.) The above input information, that is, the actual migration coordinate and the virtual migration coordinate, are respectively ( I) By substituting into the equation, calculate the difference between the actual migration start position and the virtual migration start position b' (the migration start position when no offset distortion occurs), and calculate the migration coordinates for all bands in the slot. By applying correction.

The entire band on the lane can be translated upward or downward. This offset-corrected band B' is on the original pattern. By superimposing the
(see FIG. 4), the result of offset correction can be confirmed and used as the final input information. At this time, offset correction is not yet complete (insufficient or excessive)
In this case, the desired information can be finally input by repeating the above operation again.

FIG. 4 shows an example of an electrophoretic pattern in which offset distortion has been corrected. Therefore, only the smiling phenomenon appears in the electrophoretic pattern.

In order to reduce the number of operations described above, it is preferable that the arbitrarily selected band be the topmost band. This is because by setting the position where the cursor point should be fixed to a position horizontal to the migration start position of the lane where no offset has occurred (basic lane), correction can be easily made without being influenced by the smiling phenomenon etc. This is because it can be done.

In this way, offset correction is performed for the given slots one after another.

Next, the smiling phenomenon is corrected for the offset-corrected band B'flF.

First, the cursor is placed on the migration start point of the lane where the smiling phenomenon occurs, and the position is recognized. Next, the cursor point is placed on an arbitrary band position on the lane to recognize the position, and then the cursor point is moved downward along the lane direction and fixed at an appropriate position.

Through this operation, correction information is input so that the band position becomes the position where the cursor is fixed (that is, the position where the base-specific DNA fragment would have originally migrated if the smarming phenomenon did not occur).

The smiling phenomenon occurs because the migration speed of each band differs depending on the slot when a difference in gel temperature occurs due to the edge effect of the gel support medium. By substituting the above input information, that is, the actual migration coordinates and the virtual migration coordinates, into the above equation (I), the actual migration speed a and the virtual migration speed a' (the migration speed when the smiling phenomenon did not occur) are obtained. ), and by correcting all the bands in the slot with respect to the migration coordinates, the entire band on the lane can be stretched downward at a constant rate.

By superimposing this smiling-corrected band B'' on the original pattern (see Figure 5).
, the operator can check the smile correction results and use them as the final input information. At this time, if the smile correction is not yet complete (insufficient or excessive), the desired information can finally be input by repeating the above operation again.

FIG. 5 shows an example of an electrophoresis pattern in which the smiling phenomenon has been corrected.

In order to reduce the number of operations described above, it is preferable that the arbitrarily selected band be the bottom band, because at the bottom the amount of electrophoresis delay due to smiling is maximum and the cursor point is fixed. This is because correction can be easily performed with high accuracy by setting the correct position to be horizontal (or approximately horizontal) to the position of the final band of the central lane where almost no smiling occurs.

In this way, the smiling correction is performed for each slot in sequence. Note that it is of course possible to perform the above correction in the reverse order, that is, to perform the smiling correction first and then perform the offset correction.

Each band peak based on digital signal with offset and smile correction.

are compared between slots with respect to their migration coordinates, and the bands are sorted in order of migration coordinates. The slots (1) to (4) above each have information about the terminal bases consisting of (G), (A), (T), and (C), so replace the slot number of each band with the corresponding base. By this, D.N.
Base sequence of A (e.g. A-G-C-T-A-A-G-
) can be obtained.

If desired, by displaying the offset-corrected and smiling-corrected band peaks on the screen superimposed on the original pattern, the operator can confirm the analysis results while looking at this superimposed display screen. Furthermore, the obtained DNA base sequence information may be simultaneously displayed on the screen, and in this case, it is also possible to partially correct the analysis results as necessary.

Note that the analysis results for the DNA base sequence are not limited to the above display format; for example, the intensity (2'') of each band can be displayed simultaneously as the relative amount of the electrophoretic substance, if desired. It is also possible to display it as an image together with the visible image of the autoradiograph.

Furthermore, the information obtained in this way also includes:
For example, it is also possible to perform genetic linguistic information processing such as comparing with other DNA base sequences that have already been recorded and stored.

The analysis results for the DNA base sequence determined by the above-mentioned signal processing are output from the signal processing circuit, and then
The information is then transmitted to a recording device directly or, if necessary, via a storage means such as a magnetic disk or magnetic tape.

Recording devices include, for example, those that optically record by scanning a photosensitive material with a laser beam, etc., those that record symbols and numbers displayed on a CRT etc. on a video fist printer, etc., and those that record using heat rays. Recording devices based on various principles can be used, such as those that record on materials.

In addition, although the above description has been made by taking DNA base sequencing as an example, the autoradiographic analysis method of the present invention is not limited to base-specific DNA fragments, and the sample is not limited to base-specific DNA fragments. This method can be applied to the separation and development pattern of the radiolabeled substance separated and developed in one-dimensional direction. In particular, it can be suitably used for protein trace analysis and gene screening.

[Brief explanation of the drawing]

FIG. 1(a) is a diagram showing an example of an electrophoretic pattern in which rotational distortion, offset distortion, and smiling phenomena occur in the pattern. FIG. 2 is a diagram showing an example of a migration pattern that has been subjected to pattern rotation processing. FIG. 3(a) is a diagram illustrating an example of a lane in which the migration column is distorted, and FIG. 3(b) is a diagram showing an example of a lane in which the migration column is determined by a cursor line. FIG. 4 is a diagram showing an example of a migration pattern in which offset distortion has been corrected. FIG. 5 is a diagram showing an example of an electrophoresis pattern in which the smiling phenomenon has been corrected. l: Electrophoresis row (lane) 2: Cursor line 3: Bottom position where the cursor line and lane match 4: Cursor point Patent applicant Fuji Photo Film Co., Ltd. Agent
Person Patent Attorney Yasushi Yanagawa〉〉〉〉Duck Figure 4 Figure 5.

Claims (1)

[Claims] 1. Positional information of a radiolabeled substance separated and developed on a support medium by performing signal processing on a digital signal corresponding to an autoradiograph having two-dimensional positional information of the radiolabeled substance The method for analyzing an autoradiograph comprises: 1) electrically displaying an image of the autoradiograph based on a digital signal; 2) a radioactive label determined from the displayed image; 3) A process of extracting a signal in a certain range along the separation and expansion sequence based on input information regarding the separation and expansion sequence of the substance; 3) Determining the separation and expansion site by performing one-dimensional signal processing on the extracted signal. 4) With respect to the signal corresponding to the separated deployment site, the separation deployment distance is corrected based on the input information regarding the position of one separation deployment site determined from the display image, and then the superimposed display is performed. and 5) correcting the starting position of separation deployment based on input information regarding the position of one separation deployment site determined from the display image for the signal for which the separation deployment distance has been corrected. An autoradiographic analysis method comprising: 2. After the first step and before the second step, the pattern is rotated as a whole based on the input information regarding the inclination of the separation and development pattern of the radiolabeled substance determined from the displayed image, and then the image is displayed. An autoradiograph analysis method according to claim 1, characterized in that: 3. After the fifth step, the position information of the radiolabeled substance obtained is confirmed by superimposing the position information on the autoradiograph displayed as an image. The autoradiograph analysis method described in item 1. 4. The digital signal corresponding to the autoradiograph is transmitted to the autoradiograph of the radiolabeled substance on the support medium by superimposing the support medium and the stimulable phosphor sheet containing the stimulable phosphor. Claim 1, wherein the autoradiograph is obtained by accumulating and recording on a sheet and then photoelectrically reading out the autoradiograph as photostimulated light by irradiating the phosphor sheet with excitation light. Autoradiograph analysis method. 5. A digital signal corresponding to the autoradiograph is transferred onto the photosensitive material after superimposing the support medium and the photosensitive material to photosensitively record the autoradiograph of the radiolabeled substance on the support medium on the photosensitive material. 2. The autoradiograph analysis method according to claim 1, wherein the autoradiograph analysis method is obtained by photoelectrically reading an autoradiograph visualized in . 6. The autoclave according to claim 1, wherein the radiolabeled substance is a group of biopolymer substances to which a radiolabel is attached, a derivative thereof, a decomposition product thereof, or a composite thereof. Radiograph analysis method. 7. A patent characterized in that the biopolymer substance group is a nucleic acid, a derivative thereof, a decomposition product thereof, or a composite thereof, and the positional information obtained by signal processing represents the base sequence thereof. The autoradiograph analysis method according to claim 6. 8. By performing signal processing on the digital signal corresponding to the autoradiograph that has two-dimensional positional information of the radiolabeled substance separated and developed on the support medium, the positional information of the radiolabeled substance can be converted into symbols and/or numerical values. The autoradiograph analysis method comprises: 1) electrically displaying the autoradiograph as an image based on digital signals; 3) A step of extracting a signal in a certain range along the separation expansion column based on the input information; 3) A step of determining the separation expansion region by performing one-dimensional signal processing on the extracted signals, and then superimposing and displaying it. , 4) With respect to the signal corresponding to the separation development site, the step of correcting the start position of separation development based on input information regarding the position of one separation development site determined from the display image, and then displaying the signal in a superimposed manner; 5) A step of correcting the separation deployment distance based on input information regarding the position of one separation deployment site determined from the display image for the signal whose separation deployment start position has been corrected. Autoradiograph analysis method. 9. After the first step and before the second step, the pattern is rotated as a whole based on the input information regarding the inclination of the separation and development pattern of the radiolabeled substance determined from the displayed image, and then the image is displayed. The autoradiograph analysis method according to claim 8, characterized in that: 10. After the fifth step, the position information of the radiolabeled substance obtained is confirmed by superimposing the position information on the autoradiograph displayed as an image. The autoradiograph analysis method described in Section 8. 11. The digital signal corresponding to the autoradiograph is transmitted to the autoradiograph of the radiolabeled substance on the support medium by superimposing the support medium and the stimulable phosphor sheet containing the stimulable phosphor. Claim 8, characterized in that it is obtained by accumulating and recording on a sheet, irradiating the phosphor sheet with excitation light, and photoelectrically reading out the autoradiograph as photostimulated light. Autoradiograph analysis method. 12. A digital signal corresponding to the autoradiograph is transferred onto the photosensitive material after the autoradiograph of the radiolabeled substance on the support medium is photosensitively recorded on the photosensitive material by superimposing the support medium and the photosensitive material. 9. The autoradiograph analysis method according to claim 8, which is obtained by photoelectrically reading an autoradiograph visualized in . 13. The auto according to claim 8, wherein the radiolabeled substance is a group of biopolymer substances to which a radioactive label has been added, a derivative thereof, a decomposition product thereof, or a composite thereof. Radiograph analysis method. 14. A patent characterized in that the biopolymer substance group is a nucleic acid, a derivative thereof, a decomposition product thereof, or a composite thereof, and the positional information obtained by signal processing represents the base sequence thereof. The autoradiographic analysis method according to claim 13.