 |
Description  |
|
|
BACKGROUND OF THE INVENTION
The present invention relates to a multi-colored electrophoresis pattern
reading system and, more particularly, to a multi-colored electrophoresis
pattern reading system appropriate for the comparison of a plurality of
patterns of electrophoresis by labelling each of samples with fluorescent
pigments having different fluorescent wavelengths, subjecting the samples
to electrophoresis simultaneously, and reading the resulting patterns of
electrophoresis.
Generally, electrophoresis analysis method using samples labeled with a
radioactive isotope has been employed for analyzing the sequences of
various genes, including diagnosis of diseases deriving from genes, the
structures of proteins such as amino acids, etc. The electrophoresis
analysis method is a method for analyzing samples by labelling or
replacing fragments of a sample with or by a radioactive isotope,
subjecting the fragments of the sample to electrophoresis with a gel, and
analyzing a pattern of distribution of the fragments of the sample
developed by means of electrophoresis.
Description will now be made of the diagnosis of hereditary diseases as an
example for reading and analyzing patterns of electrophoresis. The human
gene DNA consists of pairs of bases of approximately 3.times.10.sup.9 and
the sequences of the bases are generally constant among the human beings,
although there is a deviation in the sequences of the bases among
individuals. This deviation is called a polymorphism of the DNA. The
polymorphism of the DNA is seen in the non-hereditary region as well as in
the hereditary region and the polymorphism of the DNA appears in many
occasions as a polymorphism of proteins that is a phenotype of the
polymorphism of the DNA. A variety of variations as seen among the human
beings, such as the blood types, histocompatible antigens, the difference
of skin and hair colors among peoples, etc. are based on the polymorphism
of the DNA. The polymorphism of the DNA has been created on the basis of
variations that have been accumulated in the DNA of the genocytes of the
human groups up to the present time from the time when the human beings
were developed as an individual biological species in the course of the
evolution of the human beings. When such a variation exists in the site
that has the function of significance in terms of the existence as an
individual person and when a nosogenic phenotype resulting from the
variation occurs as a pathologic state, the pathologic state is called a
hereditary disease. It is said that there are currently more than 3,000
kinds of hereditary diseases in the human group.
The nosogenesis of the hereditary disease is an abnormality appeared on the
DNA sequence. However, it is recognized for the first time as a disease in
several stages ranging from a DNA through a mRNA and proteins to
pathogenic phenotype. The diagnosis as a disease is conducted usually in
the last stage and the diagnosis can be implemented at the DNA level or at
the protein level if the disease would occur simply in the course of the
several stages as described hereinabove.
The basic technique for the diagnosis of a DNA is called Southern plotting
that basically consists of six steps:
Step 1: Extraction of a DNA as a sample;
Step 2 Fragmentation of the DNA with restricting enzymes;
Step 3: Fractionation of DNA fragments by molecular weights through gel
electrophoresis;
Step 4: Migration of DNA fractions to filter;
Step 5: Hybridization of the DNA fractions with a probe DNA (obtained by
labelling a DNA having a hemeomorphous sequence of the gene to be
detected); and
Step 6: Detection of the hybrid by autoradiography.
For the diagnosis of the hereditary diseases, the DNA extracted from any
organ is employed and the sample required for that purpose is usually the
peripheral blood of the order of several milliliters. The DNA is extracted
from the leukocytes separated from the peripheral blood as the sample.
Approximately five days are usually required from step 1 to step 6. In
diagnosing the hereditary diseases, the pattern of a fraction from a
person tested is compared with the pattern of a fraction from a normal
person. The person tested is decided as normal when the pattern of the
fraction from the person tested is determined to be identical to the
pattern of the fraction from the normal person.
Recently, attempts have been made to conduct tests by using a probe DNA
labelled with a fluorescent pigment, in place of a radioactive isotope, to
excite the fluorescent pigment and to read the pattern of electrophoresis,
from the point of view of safety and other environmental problems.
However, highly sophisticated optical and signal processing techniques
capable of detecting a faint magnitude of light are required to give a
signal-to-noise ratio as equivalent as the radioactive isotope is
employed, because the quantity of the sample required for diagnosis of the
hereditary disease and determination of the sequence of bases is of the
order of approximately 10.sup.-15 mole.
Japanese Patent Laid-open Publication (kokai) No. 62,843/1986
(corresponding to U.S. Pat. No. 4,675,059) discloses an electrophoresis
apparatus of a fluorescence detecting type, capable of detecting a minute
quantity of a sample labelled with a fluorescent pigment.
A description will now be made of such an electrophoresis apparatus based
on fluorescence detection method.
FIG. 18 is a perspective view showing an outlook of a conventional
electrophoresis apparatus of a fluorescent type. The electrophoresis
apparatus comprises a combined electrophoresis and instrumentation unit 51
for implementing electrophoresis of a sample and measuring the
distribution of fluorescence, a data processor 52 for performing data
processing on the basis of measured data, and a cable 53 connecting the
combined electrophoresis and instrumentation unit 51 to the data processor
52. The electrophoresis and instrumentation unit 51 has a door 51a through
which a gel serving as a base for performing electrophoresis for DNA
fragments and a predetermined quantity of a sample (DNA fragments) for
electrophoresis are poured into the electrophoresis and instrumentation
unit 51. As the door 51a is closed and a switch for starting
electrophoresis on an operation display panel 51b is pressed to start
electrophoresis. As the electrophoresis has been started, a monitor of the
operation display panel 51b of the electrophoresis and instrumentation
unit 51 displays an operational state. The data measured by the
electrophoresis and instrumentation unit 51 is transmitted to the data
processor 52 in which the data is processed on the basis of a
predetermined program stored in advance. The data processor 52 comprises a
computer body 54, a keyboard 55 for entering an instruction from the
operator, a display unit 56 for displaying the processing state and
results, and a printer 57 for recording the data-processed results.
FIG. 19 is a block diagram showing the configuration of the inside of the
electrophoresis and instrumentation unit. As shown in FIG. 19, an overall
configuration of the combined electrophoresis and instrumentation unit 51
(FIG. 18) comprises an electrophoresis unit section 63 and a signal
processor unit section 64. The electrophoresis unit section 63 comprises
an electrophoresis section 5 for implementing electrophoresis, a first
electrode 2a and a second electrode 2b each for applying voltage to the
electrophoresis section 5, a support plate 3 for supporting the
electrophoresis section 5 as well as the first and second electrodes 2a
and 2b, a power source unit 4 for applying voltage to the electrophoresis
section 5, a light source 11 for emitting light for exciting a fluorescent
substance, an optical fiber 12 for leading the light from the light source
11, a light collector 14 of an optic system for condensing and collecting
fluorescence 13 generated by the fluorescent substance, an optical filter
15 for selectively passing the light having a particular wavelength
therethrough, and an optical sensor 16 for converting the condensed light
into electric signals. The signal processor unit section 64 comprises an
amplifier 17 for amplifying the electric signals from the optical sensor
16, an analog-digital converting circuit 18 for converting analog signals
of the electric signals into digital data, a signal processing section 19
for pre-processing the digital data, for example, by performing addition
average processing or the like, an interface 20 for implementing interface
processing for feeding the pre-processed data to an external data
processor, and a control circuit 10 for implementing overall control of
the electrophoresis unit section and the signal processing system. The
digital signal OUT generated from the signal processor unit section 64 is
transmitted to the data processing unit 52 (FIG. 18), thereby implementing
the data processing such as analysis processing and so on.
A description will now be made of the operation of the electrophoresis
apparatus with reference to FIGS. 18 and 19.
After the door 51a of the electrophoresis and instrumentation unit 51 is
opened, a gel is poured into the electrophoresis section 5 disposed within
the combined electrophoresis and instrumentation unit 51 and thereafter a
sample of DNA fragments labelled with a fluorescent substance is poured
thereinto. As a switch of the instrument panel 51b is pressed to give an
instruction to start electrophoresis, then voltage is applied from the
first and second electrodes 2a and 2b of the power source unit 4 to the
electrophoresis section 5, thereby starting the electrophoresis. The
electrophoresis allows the sample labelled with the fluorescent substance
to migrate, for example, in lanes 71, 72, 73 and 74, as shown in the
schematic representation 70 of FIG. 22, gathering the molecules having the
same molecular weights together forming bands 66. The molecules having
lower molecular weights migrate faster than those having higher molecular
weights so that the former migrates in a distance longer than the latter
within the same time unit. The bands 66 are detected in a manner as shown
in FIG. 20a by leading light from the light source through the optical
fiber 12 to a light path 61 and irradiating the gel on the light path 61
with the light, exciting the labelled fluorescent substance concentrated
on the bands 66 in the gel to generate fluorescence 13, and detecting the
fluorescence 13. The fluorescence 13 generated contains the fluorescent
substance in the concentration as extremely low as approximately
10.sup.-16 mole per band, although the quantity of fluorescence may depend
upon an extinction coefficient of the fluorescent substance used, quantum
efficiency thereof, intensity of exciting light, etc. For instance,
fluorescein isothiocyanate has a peak of the wavelength of excitation at
490 nm, a peak of its fluorescent wavelength of 520 nm, an extinction
coefficient of 7.times.104 mole.sup.-1. mole.sup.-1, and a quantum
efficiency of approximately 0.65. If fluorescein isothiocyanate employed
exists in the concentration of 10.sup.-16 mole per band, the fluorescence
generated contains photons of the order as low as 10.sup.10 /S, when
calculated by postulating the use of argon ion laser of a wavelength of
488 nm at output of 1 mW as a fluorescent substance, although it may vary
to some extent with the thickness of the gel or the like. Hence, an
extremely faint magnitude of fluorescence is required to be detected.
Referring to a front view as shown in FIG. 20a and to a longitudinally
sectional view as shown in FIG. 20b, the electrophoresis section 5
comprises a gel member 5a composed of polyacrylamide or the like and gel
supporting members 5b and 5c, each made of glass for supporting and
interposing the gel member 5a from the both sides. A sample of DNA
fragments is poured into the gel member 5a of the electrophoresis section
5 from its upper portion and the electrophoresis is carried out by
applying voltage for electrophoresis to the first electrode 2a and the
second electrode 2b (FIG. 18). While the electrophoresis is being carried
out, the fluorescent substance contained in the bands of the pattern of
electrophoresis in the gel member 5a along the light path 61 is irradiated
with rays of light sent out from the light source, such as laser light,
which pass through the optical fiber 12 onto the light path 61 of the gel
member 5e. This allows the fluorescent substance present on the light path
61 to be excited to emit fluorescence 13 that is led to a light collector
14 of optics consisting of a combination of lenses and then selected by
the optical filter 15 after having been condensed, followed by conversion
into electric signals by means of a one-dimensional optical sensor 16. In
order to convert a faint quantity of light into electric signals in an
efficient fashion, the light is amplified to 10.sup.4 to 10.sup.5 times
with an image intensifier or the like, and the image is converted into
electric signals by the optical sensor 16, such as a one-dimensional
optical sensor of CCD or the like. The electric signals converted by the
optical sensor 16 are then amplified to signals of a desired level by the
amplifier 17, and the analog signals are converted into digital signals by
the analog-digital converting circuit 18, followed by transmission to the
signal processing section 19. The digital signals transmitted from the
analog-digital converting circuit 18 are then subjected to signal
processing, such as addition average processing, or the like, in order to
improve the signal-to-noise ratio (an S/N ratio), and the resulting
digital data is transmitted to the data processor unit section 52 through
the interface 20.
FIGS. 21a and 21b are schematic representations for describing an example
of signals of a pattern indicative of the fluorescent intensity of DNA
fragments transmitted from the electrophoresis and instrumentation unit
51. For instance, as shown in FIG. 21a, the fluorescent substance present
on the light path 61 is excited upon irradiation of the gel member 5a of
the electrophoresis section 5 with the laser light in the course of
electrophoresis, thereby emitting fluorescence. The fluorescence is
detected at predetermined positions of each lane in the direction of
electrophoresis, as indicated by 62, as the time of electrophoresis
elapses. In other words, the fluorescence is detected as the bands 66 of
each lane pass through the positions of the light path 61, thereby
detecting a pattern signal of fluorescence intensity in each of the lanes,
as shown in FIG. 21b. As a peak of the fluorescence intensity is given
When each of the bands 66 passes through the positions of the light path
61, the pattern signal of the fluorescence intensity as shown in FIG. 21b
represents a pattern signal indicating the magnitude of fluorescence
intensity of the bands 66 located in the direction of electrophoresis, as
indicated by 62. In other words, the pattern signal constitutes a profile
wave form proportional to the concentration of fluorescence, and a
sequence of the bases of a DNA fragment may be determined by deciding a
peak value of the pattern signal.
The computer body 54 of the data processing unit 52 implements data
processing for comparing molecular weights and determining a sequence of
bases of a DNA fragment on the basis of data of the pattern indicative of
fluorescence intensity in response to data of the pattern signals for the
fluorescence intensity of the DNA fragments transmitted from the
electrophoresis and instrumentation unit 51. The sequence of the bases and
so on determined by the data processing is symbolized and then generated,
thereby displaying the symbolized data on a display screen by the display
unit 56 or printing it out by the printer 57.
The aforesaid embodiment is directed to an example of the apparatus in
which the fluorescent pigment is employed for labelling the sample.
Japanese Patent Laid-open Publication (kokai) No. 167,649/1989 discloses
another example of an apparatus capable of reading a fluorescent pattern
of electrophoresis. This apparatus is of such a type as reading a
fluorescent pattern of the electrophoresis section as a whole after the
end of electrophoresis, unlike the aforesaid electrophoresis apparatus of
such a type as reading the distribution of the fluorescent pattern passing
through a reading section in the course of electrophoresis.
It is to be noted herein that the gel electrophoresis method employed for
the electrophoresis pattern reading apparatuses on the basis of the
fluorescence detection method is the same as the gel electrophoresis
method which has been employed for the conventional apparatuses in which
the sample is labeled with the radioactive isotope. The gel
electrophoresis method may cause a warp in the pattern of electrophoresis
because a speed of migration of bands may vary with the position of an
electrophoresing plate due to irregularities in temperatures within the
gel and for other reasons, thereby causing a warp in the pattern of
electrophoresis. Hence, for example, when electrophoresis of two kinds of
samples or electrophoresis in a two-dimensional way is to be performed
with the purpose to compare two kinds of patterns of electrophoresis for
the diagnosis of hereditary diseases, the positions of the electrophoresed
bands may be deviated between the results of electrophoresis due to the
warp and a comparison between the two patterns of electrophoresis may be
rendered difficult, even if either of the methods for labelling the
samples with the radioactive isotope or with the fluorescent substance
would be adopted, as long as the conventional gel electrophoresis method
is employed. Further, implementation of the correction of such patterns of
electrophoresis by means of data processing is also rendered laborious and
complex.
Further, as the electrophoresis and instrumentation unit for implementing
electrophoresis and simultaneously measuring the distribution of the
fluorescent substance passing through the reading unit adopts
two-dimensional electrophoresis, it requires the one-dimensional
electrophoresis to be implemented by one device and the two-dimensional
electrophoresis to be conducted by another device, so that this operation
is laborious and complex.
SUMMARY OF THE INVENTION
The object of the present invention is to provide a multi-colored
electrophoresis pattern reading system suitable for a comparison between a
plurality of patterns of electrophoresis by labelling each of plural
samples with a fluorescent pigment having a different fluorescence
wavelength, subjecting the plural samples to electrophoresis
simultaneously, and reading the resulting patterns of electrophoresis.
Another object of the present invention is to provide a multi-colored
electrophoresis pattern reading system capable of reading the fluorescent
patterns of the patterns of electrophoresis and comparing the patterns of
electrophoresis without undergoing a warp resulting from electrophoresis.
In order to achieve the aforesaid objects, the present invention consists
of a multi-colored electrophoresis pattern reading system capable of
labelling each of plural samples separately with each of plural
fluorescent substances having a different fluorescent wavelength,
subjecting the plural samples to electrophoresis to develop a pattern of
electrophoresis, exciting the fluorescent substances labelled on the
respective plural samples to emit fluorescence, and reading a fluorescent
pattern emitting the fluorescence, characterized by: a light source means
for irradiating the pattern of electrophoresis with irradiating light for
exciting the fluorescent substance labelled on the sample; a light
scanning means for scanning the irradiating light from the light source
means and irradiating a gel in the direction of thickness of the gel with
the irradiating light; a light receiving means for receiving the
fluorescence resulting from the pattern of electrophoresis by separating
from scattered light resulting from a reading surface on the basis of a
position relationship of a light receiving path by setting a light
receiving surface in a direction different from an optical axis of
irradiating light; a fluorescent-wavelength separating means for
separating fluorescent having a predetermined wavelength from optical
signals received by the light receiving means by the aid of an optical
filter capable of controlling an angle of incidence relative to the
optical axis of the optical filter; an optoelectric conversion means for
generating electric signals by subjecting the optic signals passed through
the fluorescent-wavelength separating means to optoelectric conversion;
and a signal processing means for processing the electric signals from the
optoelectric conversion means to thereby convert them into a predetermined
form of data representation.
The light source means is a light source for generating irradiating light
for emitting fluorescence by exciting two or more fluorescent substances
and more labelled separately on the samples. In order to give light of
wavelength for exciting each of the fluorescent substances, for example, a
plurality of light sources may be provided to generate a mixture of light
from the plurality of the light sources. The light source means may be a
single light source when the light resulting from the single light source
has a predetermined range of wavelengths. The light scanning means is
arranged to scan the irradiating light from the light source means and
radiate in the direction of thickness of the gel. The light receiving
means has its light receiving surface set in the direction different of
the optical axis of the irradiating light and is designed so as to receive
the fluorescence from the pattern of electrophoresis separated from the
scattered light from the reading surface due to the position relationship
of a light receiving path. The fluorescent-wavelength separating means has
the optical filter capable of controlling the angle of incidence of
fluorescence relative to the optical axis of the optical filter and it is
to separate the predetermined fluorescent wavelength from the optical
signals received by the light receiving means by taking advantage of
dependence of the optical filter upon a pass band angle by controlling the
angle of incidence of fluorescence to the optical filter. The optoelectric
conversion means generates the electric signals by optoelectrically
converting each of the optical signals separated by the
fluorescent-wavelength separating means and the signal processing section
processes the electric signals from the optoelectric conversion means,
thereby converting them into the predetermined form of data
representation.
The signal processing section has an integral circuit composed of a
condenser and a switch for controlling an integral operation and it can
control the integral operation of the integral circuit in synchronization
with the scanning of the irradiated light by the optically scanning
mechanism, and the integral time and the number of scans for reading may
be altered in accordance with the angle of incidence of the optical
signals controlled by the optical filter of the fluorescent-wavelength
separating section. This operation allows the integral operation for the
faint electric signals so as to correspond to the scanning of the
irradiated light. At this time, the speed of the integral operation
corresponds to the speed of scanning the irradiated light, so that the
pattern of electrophoresis can be read by amplifying faint outputs of
fluorescence in an efficient way.
The multi-colored electrophoresis pattern reading system having the
configuration as described above can read the distribution of the
fluorescent substances by the difference in wavelength of fluorescence
inherent in the fluorescent pigments from the patterns of electrophoresis
of the samples. Further, the multi-colored electrophoresis pattern reading
system according to the present invention allows all the samples,
including the comparative sample, to undergo an equal degree of a warp due
to electrophoresis, so that the results of electrophoresis can be read
without paying attention to the warp originating from electrophoresis.
Other objects, features and advantages of the present invention will become
apparent in the course of the description of the preferred embodiments,
which follows, with reference to the accompanying drawings.
BRIEF DESCRIPTION OF THE DRAWINGS
FIG. 1 is a schematic representation showing an overall configuration of
the electrophoresis pattern reading system of a fluorescent type according
to an embodiment of the present invention.
FIG. 2 is a block diagram showing the configuration of the essential
portion of the instrumentation section body.
FIG. 3 is a view showing the position in which an electrophoresis unit to
be mounted to the instrumentation section body.
FIG. 4 is a view showing the light scanning mechanism for scanning a gel
surface with laser beams by using a vibrating mirror.
FIG. 5 is a graph showing the relationship between the angles of rotation
of the vibrating mirror and the distance in which spot light of the laser
beams moves.
FIG. 6 is a block diagram showing the configuration of the essential
portion of the control circuit for controlling a mirror driver for
controlling the rotation of the vibrating mirror.
FIG. 7 is a schematic representation showing the detail configuration of
the optic system in a light collector and an optoelectric conversion
section.
FIG. 8 is a schematic representation showing an example of a mechanism for
driving the rotation of an optical filter capable of changing an angle of
incidence of fluorescence relative to the optical axis of the optical
filter.
FIG. 9 is a schematic representation showing another example of the
configuration of a simplified optoelectric conversion section.
FIG. 10 is a circuit diagram showing the circuit configuration of an
amplifier containing an integral circuit.
FIG. 11 is a time chart showing the timing of reading operations by the
amplifier.
FIG. 12 is a schematic representation showing a typical sequence
representing the relationship among the angle characteristic .THETA. of
the optical filter 24d, the distribution X of the wavelengths of the
fluorescent intensity at the pixels measured, the noise .eta. at the time
of measurement, and the measured value .PSI..
FIG. 13 is a schematic representation showing an example of the method for
reading a sample labelled with a fluorescent probe and transcribed on a
thin film.
FIG. 14 is a schematic representation showing an example of correcting the
scanning by signal processing by the optically scanning mechanism for
scanning the mirror at equally angular speeds.
FIG. 15 is a perspective view describing the electrophoresis and reading
unit for reading the patterns of electrophoresis as well as for
implementing electrophoresis.
FIG. 16 is a flowchart showing the processing for determining the sequence
of bases of a DNA fragment.
FIG. 17 is a schematic representation showing a system configuration in
which a plurality of reading units of FIG. 15 are interconnected with each
other.
FIG. 18 is a perspective view showing an outlook of a conventional
electrophoresis apparatus of a fluorescent type.
FIG. 19 is a block diagram showing the configuration of the inside of the
electrophoresis and instrumentation unit of the conventional
electrophoresis pattern reading apparatus.
FIGS. 20a and 20b are elevational view and a longitudinally sectional view
showing the electrophoresis unit, respectively, in order to describe the
principle of the operations for detecting the pattern of electrophoresis
by the fluorescence method.
FIGS. 21a and 21b are schematic representations showing an example of
pattern signals of fluorescent intensity of DNA fragments to be generated
from the electrophoresis and instrumentation unit.
FIG. 22 is a schematic representation showing an example of distribution of
DNA fragments electrophoresed.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
FIG. 1 is a schematic representation showing the overall configuration of
the electrophoresis pattern reading system of a fluorescent type according
to an embodiment of the present invention.
As shown in FIG. 1, the multi-colored electrophoresis pattern reading
system of a fluorescent type according to the present invention comprises
a combination in which an electrophoresis unit 1 is interconnected to a
reading unit 6 disposed separately from the electrophoresis unit 1. The
electrophoresis unit 1 comprises an electrophoresis unit section 5, a
first electrode 2a, a second electrode 2b, a supporting plate member 3,
and a power source 4 for electrophoresis. The electrophoresis unit section
5 consists of a gel member serving as a base for electrophoresis and a gel
support member for supporting the gel member by a glass panel or the like
for interposing the gel member and it is mounted to the first and second
electrodes 2a and 2b which in turn apply voltage for electrophoresis to
the electrophoresis unit section 5. The supporting plate member 3 is
arranged to support the electrophoresis unit section 5 as well as the
first and second electrodes 2a and 2b. The power source 4 is to supply the
voltage for electrophoresis. As described hereinabove, the electrophoresis
unit section 5 is composed of the gel member for developing a sample for
electrophoresis, such as polyacryl amide or the like, and the gel support
member for supporting the gel member interposed from both sides by the
glass plate panels or the like (see FIGS. 20a and 20b).
A sample of DNA fragments to be electrophoresed is fed from an upper
portion of the gel member of the electrophoresis unit section 5 mounted to
the electrophoresis unit 1, the voltage for electrophoresis is applied to
the first and second electrodes 2a and 2b from the power source 4, thereby
enabling electrophoresis of the sample to give a pattern of
electrophoresis. The electrophoresis unit section 5 is removed or detached
from the electrophoresis unit 1 after electrophoresis has been finished
and mounted to the reading unit 6 for reading the resulting pattern of
electrophoresis.
As the reading unit 6 is mounted to an instrumentation unit body 7 of the
instrumentation unit in a state in which the electrophoresis unit 5 is
subjected to electrophoresis or in a state in which only the gel member is
removed from the electrophoresis unit section 5, the resulting pattern of
electrophoresis is read and data is then processed by the reading unit 6.
As shown in FIG. 1, the reading unit 6 has the instrumentation unit body 7
as an essential portion, and a data processor 8, an image printer 9 and
other accessories such as a magneto-optical disk unit 58 mounted to the
instrumentation unit body 7. The data processor 8 is arranged to implement
data processing, image processing and determination processing for the
data resulting from the electrophoresis pattern read by the
instrumentation unit body 7. The image printer 9 is to process and print
the read electrophoresis pattern data out. The instrumentation unit body 7
has a reading table disposed immediately below a lid 7a mounted at the
upper portion of the instrumentation unit body for reading the pattern of
electrophoresis from the electrophoresis unit section 5 consisting of the
gel member and the gel support member, wherein the electrophoresis is
performed.
After the electrophoresis unit section 5 is detached from the
electrophoresis unit 1 after electrophoresis, the lid 7a disposed at the
upper portion of the instrumentation unit body 7 is opened and the
electrophoresis unit section 5 is then mounted to the reading table. After
mounting the electrophoresis unit section 5 to the reading table, the lid
7a is closed and a start switch for starting the reading of the pattern of
electrophoresis on an operational display panel 7b of the instrumentation
unit body 7 is pressed, thereby starting reading the pattern of
electrophoresis of the gel member of the electrophoresis unit section 5.
As the reading of the electrophoresis pattern starts, the scanning of the
irradiating light from a spot light source built in the instrumentation
unit body 7 is started and the gel member of the electrophoresis unit
section 5 is irradiated with the light for exciting a fluorescent
substance, thereby emitting fluorescence. The fluorescence emitted upon
irradiation with the light is received, and a pattern of distribution of
the fluorescent substance is measured. The data processor 8 implements
data processing of the data read and measured by the instrumentation unit
body 7 and further controls the instrumentation unit body 7 itself. The
processed data is printed out by the image printer 9. In this embodiment,
the image printer 9 to be employed is of a type capable of printing with a
plurality of colors, thereby permitting the electrophoresis pattern images
to be printed out with multiple colors so as to correspond to the samples.
FIG. 2 is a block diagram showing the configuration of the essential
portion of the instrumentation unit body 7, and FIG. 3 is a view showing
the position in which the electrophoresis unit to be mounted to the
instrumentation section body 7.
In performing the analysis of each of plural samples by means of
electrophoresis with the multi-colored electrophoresis pattern reading
system of a fluorescent type, the samples of DNA fragments labelled with
the fluorescent pigments or fluorescent substances are subjected to
electrophoresis with the electrophoresis unit 1 for a predetermined period
of time, for example, as long as approximately 5 hours. After the
electrophoresis has been finished, the electrophoresis unit section 5 is
detached from the electrophoresis unit 1 and the gel member of the
electrophoresis unit section 5 removed therefrom is then mounted to an
upper portion of the reading table 7c through the lid 7a of the
instrumentation unit body 7 of the reading unit 6, as shown in FIG. 3, in
such a state that the gel member is still interposed with the gel support
member, such as glass plates, or in such a state that the gel support
member is detached from the electrophoresis unit section 5. Then, the lid
7a is closed, thereby finishing the setting of the electrophoresis sample
to the reading unit. When no gel member is yet labelled with the
fluorescent pigment after electrophoresis, the gel member may be labelled
therewith in this stage of mounting the pattern of electrophoresis. The
gel may be dried before mounting to the reading table.
Then, operations are performed for instructing the start of reading the
pattern of electrophoresis by pressing the read starting switch of the
operation display panel 7b or by giving an instruction to start reading
from the data processor 8. In starting the reading operations through the
data processor 8, the state of mounting the electrophoresis unit section 5
to the instrumentation unit body 7 is transmitted through a control signal
line to the data processor 8 which in turn controls the operations of the
reading unit section of the instrumentation unit body 7 after the state in
which the electrophoresis unit section 5 is mounted has been confirmed. In
this case, parameters such as reading speed and so on during operations
may be set and registered in advance on the side of the data processor 8,
thereby allowing the operations for starting the reading to be performed
automatically and reducing burdens for operating the switches on the side
of an operator. The read data on the distribution of the fluorescent
pigments is transmitted to the data processor 8 which in turn implements
desired processing programmed in advance, such as processing for detecting
a peak of the intensity of fluorescence, electrophoresis distance, and so
on. The data of the processed results is printed out, when needed, by the
image printer 9 as image having a shade of color in accordance with the
intensity of fluorescence or as image in which the intensity of
fluorescence is divided by contour lines, colors or concentrations of
color. The image having the shade of color in accordance with the
intensity of fluorescence looks equal to an X-ray film image of data
obtained by labelling the sample with a radioactive isotope in
conventional manner and subjecting the sample to electrophoresis. The data
of the results after data processing may be stored, when needed, as
digital data in a magnetically or optically recording device.
Referring to FIG. 2 showing the configuration of the instrumentation unit
body 7, laser beams, as indicated by 31, emitted from the light source 21
are scanned in the direction from the front to the rear in the drawing
with the vibrating mirror 22 to be driven by the mirror driver 30 and the
gel member as an object of reading is exposed to the laser beams 31. The
spot lights of the laser beams 31 scanned by the vibrating mirror
irradiates the gel member of the electrophoresis unit section 5 in the
direction of thickness of the gel member thereof while moving. The gel
member of the electrophoresis unit section 5 emits fluorescence upon
irradiation with the spot lights of the scanned laser beams 31. The
resulting fluorescence, as indicated by 13, with the spot lights of the
| | |
