U.S. patent application number 11/296487 was filed with the patent office on 2006-04-27 for capillary electrophoretic instrument and capillary array assembly.
Invention is credited to Kazumichi Imai, Masaya Kojima, Muneo Maeshima, Satoshi Takahashi, Hiromi Yamashita.
Application Number | 20060086612 11/296487 |
Document ID | / |
Family ID | 17559301 |
Filed Date | 2006-04-27 |
United States Patent
Application |
20060086612 |
Kind Code |
A1 |
Maeshima; Muneo ; et
al. |
April 27, 2006 |
Capillary electrophoretic instrument and capillary array
assembly
Abstract
The troublesomeness during the setting of a plurality of
capillaries is eliminated by composing pairs of electrodes, which
are electrically connected to the common electrode, and
capillaries. By bringing electrodes installed in the vicinity of
each capillary disposed at the pitch of wells on the side of sample
plate (within the area of the wells) into electrical contact with a
common electrode, the capillaries and electrodes are made integral
in construction. When a voltage is applied to the electrophoretic
instrument via a common electrode portion, the voltage is applied
to the electrodes for each capillary. This enables an inexpensive
microtiter plate, etc. to be used and a multiple of capillaries to
be simultaneously inserted, attached and detached.
Inventors: |
Maeshima; Muneo; (Mito,
JP) ; Imai; Kazumichi; (Hitachinaka, JP) ;
Kojima; Masaya; (Mito, JP) ; Takahashi; Satoshi;
(Hitachinaka, JP) ; Yamashita; Hiromi; (Ishioka,
JP) |
Correspondence
Address: |
KENYON & KENYON LLP
1500 K STREET N.W.
SUITE 700
WASHINGTON
DC
20005
US
|
Family ID: |
17559301 |
Appl. No.: |
11/296487 |
Filed: |
December 8, 2005 |
Related U.S. Patent Documents
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Application
Number |
Filing Date |
Patent Number |
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10413540 |
Apr 15, 2003 |
|
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11296487 |
Dec 8, 2005 |
|
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|
09671818 |
Sep 27, 2000 |
6572752 |
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10413540 |
Apr 15, 2003 |
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Current U.S.
Class: |
204/601 |
Current CPC
Class: |
G01N 27/44743 20130101;
G01N 27/44713 20130101; G01N 27/447 20130101; G01N 27/44704
20130101; G01N 27/44782 20130101 |
Class at
Publication: |
204/601 |
International
Class: |
G01N 27/447 20060101
G01N027/447 |
Foreign Application Data
Date |
Code |
Application Number |
Sep 29, 1999 |
JP |
11-275710 |
Claims
1. A capillary assembly to be inserted into a well so that an
electrophoretic lane for a sample is formed in the well,
comprising: a capillary to be inserted into the well; a cylindrical
electrode having an inner circumference receiving the capillary
therein; and a capillary holder for integrally holding the
capillary and the electrode, wherein the inner circumference has
almost the same diameter as an outer diameter of the capillary so
that the capillary is capable of being inserted into the
cylindrical electrode, and a gap between the inner circumference
and the capillary is limited to prevent another sample from
remaining in the gap.
2. A capillary assembly according to claim 1, further comprising a
bond with which the gap is filled so that the gap is limited to
prevent another sample from remaining in the gap.
3. A capillary assembly according to claim 1, wherein the
cylindrical electrode is made of stainless steel pipe.
4. A capillary electrophoretic instrument in which a capillary
assembly is adapted to be inserted into a well so that an
electrophoretic lane for a sample is formed in the well,
comprising: a capillary to be inserted into the well; a cylindrical
electrode having an inner circumference receiving the capillary
therein; a capillary holder for integrally holding the capillary
and the electrode to form the capillary assembly; a power source
for electrically energizing the cylindrical electrode; a light
analyzer for irradiating and exciting the sample in the
electrophoretic lane with laser light, and detecting light emitted
by the excitation; and a controller for identifying a type of the
sample from an output signal of the light analyzer; wherein the
inner circumference has almost the same diameter as an outer
diameter of the capillary so that the capillary is capable of being
inserted into the cylindrical electrode, and a gap between the
inner circumference and the capillary is limited to prevent another
sample from remaining in the gap.
5. A capillary electrophoretic instrument according to claim 4,
further comprising a bond with which the gap is filled so that the
gap is limited to prevent another sample from remaining in the
gap.
6. A capillary electrophoretic instrument according to claim 4,
wherein the cylindrical electrode is made of stainless steel pipe.
Description
BACKGROUND OF THE INVENTION
[0001] 1. Field of the Invention
[0002] The present invention relates to a capillary electrophoretic
instrument having a capillary array assembly comprising a plurality
of capillaries and, more particularly, to a capillary
electrophoretic instrument (hereinafter referred to as an
electrophoretic instrument) suitable for use in such as a DNA
sequencer (a DNA base sequence analyzer) for analyzing samples of
living organism, especially through the use of a plurality of
capillaries or minute passages as a medium of electrophoretic
separation, and to a capillary array assembly used in the
instrument.
[0003] 2. Description of the Prior Art
[0004] In the base sequence determination of DNA having a very long
base sequence, a shift is occurring from a conventional flat-plate
gel method, in which a gel is sandwiched between two glass plates
and DNA, which is a sample, is caused to migrate
electrophoretically by applying a voltage across both ends of the
glass plates, to a capillary electrophoretic instrument, in which a
gel is filled in capillaries made of quartz (hereinafter referred
to as capillaries) and a sample is caused to migrate
electrophoretically by applying a voltage across both ends of the
capillary array assembly.
[0005] The above capillary electrophoretic instrument, which
permits high-speed and high-sensitivity analyses in comparison with
the flat-plate gel method and is less affected by the Joule heat
from self-heat generation by a migration current, can provide a
good resolution for an electrophoretic analysis.
[0006] In recent years, in order to increase the number of analyses
per unit time allowed in one electrophoretic instrument,
electrophoretic instruments in which a multiple of capillaries are
set and the DNA analyses of a multiple of samples can be
simultaneously performed have been coming into widespread use.
[0007] In many of these instruments, as a method for applying a
voltage to the sample loading side of each capillary during sample
loading into capillaries or during electrophoresis, a sample plate
in which samples are set and a buffer tank for electrophoresis
themselves are made of a conductor, such as a metal, or electrodes
are embedded in the sample plate and buffer tank.
[0008] As in the art described in JP-A-10-206382, there is also a
method in which an electrophoretic instrument has such an electrode
structure that an electrode covers the area surrounding the sample
loading portion of each capillary and electrophoresis is performed
by applying a high voltage to the electrophoretic instrument via a
wiring pattern connected to each electrode.
[0009] In the above-mentioned technique in which the sample plate
and buffer tank for electrophoresis themselves are made of a
conductor such as a metal, an analyst must have ready a large
number of sample plates having a voltage application structure
peculiar to each DNA analyzer as mentioned above for the NDA
analyses of a large number of samples. This has posed the problems
of increased running costs related to analyses and increased burden
on analysts.
[0010] Next, it is desirable that a general-purpose
microtiter-plate, etc. is capable of being used in an
electrophoretic instrument. Of course, however, this
microtiter-plate is not provided with an electrode portion capable
of being connected to the electrophoretic instrument. For this
reason, a technique for incorporating electrodes in an
electrophoretic instrument cannot be used.
[0011] Furthermore, in an electrophoretic instrument which has an
electrode structure portion covering the area surrounding the
sample loading portion of each capillary and is provided with a
wiring pattern connected to each electrode structure portion and to
which a high voltage is applied, the capillary replacement work is
very troublesome.
[0012] In addition, capillaries have a short life and in some
applications it is necessary to perform analyses by resetting
capillaries of different lengths. On this occasion, an analyst must
set a multiple of capillaries one after another in the
electrophoretic instrument, posing the problem of much expense in
time and effort.
[0013] Further, in this method, in order to load a sample into the
capillaries, cylindrical electrodes are beforehand brought into
contact with the sample in the sample plate, and then by driving
and moving the sample plate, the above capillaries are inserted
into the cylindrical electrodes, thereby to bring the capillaries
into contact with the sample. Therefore, in order to simultaneously
insert a plurality of capillaries with an outer diameter of several
hundreds of micrometers into cylindrical electrodes, extremely high
accuracy must be required of a driving portion of the sample plate
or the cylindrical electrodes must have an inner diameter with a
sufficient allowance.
[0014] When accuracy is given to the above driving portion by
reducing the diameter of the cylinder of above electrode, a sample
measured last time remains in a gap between the inner surface of
the cylindrical electrode and the outer surface of the capillary
due to the capillary phenomenon, posing the problem that a
good-accuracy electrophoretic analysis is impossible.
[0015] When accuracy is not required of the driving portion by
increasing the diameter of the cylinder of the electrode, the
possibility of mixing of other samples due to the above capillary
phenomenon decreases. However, this case poses the problem that
because of the large diameter of the electrode, the bottom end of
the electrode does not reach the well bottom of the sample plate
with such an inverted cone shape that the well becomes narrower
toward the bottom.
[0016] Because the bottom end of above electrode does not reach the
well bottom, it does not come into contact with a sample or a
buffer solution, with the result that in principle, sample loading
and electrophoresis are impossible. Therefore, in a case where the
electrode is to be brought into contact with the above sample and
buffer solution and a general-purpose microtiter-plate is to be
used, a minimum amount of sample must be set at a large value in
order to raise the liquid level of the sample and buffer solution,
thus posing another problem.
SUMMARY OF THE INVENTION
[0017] The present invention was made in order to solve these
problems with the prior art. Accordingly, a first object of the
present invention is to provide a capillary electrophoretic
instrument that reduces running costs related to analyses and
burdens on analysts, facilitates the replacement work and setting
of capillaries, permits good-accuracy analyses, and enables minimum
amounts of sample to be set at small values.
[0018] A second object of the present invention is to provide a
capillary array that reduces running costs related to analyses and
burdens on analysts, facilitates the replacement work and setting
of capillaries, permits good-accuracy analyses, and enables minimum
amounts of sample to be set at small values. According to an
embodiment of the present invention, there is provided a capillary
electrophoretic instrument which comprises: a capillary array
assembly comprising a capillary array having a plurality of
capillaries each of which has a bore for containing a separation
medium and forming an electrophoretic lane, each of which has a
sample loading port at one end thereof and a sample detection port
remote from the sample loading port; a plurality of electrodes each
of which is so disposed as to form a pair with the each of the
capillaries at the position near the sample loading port; an
electroconductive member, connected to a power source, for
electrically connecting the plurality of electrodes; and a
capillary array holder for holding the capillary array, the
electrodes and the electroconductive member, a plurality of sample
holders for holding a sample to be analyzed, each of which is
located at a position corresponding to each of the pairs of said
capillaries and the electrodes, and a sample moving table for
moving sand supporting the plurality of sample holders.
[0019] Further, according to another embodiment of the present
invention, there is provided a capillary array assembly that
comprises: a capillary array, which has a plurality of capillaries
forming electrophoretic lanes and constituting a sample loading
port at one end thereof and a sample detection port remote from the
sample loading port; and a capillary array assembly, which has a
plurality of electrode members installed to form a pair with each
of the above plurality of capillaries in the above sample loading
port, an electroconductive member electrophoretically connected to
the above plurality of electrode members and connected to a power
supply for applying a voltage to the above capillary array, and a
capillary array holder for holding the above capillary array,
electrode members and electroconductive member.
[0020] There is provided, as a first aspect of the present
invention stated in claim 2, a capillary electrophoretic instrument
that comprises, at least; a sample plate having a plurality of
wells for housing a sample; a buffer tank for housing a buffer
solution for effecting electrophoresis; and auto sampler on whic
the above sample plate and buffer tank are placed; a plurality of
capillaries which are each filled with a gel and which are each
inserted into each of the above wells and are brought into contact
with the sample thereby to absorb the sample and are also brought
into contact with the buffer solution thereby to form an
electrophoretic lane; a plurality of electrode members each
installed in the vicinity of the above plurality of capillaries; an
electroconductive member which comes into electrical contact with
the above plurality of capillaries; a capillary array holder for
integrally holding the above plurality of capillaries, the above
plurality of electrode members and the above electroconductive
member; a power source for applying a voltage across the loading
end and trailing end of the above capillary; and a controller for
controlling the above auto sampler and the above power source.
[0021] The capillary electrophoretic instrument of the above
construction is briefly explained.
[0022] In order to reduce burdens on analysts, the construction of
the capillary electrophoretic instrument permits the use of a
commercially available microtiter-plate and the sample plate and
buffer container are not given an electrode structure.
[0023] The electrode portion for applying a voltage to the
capillary is integrally formed with the capillary array so that the
electrode portion can be attached to the capillary electrophoretic
instrument and detached therefrom. This eliminates troublesomeness
during capillary setting.
[0024] As the above electrode portion, there are disposed metal
wires in the vicinity of the capillaries at minute intervals, for
example, at distances of about 1 mm, parallel to the relevant
capillaries.
[0025] Further, the above capillaries are inserted into the
cylindrical electrodes having almost the same inner diameter as the
outer shape of the capillaries, and the above electrodes and
capillaries are bonded to each other so that a gap through which
other samples mix in is not formed.
[0026] Further, in the above capillaries, the electrode portions
are formed by vapor-depositing a thin metal film or an
electroconductive material on outer walls of the capillaries.
[0027] The above means permits the use of a commercially available,
inexpensive microtiter-plate, etc. as the sample plate, enables a
capillary array of capillaries, which have a long life and must be
replaced according to the nature of an analysis, to be
simultaneously set, eliminates troublesomeness, and permits stable
DNA analyses by preventing the mixing of other samples.
BRIEF DESCRIPTION OF THE DRAWINGS
[0028] FIG. 1 is an explanatory diagram of a capillary
electrophoretic instrument for a DNA sequencer.
[0029] FIG. 2 is an explanatory diagram of an electrode portion in
the electrophoretic instrument shown in FIG. 1.
[0030] FIG. 3 is an explanatory diagrams of the connection of a
power source in the electrophoretic instrument shown in FIG. 2.
[0031] FIG. 4 is an explanatory diagrams of another example of
electrode portion in the electrophoretic instrument shown in FIG.
1.
[0032] FIG. 5 is an explanatory diagrams of a further example of
electrode portion in the electrophoretic instrument shown in FIG.
1.
[0033] FIG. 6 is an explanatory diagrams of an even further example
of electrode portion in the electrophoretic instrument shown in
FIG. 1.
[0034] FIG. 7 is an explanatory diagram of a capillary array.
DETAILED DESCRPTION
A FIRST EMBODIMENT
[0035] Embodiments of the capillary electrophoretic instrument
related to a first aspect of the present invention and the
capillary related to a second aspect of the present invention are
explained below by referring to FIGS. 1 to 7.
[0036] FIG. 1 is an explanatory diagram of a capillary
electrophoretic instrument for DNA sequencer. FIG. 2 is an
explanatory diagram of an electrode portion in the electrophoretic
instrument shown in FIG. 1. FIGS. 3A and 3B are explanatory
diagrams of the connection of a power source in the electrophoretic
instrument shown in FIG. 2. FIGS. 3A and 3B are explanatory
diagrams of another example of electrode portion in the
electrophoretic instrument shown in FIG. 1. FIG. 5 is an
explanatory diagram of a further example of electrode portion in
the electrophoretic instrument shown in FIG. 1. FIG. 6 is an
explanatory diagram of an even further example of electrode portion
in the electrophoretic instrument shown in FIG. 1.
[0037] First, the general construction of a capillary
electrophoretic instrument best suited to a DNA sequencer related
to the first aspect of the present invention is explained.
[0038] In FIG. 1, the numeral 1 indicates a capillary
electrophoretic instrument for a DNA sequencer (hereinafter
referred to as an electrophoretic instrument), the numeral 2 a
capillary array, the numeral 2a a capillary, the numeral 3 a
constant-temperature air bath, the numeral 5 a sample plate, the
numeral 6 a buffer tank, the numeral 7 an auto sampler capable of
freely moving in each direction of the X, Y and Z axes, the numeral
8 an array holder for fixing the capillary array (hereinafter
referred to as an array holder), the numeral 9 a gel block, the
numeral 10 an electrophoretic ground, the numeral 12 a gel-filling
syringe, the numeral 13 a light irradiation-analysis portion, the
numeral 40 a power source, and the numeral 50 a control
computer.
[0039] In FIG. 1, the capillary array comprises at least one
capillary 2a and is fixed by the array holder 8 to form an
electrophoretic portion. The above capillary array 2 is so set that
the greater part thereof is housed in the constant-temperature air
bath 3. The outer region of the above constant-temperature air bath
3 is covered with a heat-insulating material with the exception of
a part thereof, and the above part of the outer region which is not
covered with the above heat-insulating material is provided with
either a heating element or a heating/cooling element that is
brought into contact with the air in the constant-temperature air
bath.
[0040] Near the loading end of the above capillary array 2 is
disposed the sample plate 5 on which a sample is set. Further, in
the vicinity of the sample plate 5 is disposed the buffer tank 6
that houses a buffer solution for preventing an electric discharge
during an application of a voltage to the capillary array 2 and for
effecting the electrophoresis of the sample. A commercially
available microtiter-plate is used as the above sample plate 5.
Hereinafter, the sample plate 5 refers to a microtiter-plate. The
above sample plate 5 is made of a synthetic resin such as acrylic
resin and has a rectangular plane shape, and 8.times.12=96 wells
are installed in rows and columns on the sample plate 5.
[0041] Each of the above wells 21 has such a cross-sectional shape
that a tapered portion is formed from the top surface side of the
sample plate 5 toward the bottom of the well. This permits easy
operations during the injection and introduction of the sample.
[0042] The above sample plate 5 and buffer tank 6 are placed on the
auto sampler 7 that is capable of moving in each direction of X, Y
and Z axes. The auto sampler 7 is attached to the bottom surface of
a casing of electrophoretic instrument 1. The control computer 50
positions the auto sampler 7 in the fore-and-aft, horizontal and
vertical directions and controls a movement motor (not shown in the
figure) to move the auto sampler 7.
[0043] In the above capillary array 2, 8.times.2=16 capillaries 2a
are arranged in rows and columns and the interior of the
capillaries 2a is filled with a gel for electrophoretic separation.
The loading end side of the above capillary array 2 corresponding
to the above sample plate 5 is fixed by the array holder 8, and the
other loading end side of the above capillary array 2 is fixed by
being connected to the gel block 9.
[0044] The above gel block 9 in the section from the connection to
the capillary array 2 to the electrophoretic ground 10 is filled
with a gel (polymer), which is a separation medium, and the power
source 40 is connected to the grounding-electrode side of the
electrophoretic ground 10. The above electrophoretic ground 10 is
connected to the bottom surface of the casing of the
electrophoretic instrument 1.
[0045] At a pitch corresponding to the pitch of the wells 21 of
sample plate 5, the above capillaries 2a are arranged and attached
to the array holder 8. In the example shown in the figure, eight
capillaries are used. In the array holder 8 to which the above
arrays 2a are attached, the relevant capillaries and the electrode
portion (not shown in the figure), which comprises metal-wire
electrodes and a common electrode, are integrally constructed. This
integrally-constructed electrode portion will be described later.
To this electrode portion is connected the high-voltage side of the
power source 40.
[0046] Near the trailing-end portion of the above capillary array 2
is disposed the light analysis portion 13. This light analysis
portion 13 is constituted by, for example, a laser light source
(not shown in the figure) for irradiating and exciting a sample in
the capillary array 2 and a photo sensor for detecting the light
emitted by the above excitation. From signals of this photo sensor,
the base sequence of DNA is determined and the type of DNA is
identified by the control computer 50.
[0047] Incidentally, the electrophoretic instrument 1 is provided
with the gel-filling syringe 12 for replacing the gel each time one
electrophoretic operation is performed and a solenoid valve 11 for
preventing a backflow during the above gel replacement. The
operations of these members are also controlled by the control
computer 50.
[0048] In FIG. 1, the power source 40 and control computer 50 are
installed outside the electrophoretic instrument 1. However, it is
needless to say that they may be installed within the
electrophoretic instrument 1.
[0049] By referring to FIG. 2, the integrally-constructed electrode
portion comprising capillaries, metal-wire electrodes and a common
electrode in the array holder 8 in the electrophoretic instrument
of the above construction is explained below.
[0050] In FIG. 2, an integrally-constructed electrode portion is
separated into an insulating member 25a and an insulating member
25b that constitute the array holder 8, in order to make the
construction clear. The construction of the integrally-constructed
electrode portion is such that the insulating member 25a and
insulating member 25b that constitute the array holder 8 are in
mutual engagement when integrally constructed. The insulating
member 25a and insulating member 25b are arranged so that they
correspond to the well 21 of a sample plate.
[0051] In FIG. 2, the same numerals as those used in FIG. 1 are
omitted to avoid troublesomeness because they indicate the same
functions and same portions as in FIG. 1, and only new numerals are
explained. The numeral 23 indicates a metal-wire electrode, the
numeral 24 a common electrode portion, the numerals 25a and 25b
insulating members, the numeral 25c a round through hole through
which the capillary 2a passes, and the numeral 26 an insertion hole
of an electrode rod 30 to be connected to the power source 40.
[0052] As shown in FIG. 2A, the array holder 8 has a face
corresponding to the sample plate 5 and comprises the above
insulating material 25b, which is formed in a manner that an
L-shaped portion is laterally placed on the above corresponding
face, and the above insulating material 25a, which is in engagement
with the corresponding face and L-shaped portion of the above
insulating material 25b, so that the two insulating members are
mutually integrated in construction. Incidentally, the array holder
8 is attached by appropriate means on the wall surface of the
constant-temperature air bath 3 shown in the figure. The same
applies to each example of variation shown below.
[0053] The above insulating material 25b is provided with
capillaries 2a, which are disposed through the corresponding
surface to the sample plate 5 at a pitch in accordance with the
pitch of the wells 21 of the sample plate 5. A plurality of
capillaries 2a (for example, two capillaries are schematically
shown), a plurality of metal-wire electrodes 23, which are disposed
in the vicinity of the above plurality of capillaries 2a each
parallel with the relevant capillaries 2a and pass through the
above corresponding surface to the sample plate 5, and the common
electrode portion 24, which is fixed on a rising surface of the
above L-shaped portion intersecting at right angles with the above
corresponding surface to the sample plate 5. The expression "the
vicinity of the above plurality of capillaries 2a" refers to such
positions that the above plurality of capillaries 2a are
simultaneously inserted into the above wells 21. The above
plurality of metal-wire electrodes 23 are all electrically
connected to the above common electrode portion 24. This electrode
member may contact the capillaries.
[0054] In order to ensure that, when the insulating member 25a and
the insulating member 25b are engaged with each other, the above
plurality of capillaries 2a can be inserted, an exact number of
through holes through which the above plurality of capillaries 2a
pass, the number of which is equal to that of capillaries 2a, are
drilled in the insulating member 25a in accordance with the pitch
of the plurality of capillaries 2a.
[0055] When the insulating member 25a and the insulating member 25b
are engaged with each other thereby to form a mutually integrated
construction, the outer surfaces of the capillaries 2a, metal-wire
electrodes 23 and common electrode portion 24 are covered with the
insulating member 25a and the insulating member 25b which are in
mutual engagement and, therefore, from the direction of the above
sample plate 5, only the above capillaries 2a and the above
metal-wire electrodes are seen. Incidentally, when the integrated
construction is obtained, engaging faces are bonded by appropriate
means, for example, with the use of a bonding agent.
[0056] Further, when the above integrated construction is obtained,
from the above through holes 25c of above insulating member 25a a
plurality of capillaries 2a protrude outwardly and extend to form
the trailing-end portion of the electrophoretic portion. Therefore,
from the opposite side of the above sample plate 5 only the
capillaries 2a are seen. Covering the outer region with the
insulating members 25a and 25b in this manner prevents the
occurrence of an arc discharge due to a high voltage applied to the
capillaries 2a.
[0057] By referring to FIGS. 3A and 3B, the voltage application
structure of the power source in the integrated structure of array
holder is explained below. FIG. 3A is a rear view of the electrode
portion of integrated construction shown in FIG. 2. FIG. 3B is a
plan view of the electrode portion of integrated construction shown
in FIG. 2. In FIGS. 3A and 3B, the numeral 30 indicates an
electrode portion (hereinafter referred to as an electrode rod) to
be connected to the power source 40. Incidentally, in FIG. 3A, the
illustration of the electrode rod 30 is omitted in order to make
the integrally-constructed portion clear.
[0058] The common electrode portion 24 installed on a plane
intersecting at right angles with the sample plate 5 on the above
insulating member 25b has at least one insertion hole 26. The
electrode rod 30 is inserted into the above insertion hole 26 and
abuts against the above common electrode 24. On this occasion, the
electrode rod 30 is urged by an elastic member (omitted in the
drawing of FIGS. 3A and 3B) in order to lower electrical contact
resistance against the above common electrode 24. The negative
high-voltage side of the power source 40 is applied to the other
end of the abutting surface of the above electrode rod 30 against
the above common electrode portion 24, and the positive side of the
power source 40 is connected to the grounding side of the
electrophoretic ground 10.
[0059] When the above capillary array 2 is inserted into the wells
21 and the high-voltage side is applied to the common electrode
portion 24 via the electrode rod 30 inserted into the above
insertion hole 26, the above metal-wire electrodes 23 are in
electrical contact with the above common electrode portion 24 and,
therefore, the above high-voltage side is applied. Accordingly, the
above metal-wire electrodes 23 and capillaries 2a simultaneously
come into contact with the sample 21 within the wells 21 of each
sample plate or the buffer solution in the buffer tank, with the
result that sample loading into the above capillaries 2a or
electrophoresis is effected.
[0060] By referring to FIG. 4, an example of variation of the
integrally-constructed electrode portion of the array holder shown
in FIG. 2 is explained below. In FIG. 4, the same numerals as those
used in FIG. 2 are omitted to avoid the troublesomeness of
re-explanation because they indicate equivalents of the same
functions and same specifications as in FIG. 2, and only new
numerals are explained. The numeral 24a indicates an extension
electrode portion of the common electrode portion 24, and the
numeral 27 an electrical contact portion.
[0061] In FIG. 4, the array holder 8 is separated into an
insulating member 25b and an insulating member 25a that is in
engagement with the insulating member 25b, in order to make the
integrated construction clear.
[0062] The construction of the integrally-constructed electrode
portion is such that the insulating member 25b and insulating
member 25a are in mutual engagement when integrally constructed.
The capillary array is arranged so that it corresponds to the well
21 of the sample plate 21.
[0063] As shown in the drawings, the common electrode portion 24 is
disposed on a plane within the insulating member 25b corresponding
to the sample plate 5. The metal-wire electrode 23 inserted into
the insulating member 25b is joined to the above common electrode
portion 24 through an electrical contact portion 27 by means of
appropriate means such as welding and the like. In the vicinity of
a through hole 26, through which the above electrode rod 30 passes,
the extension electrode portion 24a of the above common electrode
portion 24 is installed. Thus, the extension electrode portion 24a
can provide a high-voltage application portion to the above
capillary array 2 and metal-wire electrode 23.
[0064] By referring to FIG. 5, another example of variation of the
integrally-constructed electrode portion of the array holder shown
in FIG. 2 is explained below. In FIG. 5, the same numerals as those
used in FIG. 2 are omitted to avoid the troublesomeness of
re-explanation because they indicate equivalents of the same
functions and same specifications as in FIG. 2, and only new
numerals are explained. The numeral 28 indicates a vapor-deposited
electrode that is obtained by vapor-deposition on the capillary
2a.
[0065] In FIG. 5, the array holder 8 is separated into an
insulating member 25b and an insulating member 25a that is in
engagement with the insulating member 25b, in order to make the
integrated construction clear. The capillary array and sample plate
5 are arranged so that both correspond to each other. The
construction of the integrally-constructed electrode portion is
such that the insulating member 25b and insulating member 25a of
the array holder 8 are in mutual engagement when integrally
constructed.
[0066] The difference between this example of variation and the
example shown in FIG. 2 is that the vapor-deposited electrode 28
made of platinum is used as each electrode in place of the
metal-wire electrode. The above vapor-deposited electrode 28 is
provided with a vapor-deposited portion from the end of the metal
tube on the sample side to the portion corresponding to the common
electrode portion 24. This vapor-deposited electrode 28 comes into
electrical contact with the common electrode portion 24 in the
above vapor-deposited portion.
[0067] Further, in this variation, the through hole in FIG. 2 for
engaging the insulating member 25a with the insulating member 25b
and causing the capillary 2a to protrude outwardly from the array
holder 8 is semicircular 25d in shape.
[0068] By referring to FIG. 6, a further example of variation of
the integrally-constructed electrode portion of the array holder
shown in FIG. 4 is explained below. In FIG. 6, the same numerals as
those used in FIG. 4 are omitted to avoid the troublesomeness of
re-explanation because they indicate equivalents of the same
functions and same specifications as in FIG. 4, and only new
numerals are explained. The numeral 29 indicates a cylindrical
electrode made of stainless steel pipe.
[0069] In FIG. 6, the array holder 8 is separated into an
insulating member 25b and an insulating member 25a that is in
engagement with the insulating member 25b, in order to make the
integrated construction clear. The capillary array is arranged so
that it corresponds to the sample plate 5. The construction of the
integrally-constructed electrode portion is such that the
insulating member 25b and insulating member 25a of the array holder
8 are in mutual engagement when integrally constructed. The
difference between this example of variation and the example shown
in FIG. 4 is that as the electrode for each capillary 2a, the
cylindrical electrode 29 into which a capillary is inserted is used
in place of the metal-wire electrode.
[0070] The above cylindrical electrode 29 is constructed in such a
manner that a cylindrical tube surrounding each capillary 2a is
installed from the leading end of the relevant capillary 2a on the
sample side to the common electrode portion 24 and an electrical
contact portion 27 is provided between the common electrode portion
24 and the relevant cylindrical electrode 29.
[0071] In this example of variation, the gap between the
cylindrical electrode 29 and the capillary 2a is filled in, for
example, by bonding so that other samples do not remain in this
gap.
[0072] The shape of the insulating members 25a and 25b used in each
of the above embodiments is not limited to those described above
and the insulating members 25a and 25b may have various other
shapes.
A SECOND EMBODIMENT
[0073] Next, the general construction of a capillary array related
to the second aspect of the present invention is explained.
[0074] By referring to FIG. 7, the construction of a capillary
array in which the array holder shown in FIG. 6 is used is
explained below. FIG. 7 is an explanatory diagram of a capillary
array related to the second aspect of the present invention.
[0075] As explained in connection with FIG. 6, in the above array
holder 8, the cylindrical electrode 29 surrounding each capillary
2a, which is provided from the loading end of the relevant
capillary 2a on the sample side, comes into contact, in the
electrical contact portion 27, with the common electrode portion 24
installed on a corresponding surface to the sample plate 5 (the
common electrode portion 24 is not seen from the outside in FIG.
7). As shown in the drawing of FIG. 6, a capillary array 2 as
described above and 16 cylindrical electrodes 29 are arranged in
accordance with the pitch of the wells 21 of sample plate 5 and the
cylindrical electrodes 29 are caused to pass through the insulating
member 25 of array holder 8 so that they are arranged in 8 rows and
2 columns.
[0076] A re-explanation is omitted to avoid troublesomeness. As
explained in connection with FIG. 6, the insulating member 25
comprises the two insulating members 25b and 25a which are in
mutual engagement (in FIG. 7, no distinction is made between the
two because they are engaged with each other).
[0077] Further, the construction of the capillary array 2 is such
that by means of the electrode rods (refer to FIG. 3B), which pass
through the through holes 26 made in a part or a plurality of
places of the insulating member 25 covering the common electrode
24, etc., a high voltage can be applied to the electrophoretic
portion from outside the capillary array 2 via the above common
electrode portion 24.
[0078] Sample loading or electrophoresis is effected when the
loading end of each capillary 2a that has gone through and the
cylindrical electrode 29 provided in the loading end portion are
inserted into each well 21 of sample plate 5 and the buffer tank 6
(neither of the two is shown in FIG. 7). The trailing-end portion
of each capillary 2a is provided with a light analysis portion 13
and is connected to a gel block 9.
[0079] The above unit of capillary array 2 shown in FIG. 7, i.e.,
the unit of capillary array 2 comprising 16 capillaries 2a enables
the whole including not only the capillaries 2a, but also the
cylindrical electrodes 29 and the common electrode 24 to be
simultaneously attached to the electrophoretic instrument and
detached therefrom, thereby facilitating insertion into the wells.
For this reason, the replacement of the capillary array is easy and
also because the electrodes and capillaries are integrally
constructed by one array holder, it is unnecessary to pay attention
to the accuracy of arrangement between the electrodes and the
capillaries. In the above description, the capillary array in which
the array holder shown in FIG. 6 was explained. However, it is
needless to say that the above description applies also to other
array holders.
[0080] Next, the operation of the electrophoretic instrument of the
above construction is explained with the aid of the capillary array
shown in FIG. 7.
[0081] As the preparations for the operation of the electrophoretic
instrument, by means of a pipette a sample is injected into each of
the 8.times.5=96 wells 5a of the sample plate. The lid of the
constant-temperature air bath is closed and circulating air streams
are formed by fans 127 and 128 installed in the chamber of the
above constant-temperature air bath 3. The outer surface of the
constant-temperature air bath 3 is covered with a heat-insulating
material 126 with the exception of a part thereof, and the above
part of the outer surface which is not covered with the above
heat-insulating material is provided with elements 22 capable of
heating and cooling. Further, the inner surface of the
constant-temperature air bath 3 is covered with a member of good
thermal conductivity 123. Therefore, the heat transfer from the
elements 22 capable of heating and cooling occurs rapidly on the
inner surface of the chamber.
[0082] Further, the control computer 50 controls and starts the
above fans 127 and 128. Fans that suck air from the direction of
rotation and blows out the air in the radial direction are used as
the above fans 127 and 128. Therefore, circulating air streams of
large air volume are obtained and the thickness of the
constant-temperature air bath 20 becomes small, with the result
that the above heat-insulating material 26 and elements capable of
heating and cooling on the outer surface and the member of good
thermal conductivity 123 on the inner surface, in conjunction with
each other, make uniform the temperature within the chamber of
constant-temperature air bath 20. The temperature of the whole
capillary array of electrophoretic instrument 1 is made constant
and uniform.
[0083] After this condition is obtained, the control computer 50
causes the auto sampler 7 to move a microtiter-plate back and
forth, and when each well 21 of the microtiter-plate has come under
each capillary 2a of capillary array 2, the auto sampler 7 stops.
Next, the control computer 50 causes the auto sampler 7 to ascend
and stops it in a position where the capillary 2a is inserted into
the sample in the well 21.
[0084] Next, the capillary array 2 is inserted into the sample in
the well 21. On this occasion, because the above 16 capillaries 2a,
16 electrodes 29, and the common electrode 24 that is in contact
with these electrodes 29 are covered with the insulating members
25a and 25b to form an integrally-constructed electrode portion,
each of these members can be easily and simultaneously inserted
into the sample in each corresponding well 21.
[0085] Incidentally, in a case where metal-wire electrode 23 as
shown in FIG. 2 is used, the metal-wire electrode 23 must not be
brought into contact with the capillary 2a when the capillary array
2 is not inserted into the well 21 of sample plate 5. This is
because if the metal-wire electrode 23 is brought into contact with
the capillary 2a, other samples would remain in the gap between the
two due to the capillary phenomenon, preventing good-accuracy
analyses.
[0086] With the capillary array 2 kept inserted in the well 21 of
sample plate 5, the control computer 50 controls the power source
40 so that via the electrode rod 30 inserted into the through hole
26 (refer to FIGS. 3A and 3B), a negative high voltage is applied
to a circuit formed by the electrophoretic ground 10, gel block 9,
gel in the capillary 2a, sample, and electrode 29 in this order,
with the result that the sample in the well 5a is introduced into
the capillary 2a. At this point in time, the negative high voltage
is interrupted.
[0087] The auto sampler 7 is again moved and is stopped in a
position where the bottom end of the capillary 2a is inserted into
the buffer tank 6. Next, when the above auto sampler 7 is
vertically moved, the above capillary 2a is inserted into the
buffer solution in the above buffer tank 6 and the above electrode
29 is also inserted into the buffer solution. As with the insertion
of each member into the sample, the insertion of each member into
the buffer solution also takes place simultaneously and easily.
[0088] With this condition maintained, a negative high voltage is
again applied to the circuit of electrophoretic ground 10-gel block
9-gel in the capillary 2a-sample-electrode 29. The application of
this high voltage causes the sample loaded in the capillary 2a to
be electrophoretically separated.
[0089] Incidentally, the gel polymer within the capillary array 2
is replaced with a new gel polymer each time a measurement is
carried out. This is performed by closing a solenoid valve 11,
thereby driving a gel-filling syringe 12 and filling the capillary
array 2 with the gel polymer within the syringe 12. These
operations are controlled by the control computer 50.
[0090] The light analysis portion 13 is shielded against light from
the outside, and laser light for sample excitation (not shown in
the figure) is loaded to the capillary 2a in the position of the
light analysis portion 13. Fluorescence emitted from a fluorescent
reagent which has been bonded to DNA migrating electrically in the
capillary is detected and a DNA analysis is performed by the
computer on the basis of this signal.
[0091] According to the essential features of the invention, by
providing a plurality of capillaries and electrodes to form pairs,
the capillary array can be easily aligned with the sample bed.
* * * * *