U.S. patent application number 10/811917 was filed with the patent office on 2004-11-11 for biochemical reaction cartridge.
This patent application is currently assigned to CANON KABUSHIKI KAISHA. Invention is credited to Numajiri, Yasuyuki.
Application Number | 20040223874 10/811917 |
Document ID | / |
Family ID | 32993078 |
Filed Date | 2004-11-11 |
United States Patent
Application |
20040223874 |
Kind Code |
A1 |
Numajiri, Yasuyuki |
November 11, 2004 |
Biochemical reaction cartridge
Abstract
A biochemical reaction cartridge includes an injection port for
injecting a specimen, a chamber for containing therein the
specimen, a chamber for containing a regand for treating the
specimen, nozzle ports for applying or reducing pressure by using
fluid. In the cartridge, the specimen is subjected to a sequence of
a biochemical reaction by controlling the fluid. The cartridge is
mounted in a biochemical reaction apparatus.
Inventors: |
Numajiri, Yasuyuki;
(Kawasaki-shi, JP) |
Correspondence
Address: |
FITZPATRICK CELLA HARPER & SCINTO
30 ROCKEFELLER PLAZA
NEW YORK
NY
10112
US
|
Assignee: |
CANON KABUSHIKI KAISHA
TOKYO
JP
|
Family ID: |
32993078 |
Appl. No.: |
10/811917 |
Filed: |
March 30, 2004 |
Current U.S.
Class: |
422/400 |
Current CPC
Class: |
B01L 3/502761 20130101;
B01L 2300/087 20130101; B01L 7/52 20130101; B01L 2200/027 20130101;
B01L 3/502715 20130101; B01L 2400/0487 20130101; B01L 2300/0816
20130101; B01L 2300/1822 20130101 |
Class at
Publication: |
422/058 ;
422/057 |
International
Class: |
G01N 033/00 |
Foreign Application Data
Date |
Code |
Application Number |
Mar 31, 2003 |
JP |
094241/2003(PAT.) |
Mar 31, 2003 |
JP |
097136/2003(PAT.) |
Claims
What is claimed is:
1. A biochemical reaction cartridge, comprising: an injection port
for injecting a specimen therefrom, a first chamber for containing
the specimen therein, a second chamber for containing therein a
reagent which contributes to a biochemical reaction, a passage for
passing therethrough the specimen and/or the reagent and/or a
reaction liquid, and a plurality of nozzle ports for receiving a
plurality of nozzles for applying or reducing pressure, wherein
said plurality of nozzle ports communicate with said first or
second chamber, and fluid is present between said plurality of
nozzle ports and said first or second chamber and is pressurized or
depressurized by said plurality of nozzles to move the specimen
and/or the reagent and/or the reaction liquid, thereby to effect a
sequence of a biochemical reaction within the cartridge.
2. A cartridge according to claim 1, wherein said plurality of
nozzle ports are divided into two portions, which are disposed on
two surfaces of the cartridge.
3. A cartridge according to claim 1 or 2, wherein said plurality of
nozzle ports are disposed linearly.
4. A cartridge according to claim 2, wherein the cartridge is a
substantially rectangular parallelepiped, and the two surfaces are
lateral surfaces, opposite from each other, of the
parallelepiped.
5. A cartridge according to claim 1, wherein said particles of a
magnetic material to which a target material comprising DNA, RNA or
protein is adsorbed, are used as a species of the reagent, and are
trapped during movement thereof by exerting a magnetic force of a
magnet disposed close to said passage, after an an adsorption
reaction is completed, thereby to purify the target material.
6. A cartridge according to claim 1, wherein the cartridge further
comprises a chamber containing a washing liquid and a chamber
containing waste liquid after washing.
7. A biochemical treatment apparatus, comprising: a cartridge
mounting portion for mounting a cartridge having a plurality of
chambers containing a solution for biochemically treating a
specimen, a plurality of nozzle portions each connected to an
associated passage communicating with an associated chamber of the
chambers of the cartridge, and control means for controlling a
fluid pressure in the cartridge through said nozzle portions,
wherein said control means controls the fluid pressure so that the
solution in the cartridge is moved only in the cartridge.
8. An apparatus according to claim 7, wherein a plurality of
cartridges are mountable to the apparatus.
9. An apparatus according to claim 7, wherein said plurality of
nozzle portions are separately disposed at two surfaces of the
cartridge.
10. An apparatus according to claim 7, wherein said plurality of
nozzle portions are arranged linearly.
11. A biochemical treatment process for effecting biochemical
treatment in a cartridge having a plurality of chambers containing
a solution for biochemically treating a specimen, said process
comprising: a step of connecting each of nozzles to an associated
port of passage communicating with an associated chamber of the
cartridge, and a step of injecting fluid into the cartridge to move
the liquid in the cartridge.
12. A process according to claim 11, wherein said injection step
comprises a step of injecting a hymolytic agent.
13. A process according to claim 11, wherein said injection step
comprises a step of injecting particles of a magnetic material to
which a target material comprising DNA, RNA or protein is
adsorbed.
14. A process according to claim 13, wherein said process further
comprises, after the step of injecting particles of magnetic
material, a step of trapping the particles of magnetic material
during movement thereof by exerting a magnetic force of a magnet
disposed close to the passage to purify the target material.
15. A process according to claim 14, wherein said process further
comprises, after the trapping step, a step of cleaning the target
material.
16. A biochemical reaction cartridge, comprising: a storage chamber
for accumulating a liquid, a first chamber, a first passage for
connecting said storage chamber to said first chamber to move the
liquid in said storage chamber to said first chamber, a second
chamber, and a second passage for connecting said first chamber to
said second chamber to move the liquid in said first chamber to
said second chamber, wherein a bottom position of a first
connecting portion for connecting said first chamber to said first
passage is higher than a bottom position of a second connecting
portion for connecting said first chamber to said second
passage.
17. A cartridge according to claim 16, wherein the liquid is caused
to flow to said first chamber so that said first chamber has a
maximum liquid level lower than the bottom position of the first
connecting position.
18. A cartridge according to claim 16 or 17, wherein movement of
the liquid is controlled by externally applying or reducing
pressure.
19. A cartridge according to claim 18, wherein said cartridge
comprises a pressure reducing portion for externally reducing
pressure, said pressure reducing portion being provided with a
chamber for preventing outflow of the liquid.
Description
FIELD OF THE INVENTION AND RELATED ART
[0001] The present invention relates to a technology to analyze
cell, microorganism, chromosome, nuclei acid, etc., in a specimen
by utilizing a biochemical reaction. More specifically, the present
invention relates to a biochemical reaction cartridge for use in
the analysis and a biochemical treatment apparatus for effecting
the biochemical reaction in the cartridge.
[0002] Most of analyzers for analyzing specimens such as blood uses
an immunological procedure utilizing antigen-antibody reaction or a
procedure utilizing nuclei acid hybridization. For example, protein
or single-stranded nucleic acid, such as antibody or antigen, which
specifically connects with a material or substance to be detected,
is used as a probe and is fixed on a surface of solid phase, such
as fine particles, beads or a glass plate, thus effecting
antigen-antibody reaction or nuclei acid hybridization. Then, for
example, an antigen-antibody compound or double-stranded nucleic
acid is detected by a labeled antigen or labeled nucleic acid,
which causes a specific interaction such that a labeled material
having a high detection sensitivity, such as an enzyme, a
fluorescent material or a luminescent material, is supported, thus
effecting detection of presence or absence of the material to be
detected or quantitative determination the detected material.
[0003] As an extension of these technologies, e.g., U.S. Pat. No.
5,445,934 has disclosed a so-called DNA (deoxyribonucleic acid)
array wherein a large number of DNA probes having mutually
different base sequences are arranged on a substrate in array
form.
[0004] Further, Anal. Biochem., 270(1), pp. 103-111 (1999) has
disclosed a process for preparing a protein-array, like the DNA
array, such that various species of proteins are arranged on a
membrane filter. By using these DNA and protein arrays and the
like, it has become possible to effect a test on a large number of
items at the same time.
[0005] Further, in various methods of specimen analysis, in order
to realize alleviation of contamination by specimen, promotion of
reaction efficiency, reduction in apparatus size, and facilitation
of operation, there have been also proposed disposable biochemical
reaction cartridges in which a necessary reaction is performed in
the cartridge. For example, Japanese Laid-Open Patent Application
(JP-A) (Tokuhyo) Hei 11-509094 has disclosed a biochemical reaction
cartridge, including DNA array, in which a plurality of chambers
are disposed and a solution is moved by a differential pressure so
as to permit a reaction such as extraction, amplification or
hybridization of DNA in a specimen within the cartridge. U.S. Pat.
No. 5,690,763 has disclosed a constitution for reacting a
three-dimensionally curved passage through sheet lamination, and
U.S. Pat. Nos. 6,167,910 and 6,494,230 have disclosed structures of
.mu.-TAS (micro-total analysis system) wherein a passage is
provided between a first layer and a second layer and between a
second layer and a third layer, constituting a three-layer
structure, and the respective passages are partially connected with
each other.
[0006] As a method for externally injecting a solution into the
inside of such biochemical reaction cartridges, it is possible to
utilize an external syringe or vacuum pump. Further, a a method for
moving the solution within the biochemical reaction cartridges,
those utilizing gravity, capillarity, and electrophoresis are
known. Further, as a compact micropump which can be provided inside
of the biochemical reaction cartridge, Japanese Patent No. 2832117
has disclosed one utilizing a heat generating element, JP-A
(Tokkai) 2000-274375 has disclosed one utilizing a piezoelectric
element, and JP-A (Tokuhyo) Hei 11-5-9094 has disclosed a diaphragm
pump.
[0007] As described above, it is preferable that a disposable
cartridge containing a necessary solution is used from the
viewpoints of prevention of secondary infection or contamination
and usability but the cartridge containing a pump is expensive.
[0008] Further, in the conventional biochemical reaction
cartridges, such as .mu.-TAS, there is no disclosure as to how to
use properly a manner of movement of liquid performed by only
injecting, e.g., a regand, liquid or a specimen in one direction
and a manner of movement of reaction liquid required for
reciprocating motion. Particularly, the former movement is
accompanied with such a problem that when the whole quantity of
liquid is moved, bubbles are generated after completion of the
movement, and thus the whole quantity of liquid cannot be moved
completely in the case of preventing the generation of bubbles.
SUMMARY OF THE INVENTION
[0009] An object of the present invention is to provide a
disposable biochemical reaction cartridge having a structure
capable of causing a sequence of a biochemical reaction to proceed
by moving a solution under the action of an external pump without
containing a pump and capable of preventing the solution from
flowing out of the cartridge.
[0010] Another object of the present invention is to provide a
biochemical treatment apparatus for effecting the biochemical
reaction within the cartridge by using the biochemical reaction
cartridge described above.
[0011] Another object of the present invention is to provide a
method o fusing a biochemical reaction cartridge capable of
ensuring appropriate movement in such a manner that in a
biochemical reaction cartridge for effecting movement of liquid
therein, an optimum passage is selected and used properly with
respect to movement of a reagent or a specimen only requiring
injection into a subsequent chamber and movement of a reaction
liquid requiring reciprocating motion.
[0012] According to the present invention, there is provided a
biochemical reaction cartridge, comprising:
[0013] an injection port for injecting a specimen therefrom,
[0014] a first chamber for containing the specimen therein,
[0015] a second chamber for containing therein a reagent which
contributes to a biochemical reaction,
[0016] a passage for passing therethrough the specimen and/or the
reagent and/or a reaction liquid, and
[0017] a plurality of nozzle ports for receiving therethrough a
plurality of nozzles for applying or reducing pressure,
[0018] wherein the plurality of nozzle ports communicate with the
first or second chamber, and fluid is present between the plurality
of nozzle ports and the first or second chamber and is pressurized
or depressurized by the plurality of nozzles to move the specimen
and/or the reagent and/or the reaction liquid, thereby to effect a
sequence of a biochemical reaction within the cartridge.
[0019] According to the present invention, there is also provided a
biochemical treatment apparatus, comprising:
[0020] a cartridge mounting portion for mounting a cartridge having
a plurality of chambers containing a solution for biochemically
treating a specimen,
[0021] a plurality of nozzle portions each connected to an
associated passage communicating with an associated chamber of the
chambers of the cartridge, and
[0022] control means for controlling a fluid pressure in the
cartridge through the nozzle portions,
[0023] wherein the control means controls the fluid pressure so
that the solution in the cartridge is moved only in the
cartridge.
[0024] According to the present invention, there is further
provided a biochemical treatment process for effecting biochemical
treatment in a cartridge having a plurality of chambers containing
a solution for biochemically treating a specimen, the process
comprising:
[0025] a step of connecting each of nozzles to an associated port
of passage communicating with an associated chamber of the
cartridge, and
[0026] a step of injecting fluid into the cartridge to move the
liquid in the cartridge.
[0027] According to the present invention, there is still further
provided a biochemical reaction cartridge, comprising:
[0028] a storage chamber for accumulating a liquid,
[0029] a first chamber,
[0030] a first passage for connecting the storage chamber to the
first chamber to move the liquid in the storage chamber to the
first chamber,
[0031] a second chamber, and
[0032] a second passage for connecting the first chamber to the
second chamber to move the liquid in the first chamber to the
second chamber,
[0033] wherein a bottom position of a first connecting portion for
connecting the first chamber to the first passage is higher than a
bottom position of a second connecting portion for connecting the
first chamber to the second passage.
[0034] These and other objects, features and advantages of the
present invention will become more apparent upon a consideration of
the following description of the preferred embodiments of the
present invention taken in conjunction with the accompanying
drawings.
BRIEF DESCRIPTION OF THE DRAWINGS
[0035] FIG. 1 is a perspective view of an embodiment of the
biochemical reaction cartridge according to the present
invention.
[0036] FIG. 2 is a plan view of the biochemical reaction
cartridge.
[0037] FIG. 3 is a block diagram of a treatment apparatus for
controlling movement of liquid and various reactions within the
biochemical reaction cartridge.
[0038] FIG. 4 is a flow chart of a treatment procedure.
[0039] FIG. 5 is a longitudinal sectional view of a part of a
chamber.
[0040] FIG. 6 is a longitudinal sectional view of another part of
the chamber.
[0041] FIG. 7 is a longitudinal sectional view of another part of
the chamber.
[0042] FIG. 8 is a longitudinal sectional view of a part of a
chamber according to another embodiment.
DESCRIPTION OF THE PREFERRED EMBODIMENTS
[0043] Hereinbelow, the present invention will be described more
specifically with reference to the drawings.
[0044] (Embodiment 1)
[0045] FIG. 1 is an external view of a biochemical reaction
cartridge 1 in this embodiment. Referring to FIG. 1, on the
cartridge 1, a specimen port 2 for injecting a specimen such as
blood by a syringe (injector) or the like is disposed and sealed up
with a rubber cap. On a side surface of the cartridge 1, a
plurality of nozzle ports 3 into which nozzles are injected to
apply or reduce pressure in order to move a solution in the
cartridge 1. A rubber cap is fixed on each of the nozzle ports 3.
The other side surface of the cartridge 1 has a similar
structure.
[0046] A body of the biochemical reaction cartridge 1 comprises
transparent or semitransparent synthetic resin, such as polymethyl
methacrylate (PMMA), acrylonitrile-butadiene-styrene (ABS)
copolymer, polystyrene, polycarbonate, polyester or polyvinyl
chloride. In the case where an optical measurement is not required,
the material for the body of the cartridge 1 is not required to be
transparent.
[0047] FIG. 2 is a plan view of the biochemical reaction cartridge
1. Referring to FIG. 2, on one side surface of the cartridge 1, 10
nozzle ports 3a to 3j are provided and also on the other side
surface thereof, 10 nozzle ports 3k to 3t are provided. The
respective nozzle ports 3a to 3t communicate with chambers 5, which
are portions or sites for storing the solution or causing a
reaction, through corresponding air passages 4a to 4t,
respectively.
[0048] In this embodiment, however, the nozzle ports 3n, 3p, 3q and
3s are not used, these nozzle ports do not communicate with the
chambers 5 and are used as reserve ports. More specifically, in
this embodiment, the nozzle ports 3a to 3j communicate with the
chambers 5a to 5j through the passages 4a to 4j, respectively. On
the other side surface, the nozzle ports 3k, 3l, 3m, 3o, 3r and 3t
communicate with the chambers 5k, 5l, 5m, 5o, 5r and 5t through the
passages 4k, 4l, 4m, 4o, 4r and 4t, respectively.
[0049] The specimen port 2 communicates with a chamber 7. The
chambers 5a, 5b, 5c and 5k communicate with the chamber 7, the
chambers 5g and 5o communicate with a chamber 8, and the chambers
5h, 5i, 5j, 5r and 5t communicate with a chamber 9. Further, the
chamber 7 communicate with the chamber 8 via a passage 10, and the
chamber 8 communicates with the chamber 9 via a passage 11. With
the passage 10, the chambers 5d, 5e, 5f, 5l and 5m communicate via
passages 6d, 6e, 6f, 6l and 6m, respectively. At a bottom
(undersurface) of the chamber 9, a square hole is provided. To the
square hole, a DNA microarray 12, on which several tens to several
hundreds of thousand of different species of DNA probes are
arranged in high density on a surface of solid phase, such as a
glass plate having a size of ca. square centimeter, with the probe
surfaces up, is attached.
[0050] It is possible to test a large number of genes at the same
time by effecting a hybridization reaction with the use of the
microarray 12.
[0051] The DNA probes are regularly arranged in a matrix form, and
an address (position determined by the number of row and the number
of column on the matrix) of each of the DNA probes is readily read
as information. The genes to be tested includes, e.g., genetic
polymorphism of each individual in addition to infections viruses,
bacteria and disease-associated genes.
[0052] In the chambers 5a and 5b, a first hemolytic agent
containing EDTA (ethylenediaminetetraacetic acid) for destructing
cell wall and a second hemolytic agent containing a protein
modifying agent such as a surfactant are stored, respectively.
[0053] In the chamber 5c, particles of magnetic material coated
with silica by which DNA is adsorbed are stored. In the chambers 5l
and 5m, a first extraction cleaning liquid and a second extraction
cleaning liquid which are used for purifying DNA at the time of
extraction of DNA are stored, respectively.
[0054] An eluent, comprising a buffer of low-concentration salt,
for eluting DNA from the magnetic particles is stored in the
chamber 5d, a mixture liquid for PCR (polymeraze chain reaction)
comprising a primer, polymerase, a dNTP (deoxyribonucleotide
triphosphate), a buffer, Cy-3dUTP containing a fluorescent agent,
etc., is stored in the chamber 5g. In the chambers 5h and 5j, a
cleaning agent containing a surfactant for cleaning a
fluorescence-labeled specimen DNA, which is not subjected to
hybridization, and a fluorescence label is stored. In the chamber
5i, alcohol for drying the inside of the chamber 9 including the
DNA microarray 12 is stored.
[0055] The chamber 5e is a chamber in which dust other than DNA of
blood accumulates, the chamber 5f is a chamber in which waste of
the first and second extraction cleaning liquids in the chambers 5l
and 5m accumulate, the chamber 5r is a chamber in which waste
liquid of the first and second cleaning liquids accumulate, and the
chambers 5k, 5o and 5t are blank chambers provided for preventing
the solution to flow into the nozzle ports.
[0056] When the liquid specimen such a blood is injected into the
biochemical reaction cartridge described above and the biochemical
reaction cartridge 1 is set in a treatment apparatus described
later, extraction and amplification of DNA or the like are
performed within the cartridge 1. Further, hybridization between
the amplified specimen DNA and DNA probes on the DNA microarray
disposed in the cartridge and cleaning of the fluorescence-labeled
specimen DNA, which is not hybridized, and the fluorescence label
are performed.
[0057] FIG. 3 is a schematic view of the treatment apparatus for
controlling movement of the solution within the biochemical
reaction cartridge and various reactions.
[0058] On a table 13, the biochemical reaction cartridge 1 is
mounted. Further, on the table 13, an electromagent 14 to be
actuated at the time of extracting DNA or the like from the
specimen in the cartridge 1, a Peltier element 15 for effecting
temperature control at the time of amplifying DNA from the specimen
through a method such as PCR (polymerase chain reaction), and a
Peltier element 16 for effecting temperature control at the time of
performing hybridization between the amplified specimen DNA and the
DNA probe on the DNA microarray within the cartridge 1 and at the
time of cleaning or washing the specimen DNA which is not
hybridized, are disposed and connected to a control unit 17 for
controlling the entire treatment apparatus.
[0059] At both side surfaces of the table 13, an electric
(motor-driven) syringe pumps 18 and 19 and pump blocks 22 and 23
each of which is a port for discharging or sucking in air by these
pumps 18 and 19 and is provided with 10 pump nozzles 20 or 21 on
its side surface, are disposed. Between the electric syringe pumps
18 and 19 and the pump nozzles 20 and 21, a plurality of electric
switching (selector) valves (not shown) are disposed and connected
to the control unit 17 together with the pumps 18 and 19. The
control unit 17 is connected to an input unit 24 to which inputting
by a tester is performed. The control unit 17 controls the pump
nozzles 20 and 21 so that each of the respective 10 pump nozzles is
selectively opened and closed with respect to the electric syringe
pumps 18 and 19, respectively.
[0060] In this embodiment, when the tester injects blood as a
specimen into the cartridge 1 through the rubber cap of the
specimen port 2 by a syringe or an injector, the blood flows into
the chamber 7. Thereafter, the tester places the biochemical
reaction cartridge 1 on the table 13 and moves the pump blocks 22
and 23 i directions of arrows indicated in FIG. 3 by operating an
unshown lever, whereby the pump nozzles 20 and 21 are injected into
the cartridge 1 through the corresponding nozzle ports 3 at the
both side surfaces of the cartridge 1.
[0061] Further, the nozzle ports 3a to 3t are concentrated at two
surfaces, i.e., both side surfaces, of the biochemical reaction
cartridge 1, so that it is possible to simplify shapes and
arrangements of the electric syringe pumps 18 and 19, the electric
switching valves, the pump blocks 22 and 23 containing the pump
nozzles, etc. Further, by effecting such a simple operation that
the cartridge 1 is sandwiched between the pump blocks 22 and 23 at
the same time while ensuring necessary chambers 5 and passages, it
is possible to inject the pump nozzles 20 and 21 and simplify the
structure of the pump blocks 22 and 23. Further, all the nozzle
ports 3a to 3t are disposed at an identical level, i.e., are
arranged linearly, whereby all the heights of the passages 4a to 4t
connected to the nozzle ports 3a to 3t become equal to each other.
As a result, preparation of the passages 4a to 4t becomes easy.
[0062] Further, in the treatment apparatus shown in FIG. 3, in the
case where the length of the pump blocks 22 and 23 is increased n
times the original length with respect to n biochemical reaction
cartridges 1, when the n cartridge 1 are arranged in series, it is
possible to perform a necessary step to all the n cartridges 1 at
the same time. As a result, a biochemical reaction can be performed
in the large number of biochemical reaction cartridges with a very
simple apparatus structure.
[0063] Treatment starts when the tester inputs a command of
procedure entry at the input unit 24. FIG. 4 is a flow chart for
explaining a treatment procedure in the treatment apparatus in this
embodiment.
[0064] Referring to FIG. 4, in a step S1, the control unit 24 opens
only the nozzle ports 3a and 3b, and air is discharged form the
electric syringe pump 18 and sucked in the cartridge 1 from the
electric syringe pump 19, whereby the first hemolytic agent 1 is
injected from the chamber 5a into the chamber 7 containing blood.
At this time, by controlling suction of air from the pump 19 so as
to start 10-20 msec after initiation of air discharge from the pump
18, the solution can flow smoothly without causing splash or
scattering thereof at its leading end although it depends on a
viscosity of the hemolytic agent and a resistance of the
passage.
[0065] As described above, by shifting timing of supply and suction
of air so as to control a manner of pressure application and
pressure reduction, it is possible to cause the solution to flow
smoothly. In a preferred embodiment, the solution can be caused to
flow further smoothly by effecting such a control that a degree of
suction of air is linearly increased from the initiation of air
discharge from the pump 18. This is true in the case of subsequent
liquid movement.
[0066] The air supply control can be readily realized by using the
electric syringe pumps 18 and 19. More specifically, after only the
nozzle ports 3a and 3o are opened, discharge and suction of air are
repeated alternately by the pumps 18 and 19 to cause repetitive
flow and flowback of the solution of the chamber 7 in the passage
10, thus stirring the solution. Alternatively, the solution can be
stirred while continuously discharging air from the pump 19 to
generate bubbles.
[0067] FIG. 5 is a sectional view of the biochemical reaction
cartridge 1 shown in FIG. 2 along a cross section intersecting the
chambers 5a, 7 and 5k, and shows such a state that the nozzle port
3a is pressurized by injecting therein the pump nozzle 20 and the
nozzle port 3k is reduced in pressure by injecting therein the pump
nozzle 21, whereby the first hemolytic agent in the chamber 5a
flows into the chamber 7 through the passage 6a. In FIG. 5, in
order to clarify a height (level) relationship, a cross section of
the passage 10 is also shown.
[0068] A volume of the first hemolytic agent in the chamber 5a is
determined so that it ensures a requirement. Further, dimensions
and positions of the chambers 5a and 7 are determined so that the
liquid level in the chamber 7 is lower than a height (vertical
position) of a bottom surface 25 of a connecting portion between
the passage 6a and the chamber 7 when the first hemolytic agent
flows into the chamber 7.
[0069] Referring again to FIG. 4, in a step S2, only the nozzle
ports 3b and 3k are opened and the second hemolytic agent in the
chamber 5b is caused to flow into the chamber 7 in the same manner
as in the case of the first hemolytic agent. Similarly, in a step
53, the magnetic particles in the chamber 5c are caused to flow
into the chamber 7. In the steps S2 and S3, stirring is performed
in the same manner as in the step S1. In the step S3, DNA resulting
from dissolution of cells in the steps S1 and S2 attaches to the
magnetic particles.
[0070] Cross sectional shapes of the chambers 5b and 5c and the
passages 6b and 6c are the same as those of the chamber 5a and the
passage 6a. Volumes of the second hemolytic agent and the magnetic
particle solution are determined so that they ensure their
requirements. Further, dimensions and positions of the chambers 5b,
5c and 7 are determined, similarly as in the step S1, so that the
liquid level in the chamber 7 is lower than height of bottom
surfaces of connecting portions between the passages 6b and 6c and
the chamber 7.
[0071] Incidentally, in this embodiment, the biochemical reaction
cartridge 1 is prepared through ultrasonic fusion bonding of three
injection molded parts 1A, 1B and 1C defined by chain double-dashed
lines indicated in FIG. 5. For convenience of preparation of the
parts, the passages 6a, 6b and 6c are identical in height (vertical
position) to each other. Accordingly, the associated connection
portions are also at the same height. Further, the chambers having
the same height as the chambers 5a, 5b and 5c are the chamber 5k
shown in FIG. 1 and the chambers 5g and 5o shown in FIG. 2.
[0072] By doing so, the reagent is caused to flow from a higher
position than the chamber to be moved, so that it is possible to
smoothly move reliably the entire amount of the reagent stored in
the storage chamber with less resistance. Further, there is such a
case that avoidance of generation of bubbles is desired with
respect to some reagents. In such case, when the movement of the
reagent is performed as described above, the entire amount of the
solution can be moved with a simple structure while avoiding the
generation of bubbles without monitoring completion of movement of
the solution.
[0073] Thereafter, in a step S4, an electromagnet 14 is turned on
and only the nozzle ports 3e and 3k are opened. Then, air is
discharged from the electric syringe pump 19 and sucked in form the
pump 18 to move the solution from the chamber 7 to the chamber 5e.
At the time of movement, the magnetic particles and DNA are trapped
in the passage 10 on the electromagnet 14. The suction and
discharge by the pumps 18 and 19 are alternately repeated to
reciprocate the solution two times between the chambers 7 and 5e,
whereby a trapping efficiency of DNA is improved. The trapping
efficiency can be further improved by increasing the number of
reciprocation. In this case, however, it takes a longer treating
time by that much.
[0074] As described above, DNA is trapped in a flowing state on
such a small passage having a width of about 1-2 mm and a height of
about 0.2-1 mm by utilizing the magnetic particles, so that DNA can
be trapped with high efficiency. This is also true for RNA and
protein.
[0075] FIG. 6 is a sectional view of the cartridge 1 shown in FIG.
2 along a cross section intersecting the chambers 5e, 7 and 5k, and
shows a height relationship between the chambers 5e and 7 and the
passage 6e. The passage 6e connects the bottom portions of the
chambers 5e and 7, so that the movement direction of the solution
is changed to an opposite direction when the suction by the pump 18
and the discharge by the pump 19 are inverted. As a result, when
the suction and the discharge is alternately repeated, it is
possible to reciprocate the solution any number of times between
the chambers 7 and 5e.
[0076] Then, in a step S5, the electromagnet 14 is turned off, and
only the nozzle ports 3f and 3l are opened. Thereafter, air is
discharged from the electric syringe pump 19 and sucked in from the
pump 18 to move the first extraction cleaning liquid from the
chamber 5l to the chamber 5f. At this time, the magnetic particles
and DNA trapped in the step S4 are moved together with the
extraction cleaning liquid, whereby cleaning is performed. After
the reciprocation of two times is performed in the same manner as
in the step S4, the electromagnet 14 is turned on, and the
reciprocation of two times is similarly performed to recover the
magnetic particles and DNA in the passage 10 on the electromagnet
14 and return the solution to the chamber 5l.
[0077] In a step S6, cleaning is further performed in the same
manner as in the step S5 by using the second extraction cleaning
liquid in the chamber 5m in combination with the nozzle ports 3f
and 3m.
[0078] In a step 7, only the nozzle ports 3d and 3o are opened
while the electromagnet 14 is kept on, and air is discharged from
the pump 18 and sucked in from the pump 19, whereby the eluent in
the chamber 5d is moved to the chamber 8.
[0079] At this time, the magnetic particles and DNA are separated
by the action of the eluent, so that only the DNA is moved together
with the eluent to the chamber 8, and the magnetic particles remain
in the passage 10. Thus, extraction and purification of the DNA are
performed. As described above, the chamber containing the
extraction cleaning liquid and the chamber containing waste liquid
after the cleaning are separately provided, so that it becomes
possible to effect extraction and purification of the DNA in the
biochemical reaction cartridge 1.
[0080] Next, in a step S8, only the nozzle ports 3g and 3o are
opened, and air is discharged from the electric syringe pump 18 and
sucked in from the pump 19 to cause the PCR agent in the chamber 5g
to flow into the chamber 8. Further, only the nozzle ports 3g and
3t are opened, and air discharge and suction by the pumps 18 and 19
are repeated alternately to cause the solution in the chamber 8 to
flow. Thereafter, the returning operation is repeated to effect
stirring. Then, the Peltier element 15 is controlled to retain the
solution in the chamber 8 at 96.degree. C. for 10 min. Thereafter,
a cycle of heating at 96.degree. C./10 sec, 55.degree. C./10 sec,
and 72.degree. C./1 min. is repeated 30 times, thus subjecting the
eluted DNA to PCR to amplify the DNA.
[0081] In a step S9, only the nozzle ports 3g and 3t are opened,
and air is discharged from the electric syringe pump 18 and sucked
in from the pump 19 to move the solution in the chamber 8 to the
chamber 9. Further, by controlling the Peltier element 16, the
solution in the chamber 9 is kept at 45.degree. C. for 2 hours to
effect hybridization. At this time, discharge and suction of air by
the pumps 18 and 19 are repeated alternately to move the solution
in the chamber 9 to he passage 6t. Thereafter, the hybridization
proceeds while effecting stirring by repeating the returning
operation.
[0082] In a step S10, while keeping the temperature at 45.degree.
C., only the nozzle ports 3h and 3r are opened, and air is
discharged from the electric syringe pump 18 and sucked in from the
pump 19 to cause the first cleaning liquid in the chamber 5h to
flow into the chamber 5r through the chamber 9 while moving the
solution in the chamber 9 to the chamber 5r. The suction and
discharge by the pumps 18 and 19 are repeated alternately to
reciprocate the solution two times between the chambers 5h, 9 and
5r and finally return the solution to the chamber 5h. Thus, the
fluorescence-labeled specimen DNA and the fluorescence label which
are not hybridized are cleaned.
[0083] FIG. 7 is a sectional view of the biochemical reaction
cartridge 1 shown in FIG. 2 along a cross section intersecting the
chambers 5h, 9 and 5r. The cartridge 1 is pressurized by injecting
the pump nozzle 20 into the nozzle port 3h and is reduced in
pressure by injecting the pump nozzle 21 into the nozzle port 3r.
FIG. 7 illustrates such a state that the first cleaning liquid is
caused to flow into the chamber 5r through the chamber 9.
[0084] Referring again to FIG. 4, in a step S11, while keeping the
temperature at 45.degree. C., the cleaning is further effected in
the same manner as in the step S10 by using the second cleaning
liquid in the chamber 5j in combination with the nozzle ports 3j
and 3r, and the solution is finally returned to the chamber 5j. As
described above, the chambers 5h and 5j containing the cleaning
liquids and the chamber 5r containing waste liquid after the
cleaning are separately provided, so that it becomes possible to
effect extraction and purification of the DNA microarray 12 in the
biochemical reaction cartridge 1.
[0085] In a step 12, only the nozzle ports 3i and 3r are opened,
and air is discharged from the electric syringe pump 18 and sucked
in from the pump 19 to move alcohol in the chamber 5i to the
chamber 5r through the chamber 9. Thereafter, only the nozzle port
3i and 3t are opened, and air is discharged from the pump 18 and
sucked in from the pump 19 to dry the chamber 9.
[0086] When the tester operates a lever (not shown), the pump
blocks 22 and 23 are moved away from the biochemical reaction
cartridge 1. As a result, the pump nozzles 20 and 21 are removed
from the nozzle ports 3 of the cartridge 1. Then, the tester mounts
the cartridge 1 in a reader for DNA array, such a known scanner to
effect measurement and analysis.
[0087] (Embodiment 2)
[0088] FIG. 8 is a sectional view of a biochemical reaction
cartridge 1 of this embodiment, and illustrates a cross section
intersecting the chambers 5a, 7 and 5k shown in FIG. 2 of
Embodiment 1. Further, FIGS. 1 to 4 and 7 in Embodiment 1 are also
applicable to this embodiment.
[0089] The biochemical reaction cartridge 1 is pressurized by
injecting the pump nozzle 20 into the nozzle port 3a and reduced in
pressure by injecting the pump nozzle 21 into the nozzle port 3k.
FIG. 8 illustrates such a state that a first hemolytic agent in the
chamber 5a is caused to flow into the chamber 7 containing blood
through the passage 6a. In order to clarify a height relationship,
a cross section of the passage is also indicated.
[0090] In this embodiment, the passage connecting the chambers 5a
and 7 extends in not only a horizontal direction but also a
vertical direction, so that a (vertical) height of a bottom surface
25 of the connection portion between the passage 6a and the chamber
7 is increased, i.e., a permissible liquid level is increased. As a
result, a mount of a solution to be contained in the chamber 7 is
made larger. If it is not necessary to increase the solution
amount, the height of the biochemical reaction cartridge 1 can be
decreased.
[0091] Further, in the case of preparing the biochemical reaction
cartridge 1 through the injection molding, the vertical portion of
the passage 6a is required in this embodiment. However, it can be
provided by using two injection molded parts A and B defined a
chain double-dashed line shown in FIG. 8. Alternatively, it is also
possible to bond two sheet parts to each other. In this case, the
passage 6a may be tilted to have an oblique surface.
[0092] In the above embodiments (Embodiments 1 and 2), the movement
from the storage chamber is performed with respect to the reagent
but may also be performed with respect to liquid specimen or
cleaning liquid. Further, in the above embodiments, the movement of
liquid is performed by utilizing pressure application and reduction
of air but may also be performed in other manners such that the
cartridge 1 is opened at one side surface and only pressurized or
reduced in pressure at the other side surface, that a pump which
directly moves a solution to be moved is used, and that electrical
movement or movement by utilizing a magnetic force is adopted.
Further, in the above embodiments, a predetermined amount of the
solution is stored in the storage chamber and all the amount of the
solution is moved but, the amount of the moving solution may also
be controlled by a liquid amount sensor or a flow rate sensor.
[0093] As described hereinabove, the biochemical reaction cartridge
according to the present invention moves the solution only therein
by an external pump without incorporating a pump to cause an
necessary reaction to proceed, so that it becomes possible to
provide a disposable cartridge which does not cause outflow of the
solution therefrom with an inexpensive structure. As a result,
possibilities of secondary infection and contamination are
eliminated. Further, the cartridge incorporates therein the
necessary solution, so that it is not necessary to prepare a
reagent and cleaning liquids. As a result, it becomes possible to
realize elimination of labor and prevent an error in selection of
the reagent.
[0094] Further, according to the present invention, air pressure
within the cartridge is controlled by the (external) pump on the
treatment apparatus side to move the solution only within the
cartridge, thus causing a necessary biochemical reaction.
Accordingly, it becomes possible to effect the biochemical reaction
within the cartridge by using the inexpensive biochemical reaction
cartridge.
[0095] Further, the biochemical reaction cartridge according to the
present invention can effect movement with reliability and simple
structure by properly using an optimum passage with respect to both
of movement, for a reagent or specimen, which can be performed only
by causing the reagent or specimen to flow into a subsequent
chamber, and movement of a reaction liquid requiring reciprocating
motion. Further, such an effect that it is possible to move most
efficiently a liquid, such as a reagent or an liquid specimen, to a
subsequent chamber without causing generation of bubbles, can be
attained.
[0096] While the invention has been described with reference to the
structures disclosed herein, it is not confined to the details set
forth and this application is intended to cover such modifications
or changes as may come within the purposes of the improvements or
the scope of the following claims.
* * * * *