U.S. patent application number 13/995576 was filed with the patent office on 2013-10-17 for detection instrument, and detection system.
This patent application is currently assigned to NEC SOFT, LTD.. The applicant listed for this patent is Iwao Waga. Invention is credited to Iwao Waga.
Application Number | 20130273645 13/995576 |
Document ID | / |
Family ID | 46313869 |
Filed Date | 2013-10-17 |
United States Patent
Application |
20130273645 |
Kind Code |
A1 |
Waga; Iwao |
October 17, 2013 |
DETECTION INSTRUMENT, AND DETECTION SYSTEM
Abstract
The present invention provides a detection instrument capable of
easily detecting an intended detection object without any skilled
technique. The detection instrument (1) of the present invention
includes a detection portion (12), a detection reagent which
develops a color by specifically reacting with a detection object
in a sample is placed in the detection portion (12), positional
information of the detection reagent in the detection portion (12)
is information on the detection object, and color development of
the detection reagent can be optically read. It is preferred that a
bar code is formed in the detection portion (12), and the detection
reagent is placed as a part of the bar code.
Inventors: |
Waga; Iwao; (Tokyo,
JP) |
|
Applicant: |
Name |
City |
State |
Country |
Type |
Waga; Iwao |
Tokyo |
|
JP |
|
|
Assignee: |
NEC SOFT, LTD.
Koto-ku, Tokyo
JP
|
Family ID: |
46313869 |
Appl. No.: |
13/995576 |
Filed: |
December 19, 2011 |
PCT Filed: |
December 19, 2011 |
PCT NO: |
PCT/JP2011/079402 |
371 Date: |
June 19, 2013 |
Current U.S.
Class: |
435/287.2 |
Current CPC
Class: |
B01L 3/545 20130101;
B01L 3/5023 20130101; B01L 2300/0627 20130101; C12Q 1/6823
20130101; G01N 21/78 20130101; G01N 33/56983 20130101; G01N 33/5302
20130101; C12Q 1/6837 20130101; G01N 33/56911 20130101; B01L
2300/021 20130101; C12Q 1/6823 20130101; C12Q 2521/543 20130101;
C12Q 2565/513 20130101; C12Q 2525/161 20130101; C12Q 2525/161
20130101; C12Q 2563/125 20130101; C12Q 2525/205 20130101; C12Q
2563/125 20130101; C12Q 2525/205 20130101; C12Q 2565/513 20130101;
C12Q 2521/543 20130101; G01N 33/581 20130101; C12Q 1/6837
20130101 |
Class at
Publication: |
435/287.2 |
International
Class: |
G01N 33/58 20060101
G01N033/58 |
Foreign Application Data
Date |
Code |
Application Number |
Dec 20, 2010 |
JP |
2010-283718 |
Oct 18, 2011 |
JP |
2011-229162 |
Claims
1. A detection instrument comprising: a detection portion, wherein
a detection reagent which develops a color by specifically reacting
with a detection object in a sample is placed in the detection
portion, positional information of the detection reagent in the
detection portion is information on the detection object, and color
development of the detection reagent can be optically read.
2. The detection instrument according to claim 1, wherein a bar
code is formed in the detection portion, and the detection reagent
is placed as a part of the bar code.
3. The detection instrument according to claim 1, wherein a bar
code is formed in the detection portion, besides the bar code, the
detection reagent is placed on a part of or the entire bar code,
and the part of or the entire bar code is indistinguishable by
color development of the detection reagent.
4. The detection instrument according to claim 1, wherein the
detection reagent comprises a nucleic acid element, the nucleic
acid element comprises a first nucleic acid portion and a second
nucleic acid portion, the first nucleic acid portion is a binding
portion which can bind to the detection object, the second nucleic
acid portion is a labeling portion which can distinguish between
binding and non-binding of the first nucleic acid portion and the
detection object, and the labeling portion can cause the detection
reagent to or not to develop a color according to the
distinguishing between the binding and non-binding.
5. The detection instrument according to claim 4, wherein the
detection reagent comprises a labeling substance, when the
detection object does not bind to the first nucleic acid portion,
the second nucleic acid portion can bind to the labeling substance,
and when the detection object binds to the first nucleic acid
portion, the second nucleic acid portion cannot bind to the
labeling substance.
6. The detection instrument according to claim 5, wherein a
secondary structure of the second nucleic acid portion changes by
binding of the detection object to the first nucleic acid portion,
and the labeling substance binding to the second nucleic acid
portion is released from the second nucleic acid portion by the
change of the secondary structure.
7. The detection instrument according to claim 6, wherein the
labeling substance is an enzyme, the detection reagent contains a
chromogenic substrate which develops a color by an enzyme reaction
with the enzyme, when the enzyme binds to the second nucleic acid
portion, the enzyme reaction with the enzyme is inhibited, and when
the enzyme is released from the second nucleic acid portion, the
enzyme reaction is released from being inhibited.
8. The detection instrument according to claim 4, wherein the
second nucleic acid portion can cause an enzyme reaction, the
detection reagent comprises a chromogenic substrate which develops
a color by an enzyme reaction with the second nucleic acid portion,
when the detection object does not bind to the first nucleic acid
portion, the enzyme reaction with the second nucleic acid portion
is inhibited, and when the detection object binds to the first
nucleic acid portion, the enzyme reaction is released from being
inhibited.
9. The detection instrument according to claim 4, wherein the first
nucleic acid portion and the second nucleic acid portion are a
single-stranded nucleic acid obtained by linking to each other.
10. The detection instrument according to claim 4, wherein the
nucleic acid is RNA.
11. The detection instrument according to claim 4, wherein the
first nucleic acid portion and the second nucleic acid portion are
aptamers.
12. The detection instrument according to claim 1, wherein a
positive control which can be read optically is placed in the
detection portion.
13. A detection system comprising: the detection instrument
according to claim 1; and an optical reader, wherein the detection
object is detected by reading color development of the detection
reagent in the detection instrument by the optical reader.
14. The detection system according to claim 13, wherein the
detection instrument is the detection instrument according to claim
2, and the optical reader is a bar-code reader.
15. The detection system according to claim 13, further comprising:
a server, wherein the optical reader is connectable to the server
via a communication network outside of the detection system.
Description
TECHNICAL FIELD
[0001] The present invention relates to a detection instrument and
a detection system.
BACKGROUND ART
[0002] Recently, cases involving food such as misrepresentation
cases and pesticide contamination cases have occurred. Further,
cases of food poisoning caused by microorganisms such as an
Escherichia coli O157 strain and Salmonella enterica, and
food-borne infectious diseases such as a Creutzfeldt-Jakob disease
caused by a pathogenic protein and the like have occurred.
Furthermore, accompanying the arrival of an aging society, public
health consciousness is spreading. Because of this, public
awareness on food is high, and safe and healthy food is required.
In order to ensure the quality of food, a system for inspecting the
quality of food is necessary in a food production stage, a food
distribution stage, and a food consumption stage. On the other
hand, as a method for specifically detecting a specific substance,
there is a method utilizing an antigen-antibody reaction. For
example, the patent document 1 discloses a method for inspecting
components in food, using an antigen-antibody reaction.
PRIOR ART DOCUMENTS
Patent Document
[0003] Patent Document 1: JP 2009-133712 A
SUMMARY OF INVENTION
Problem to be Solved by the Invention
[0004] Sanitation inspections and contamination inspections are
carried out in the food production stage because of the requirement
by the food sanitation law and the voluntary management by food
companies and the like. However, skilled techniques are required in
order to carry out these inspections of food quality. Therefore, it
is rare that food quality is inspected in a distribution stage and
a consumption stage. That is, it is impossible to inspect food
quality by part-time workers having no skilled technique in
supermarkets, department stores, convenience stores, and
restaurants in the same manner as in food factories. The same
applies to inspections of chemical substances such as pesticides in
a food production stage. It is required to remove such difficulties
in inspections not only in the food field, but also in all of
fields relating to public health such as the medical field and the
agricultural field.
[0005] Hence, the present invention is intended to provide a
detection instrument and a detection system, capable of easily
detecting an intended detection object without any skilled
technique.
Means for Solving Problem
[0006] The detection instrument according to the present invention
is a detection instrument including: a detection portion, wherein a
detection reagent which develops a color by specifically reacting
with a detection object in a sample is placed in the detection
portion, positional information of the detection reagent in the
detection portion is information on the detection object, and color
development of the detection reagent can be optically read.
[0007] The detection system according to the present invention is a
detection system including: the detection instrument according to
the present invention; and an optical reader, wherein the detection
object is detected by reading color development of the detection
reagent in the detection instrument by the optical reader.
Effects of the Invention
[0008] The present invention uses an easy method of optically
reading color development of a detection reagent. Thus, according
to the present invention, everyone can easily detect an intended
detection object without any skilled technique.
BRIEF DESCRIPTION OF DRAWINGS
[0009] FIG. 1A is a plan view showing an example of a configuration
of a detection instrument of Embodiment 1 according to the present
invention. FIG. 1B is a plan view showing another example of a
configuration of a detection instrument of Embodiment 1 according
to the present invention.
[0010] FIGS. 2A and 2B are plan views illustrating an example of
detection of a detection object in a detection portion of the
detection instrument of Embodiment 1.
[0011] FIGS. 3A to 3D are plan views showing another example of
detection of a detection object in a detection portion of the
detection instrument of Embodiment 1.
[0012] FIGS. 4A to 4D are plan views showing yet another example of
detection of a detection object in a detection portion of the
detection instrument of Embodiment 1.
[0013] FIGS. 5A and 5B are diagrams schematically showing a
detection method for detecting a detection object using a detection
instrument of Embodiment 1-1a according to the present
invention.
[0014] FIGS. 6A and 6B show a specific example of the detection
method in Embodiment 1-1a.
[0015] FIGS. 7A and 7B are diagrams schematically showing a
detection method for detecting a detection object using a detection
instrument of Embodiment 1-1b according to the present
invention.
[0016] FIGS. 8A to 8D show a specific example of the detection
method in Embodiment 1-1b.
[0017] FIG. 9 is a schematic view illustrating antibodies
immobilized on a detection portion in a detection instrument of
Embodiment 1-2 according to the present invention.
[0018] FIG. 10 is a schematic view illustrating the state of an
antigen-antibody reaction in the detection instrument of Embodiment
1-2.
[0019] FIG. 11 is a schematic view showing an example of a
configuration of a detection system of Embodiment 2 according to
the present invention.
[0020] FIG. 12 is a schematic view showing an example of a
configuration of a detection system of Embodiment 3 according to
the present invention.
[0021] FIGS. 13A to 13C are views showing the respective detection
instruments of Embodiments 4 to 6 according to the present
invention.
DESCRIPTION OF EMBODIMENTS
[0022] In the present invention, the concept of "color development"
encompasses coloring. In the present invention, the concept of
"detection" encompasses quantitative analysis, semi-quantitative
analysis, and qualitative analysis.
[0023] The embodiments of the present invention are described. The
present invention, however, is not limited by the following
embodiments.
Embodiment 1
[0024] FIG. 1A shows an example of a configuration of a detection
instrument according to the present invention. As shown in FIG. 1A,
this detection instrument 1 is configured so that a detection
portion 12 is formed in a main body 11. A detection reagent which
develops a color by specifically reacting with a detection object
in a sample is placed in the detection portion 12.
[0025] In the detection portion 12, positional information of the
detection reagent is information on the detection object.
[0026] It is preferred that a bar code is formed in the detection
portion 12 of the detection instrument according to the present
invention as shown in FIG. 1A, and the detection reagent is placed
as a part of the bar code. The detection portion (bar code) 12 may
include, besides information on the detection object, information
such as information on the detection reagent (information on the
kind of the detection reagent and the like) other than the
positional information and information on a site at which detection
is performed using the detection information, for example. With
this embodiment, it is possible to detect the detection object with
reference to information on the kind of the detection reagent,
information on the site at which detection is performed, and the
like. The bar code is not limited to a one-dimensional code shown
in FIG. 1A and may be, for example, a two-dimensional code such as
a QR code (registered trademark) shown in FIG. 1B.
[0027] The detection instrument shown in FIG. 1A can be produced by
forming a detection portion (bar code) 12 in a main body 11 of a
detection instrument 1. A method for forming the detection portion
(bar code) 12 is not particularly limited and can be, for example,
a method such as printing using a printer such as an ink-jet
recording device or a laser printer. In this case, for example, by
causing the detection reagent to be contained in an ink or a toner
at the time of printing a part of the detection portion (bar code)
12, the detection reagent is placed as the part of the detection
portion (bar code) 12.
[0028] The detection reagent is not particularly limited as long as
it develops a color by specifically reacting with a detection
object in a sample, and examples thereof include a nucleic acid
element described below, an antibody, and an antigen.
[0029] The method for using the detection instrument shown in FIG.
1A is, for example, as follows. Note here that the following method
is a mere example, and the present invention is not limited by
this.
[0030] First, the detection portion (bar code) 12 is brought into
contact with a sample. At that time, when a detection object is
present in the sample, the detection reagent specifically reacts
with the detection object and thus develops a color.
[0031] In the present invention, the sample is not particularly
limited, and examples thereof include food (including beverages),
pharmaceuticals, chemicals, the ground, animals, plants,
microorganisms, virus, water (e.g., tap water, discharged water,
river water, seawater, rainwater, and snow), garbage, and
waste.
[0032] In the present invention, the detection object is not
particularly limited, and examples thereof include high-molecular
compounds, low-molecular compounds, organic substances, and
inorganic substances. Examples of the high-molecular compounds or
the organic substances include microorganisms, virus,
polysaccharides, proteins, nucleic acids, and resins. Examples of
the low-molecule compounds include pesticides, pharmaceuticals,
chemicals, oligosaccharides, monosaccharides, lipids,
oligopeptides, amino acids, vitamins, and bioactive substances.
Examples of the inorganic substances include minerals, mineral
acids, and metals.
[0033] An example of detecting the detection object in the
detection portion is described with reference to FIGS. 2A and 2B.
In FIGS. 2A and 2B, the identical parts to those in FIGS. 1A and 1B
are denoted by identical reference numerals. In FIG. 2A, a bar code
is formed in the detection portion 12, and a detection reagent 1a
is placed as a part of the bar code. In the detection portion (bar
code) 12, positional information of the detection reagent 1a is
information on the detection object. The detection reagent 1a is
placed adjacent to a bar 1b in the detection portion (bar code) 12.
At the time when a sample is brought into contact with the
detection portion (bar code) 12, when the detection object is
present in the sample, the detection reagent 1a placed adjacent to
the bar 1b specifically reacts with the detection object and
develops a color as shown in FIG. 2B. For example, when the
detection object is an Escherichia coli O157 strain, a detection
reagent which develops a color by contact with an Escherichia coli
O157 strain is used as the detection reagent 1a. As described
above, when the detection object is present in the sample, the
thickness of the bar 1b is increased by color development of the
detection reagent 1a as shown in FIG. 2B. On the other hand, when
the detection object is not present in the sample, the bar 1b is
thin as it is (in the state shown in FIG. 2A). Whether or not the
thickness of the bar 1b is increased is read by a bar-code reader
and detected.
[0034] Another example of detection of the detection object in the
detection portion is described with reference to FIGS. 3A to 3D. In
FIGS. 3A to 3D, the identical parts to those in FIGS. 1A to 2B are
denoted by identical reference numerals. In FIG. 3A, a bar code is
formed in the detection portion 12, and three detection reagents
1a, 2a, and 3a are placed as parts of the bar code. For example, a
detection reagent for an Escherichia coli O157 strain as the
detection reagent 1a, a detection reagent for Salmonella enterica
as the detection reagent 2a, a detection reagent for Staphylococcus
as the detection reagent 3a are placed. In the detection portion
(bar code) 12, three pieces of positional information of the
detection reagents 1a, 2a, and 3a are the respective pieces of
information on the three detection objects (Escherichia coli O157
strain, Salmonella enterica, and Staphylococcus). The detection
reagents 1a, 2a, and 3a are placed adjacent to the respective bars
1b, 2b, and 3b in the detection portion (bar code) 12. At the time
when a sample is brought into contact with the detection portion
(bar code) 12, when all of Escherichia coli O157 strain, Salmonella
enterica, and Staphylococcus are present in the sample, the three
detection reagents 1a, 2a, and 3a specifically react with the
respective three detection objects (Escherichia coli O157 strain,
Salmonella enterica, and Staphylococcus) and develop a color as
shown in FIG. 3B. When an Escherichia coli O157 strain and
Salmonella enterica are present in the sample, two detection
reagents 1a and 2a specifically react with the respective two
detection objects (Escherichia coli O157 strain and Salmonella
enterica) and develop a color as shown in FIG. 3C. When only an
Escherichia coli O157 strain is present in the sample, the
detection reagent 1a specifically reacts with the detection object
(Escherichia coli O157 strain) and develops a color as shown in
FIG. 3D. As described above, when any of the three detection
objects (an Escherichia coli O157 strain, Salmonella enterica, and
Staphylococcus) is present in the sample, the thickness of a
corresponding bar among the bars 1b, 2b, and 3b is increased as
shown in FIGS. 3B to 3D. On the other hand, when none of the three
detection objects (an Escherichia coli O157 strain, Salmonella
enterica, and Staphylococcus) is present in the sample, the bars
1b, 2b, and 3b are thin as they are (in the state shown in FIG.
3A). Then, whether or not any of the thicknesses of the bars 1b,
2b, and 3b is increased is read by a bar-code reader and detected.
In FIGS. 3A to 3D, the number of detection reagents is set to
three, and the number of detection reagents, however, can be
increased or decreased if necessary. With this configuration, a
plurality of detection objects can be detected simultaneously.
[0035] In the detection instrument according to the present
invention shown in FIG. 1A, besides the bar code formed in the
detection portion 12, the detection reagent may be placed on a part
of or the entire bar code, and the part of or the entire bar code
may be indistinguishable by color development of the detection
reagent. The present embodiment is described with reference to
FIGS. 4A to 4D. In FIGS. 4A to 4D, the identical parts to those in
FIGS. 1A to 3D are denoted by identical reference numerals. In FIG.
4A, a bar code is formed in a detection portion 12, and besides the
bar code, eleven detection reagents 1a are placed on the respective
parts of the barcode. In the detection portion 12, positional
information of the eleven detection reagents 1a is information on
the detection object. At the time when a sample is brought into
contact with the detection portion 12, when the detection object is
present in the sample, the detection reagents 1a specifically react
with the detection object and develop a color as shown in FIG. 4B.
For example, in the case where the detection object is an
Escherichia coli O157 strain, a detection reagent which develops a
color by contact with an Escherichia coli O157 strain is used as
the detection reagents 1a. It is checked and detected that a part
of the bar code is indistinguishable by color development of the
detection reagent 1a using a bar-code reader. In FIG. 4C, a bar
code is formed in the detection portion 12, and besides the bar
code, a detection reagent 1a is placed on the entire bar code. In
the detection portion 12, positional information of the detection
reagent 1a is information on the detection object. At the time when
a sample is brought into contact with the detection portion 12,
when the detection object is present in the sample, the detection
reagent 1a specifically reacts with the detection object and
develops a color as shown in FIG. 4D. For example, in the case
where the detection object is an Escherichia coli O157 strain, a
detection reagent which develops a color by contact with an
Escherichia coli O157 strain is used as the detection reagent 1a.
It is checked and detected that the entire bar code is
indistinguishable by color development of the detection reagent 1a
using a bar-code reader.
Embodiment 1-1
[0036] The present embodiment is an example of a detection
instrument in the case where the detection reagent contains a
nucleic acid element. The detection instrument of the present
embodiment is a detection instrument wherein the detection reagent
contains a nucleic acid element, the nucleic acid element contains
a first nucleic acid portion and a second nucleic acid portion, the
first nucleic acid portion is a binding portion which can bind to
the detection object, the second nucleic acid portion is a labeling
portion which can distinguish between binding and non-binding of
the first nucleic acid portion and the detection object, and the
labeling portion can cause the detection reagent to or not to
develop a color according to the distinguishing between the binding
and non-binding.
[0037] <Nucleic Acid>
[0038] In the present embodiment, the kind of the nucleic acid is
not particularly limited, and examples thereof include a single
strand such as a single-stranded RNA or a single-stranded DNA and a
double-stranded nucleic acid such as a double-stranded RNA or a
double-stranded DNA. In the present embodiment, the nucleic acids
may be used alone or in a combination of two or more of them. The
nucleic acid may have a secondary structure formed by
self-annealing, for example. The secondary structure can be, for
example, a stem-loop structure.
[0039] The nucleic acid may have, for example, a naturally-derived
nucleic acid sequence or a synthesized nucleic acid sequence. A
method for synthesizing the nucleic acid is not at all limited and
can be, for example, a method in which a nucleic acid is chemically
synthesized from terminal bases using dNTP or the like as a
material by a DNA synthesizer or an RNA synthesizer. Examples of
the nucleic acid include DNA and RNA as mentioned above. The
nucleic acid may contain peptide nucleic acid such as PNA, for
example. The nucleic acid may contain a natural nucleic acid
(nonartificial nucleic acid) such as A, C, G, T, or U or may
contain an artificial nucleic acid such as 2'-fluorouracil,
2'-aminouracil, 2'-O-methyluracil, or 2-thiouracil, for
example.
[0040] In the detection instrument according to the present
embodiment, the nucleic acid element preferably is a
single-stranded nucleic acid obtained by linking the first nucleic
acid portion and the second nucleic acid portion. As described
above, when the nucleic acid element is a single-stranded nucleic
acid, the secondary structure changes by binding of a detection
object to the first nucleic acid portion, and this change is prone
to cause a change in secondary structure of the second nucleic acid
portion. The present invention, however, is not limited by this,
the first nucleic acid portion and the second nucleic acid portion
may be linked via a linker.
[0041] The nucleic acid preferably is an aptamer, for example. The
aptamer generally means a nucleic acid molecule which can
specifically bind to a specific target substance.
[0042] The aptamer may be, for example, DNA or RNA as mentioned
above and may be a single-stranded nucleic acid such as a
single-stranded RNA or a single-stranded DNA or a double-stranded
nucleic acid such as a double-stranded RNA or a double-stranded
DNA. The aptamer may be a naturally-derived nucleic acid sequence
or a synthesized nucleic acid sequence and may contain peptide
nucleic acid such as PNA, the natural nucleic acid (nonartificial
nucleic acid), or the artificial nucleic acid.
[0043] A method for producing an aptamer is not particularly
limited, and it can be produced by the above-mentioned
conventionally known method, for example. As mentioned above, for
example, in the case of introducing an aptamer which can bind to a
target substance into an animal in the present embodiment, the
conventionally known SELEX (Systematic Evolution of Ligands by
Exponential Enrichment) method or the like can be employed in the
aptamer production, for example.
[0044] The preparation of an aptamer by the SELEX method is not
particularly limited and can be carried out as follows, for
example. First, a nucleic acid pool containing a plurality of
nucleic acids is provided. Then, the nucleic acid library and a
target substance are bound to (associated with) each other to form
complexes between them. Thereafter, only a nucleic acid pool
involved in formation of the complexes is collected from the
complexes. Thus, nucleic acid aptamers which can specifically bind
to the secondary reagent can be prepared. A method for preparing
nucleic acid aptamers which can specifically bind to a target
substance using the SELEX method is shown below as an example. The
present invention, however, is not at all limited by this
method.
[0045] The nucleic acid pool is, for example, a library (mixture)
of nucleic acids each having a random region. Examples of the
nucleic acids in the library include polynucleotides such as RNAs
and DNAs. The random region is, for example, a region in which
bases of A, G, C, and U or bases of A, G, C, and T are randomly
linked, and the length thereof is, for example, in the range from
20 to 120 mer. The nucleic acid pool includes preferably from
4.sup.20 to 4.sup.120 types (about from 10.sup.12 to 10.sup.72
types) of nucleic acids, more preferably from 4.sup.30 to 4.sup.60
types (about from 10.sup.18 to 10.sup.36 types) of nucleic
acids.
[0046] It is only necessary that each of the polynucleotides
contained in the nucleic acid pool has the random region, for
example, and the other configuration is not particularly limited.
It is preferred that each of the polynucleotides has, for example,
in addition to the random region, a primer region utilized in
nucleic acid amplification described below, a polymerase
recognition region that can be recognized by a polymerase, and the
like, at at least one of the 5'-end and 3'-end of the random
region. The polymerase recognition region can be decided as
appropriate according to the type of polymerase to be used in the
nucleic acid amplification described below in preparation of
aptamers, for example. In the case where the nucleic acid pool is
an RNA pool, the polymerase recognition region is, for example,
preferably a DNA-dependent RNA polymerase recognition region
(hereinafter, also referred to as an "RNA polymerase recognition
region".), and specifically, a T7 promoter that is a T7 RNA
polymerase recognition region. A specific example of the RNA pool
can be, for example, an RNA pool containing RNAs each having a
structure in which, from the 5'-end side thereof, the RNA
polymerase recognition region and the primer region (hereinafter,
also referred to as a "5'-end side primer region".) are linked in
this order, the random region is linked to the 3'-end side of the
5'-end side primer region, and the primer region (hereinafter, also
referred to as a "3'-end side primer region".) is linked to the
3'-end side of the random region. It is preferred that the 5'-end
side primer region in the RNA is, for example, a sequence
complementary to the 3'-end of a DNA antisense strand synthesized
using the RNA as a template, i.e., a sequence that is the same as a
sequence of a primer that can bind to the 3'-end of the antisense
strand. Moreover, the RNA pool may further include a region that
assists the binding to a secondary reagent which is a target
substance, for example. Each of the polynucleotides in the nucleic
acid pool may have a different random region or a random region a
part of which is a common sequence. The respective regions in each
of the polynucleotides may be directly adjoined to one another or
may be indirectly adjoined through intervening sequences.
[0047] A method for preparing the nucleic acid pool is not
particularly limited, and a known method can be employed. In the
case where the nucleic acid pool is an RNA pool, the nucleic acid
pool can be prepared using an initial pool containing DNAs and, as
templates, the DNAs, for example. Hereinafter, a DNA strand used as
a template of RNAs in a nucleic acid pool is also referred to as an
antisense strand, and a DNA strand having a sequence of any of the
RNAs with U replaced by T is also referred to as a sense strand. It
is preferred that the initial pool containing DNAs contains, for
example, any of DNAs (antisense strands) each obtained by replacing
U in a strand complementary to each random region in the RNA pool
by T and DNAs (sense strands) each having a sequence obtained by
replacing U in each random region by T. Nucleic acid amplification
is conducted using each of the DNAs in this initial pool as a
template and a DNA-dependent DNA polymerase. Thereafter, a
transcription reaction is conducted using each of obtained DNA
amplification products as a template and a DNA-dependent RNA
polymerase. Thus, a nucleic acid pool containing RNAs can be
prepared.
[0048] It is also possible to prepare a nucleic acid pool
containing RNAs by nucleic acid amplification through a preparation
of an initial pool containing DNAs each obtained by replacing U in
each random region of each of the RNAs by T and annealing of
primers each having an RNA polymerase recognition region and a
sequence complementary to a 5'-end side primer region, using the
initial pool as a template.
[0049] Then, the nucleic acid pool and a target react with each
other. Thus, a composite of the nucleic acid pool and the target is
formed. In the preparation of aptamers, a target substance that
reacts with the nucleic acid pool may be, for example, the
above-described target substance or a degradate thereof A binding
form between the nucleic acid pool and the target is not
particularly limited and can be, for example, binding via
intermolecular force such as a hydrogen bond. A treatment for
binding the nucleic acid pool and the target substance can be, for
example, a method in which the both are incubated for a certain
period of time in a solvent. The solvent is not particularly
limited and preferably the one can maintain the binding of the both
and the like. Examples of the solvent include various buffer
solutions.
[0050] Subsequently, the composite of the nucleic acid pool and the
target is collected. A reaction solution in which the both are
caused to react with each other in order to form a composite
contains, besides the composite, a nucleic acid pool (hereinafter
referred to as an "unreacted nucleic acid pool") that does not
involved in formation of the composite, for example. Therefore, for
example, it is preferred that the composite and the unreacted
nucleic acid pool are separated from the reaction solution. A
method for separating the composite and the unreacted nucleic acid
pool from each other is not particularly limited and can be, for
example, a method utilizing the difference in adsorbability between
the target substance and the nucleic acid pool or the difference in
molecular weight between the composite and the nucleic acid
pool.
[0051] As the former method utilizing the difference in
adsorbability, the following method is illustrative, for example.
That is, first, a carrier having adsorbability to the target
substance and the reaction solution containing the composite are
brought into contact with each other. At that time, the unreacted
nucleic acid pool is not adsorbed to the carrier. In contrast, the
composite of the target substance and the nucleic acid pool is
adsorbed to the same. Thus, the unreacted nucleic acid pool and the
composite can be separated from each other. Therefore the composite
adsorbed to the carrier can be collected after removing the
unreacted nucleic acid pool. It is preferred that the carrier is
washed in order to completely remove the unreacted nucleic acid
pool prior to collection of the composite from the carrier, for
example. The carrier having adsorbability to the target substance
is not particularly limited and can be selected as appropriate
according to the type of the target, for example. In the case where
the target substance is, for example, a protein such as an
antibody, the carrier having adsorbability can be, for example, a
nitrocellulose film.
[0052] As the latter method utilizing the difference in molecular
weight, a method using a carrier can be illustrative, for example.
The carrier can be, for example, a carrier having pores each with a
pore size with which the nucleic acid pool is allowed to pass
therethrough, but the composite is not allowed to pass
therethrough. By utilizing such carrier, the composite and the
unreacted nucleic acid pool can be separated from each other. The
separation may be, for example, electrical separation using an
agarose gel, a polyacrylamide gel, or the like.
[0053] Besides these methods, a method using a target substance
immobilized on a carrier in formation of composite can be used, for
example. That is, the target substance is previously immobilized on
a carrier, and the carrier and the nucleic acid pool are brought
into contact with each other. Thus, a composite of the immobilized
target substance and the nucleic acid pool is formed. Then, an
unreacted nucleic acid pool binding to no immobilized target
substance is removed, and thereafter the composite of the target
substance and the nucleic acid pool is dissociated from the
carrier. A method for immobilizing the target substance on the
carrier is not at all limited, and a known method can be employed.
Specifically, the method can be, for example, a method in which the
target substance is previously bound to a label, and a carrier
having a ligand with the label and the target substance binding to
the label are brought into contact with each other. The label can
be, for example, a His-tag. Examples of the ligand include metal
ions such as a nickel ion (Ni.sup.2+) and a cobalt ion (Co.sup.2+).
Specific examples of the carrier include Ni-agarose and
Ni-sepharose based on the metal ions.
[0054] Then, a nucleic acid pool involved in formation of the
composite is collected from the collected composite. The nucleic
acid pool involved in formation of the composite can be collected
by releasing a bond between the target substance and the nucleic
acid pool, for example.
[0055] Subsequently, nucleic acid amplification of the collected
nucleic acid pool involved in formation of the composite is
conducted. A method for amplifying the nucleic acid pool is not
particularly limited, and the nucleic acid pool can be amplified by
a known method according to the type of the nucleic acid pool, for
example. In the case where the nucleic acid pool is an RNA pool,
for example, first, cDNAs are prepared by a reverse transcription
reaction using an RNA-dependent DNA polymerase, and nucleic acid
amplification of DNAs is conducted by a PCR or the like using the
each of the cDNAs as a template. Then, using each of amplification
products thus obtained as a template and using, for example, a
DNA-dependent RNA polymerase, a transcription of RNAs is conducted.
Thus, the RNA pool involved in formation of the composite can be
amplified.
[0056] When the RNA pool contains an RNA polymerase recognition
region, a 5'-end side primer region, a random region, and a 3'-end
side primer region, the nucleic acid amplification can be conducted
by an amplification method utilizing these regions, for example. In
a reverse transcription reaction for preparing the cDNAs using each
of the RNAs as a template, it is preferred that a polynucleotide
having a sequence complementary to the 3'-end side primer region
contained in the RNA pool is used as a primer, for example.
Further, in amplification of DNAs using each of the cDNAs as a
template, it is preferred that a polynucleotide having the 5'-end
side primer region and a polynucleotide having a strand
complementary to the 3'-end side primer region are used as primers,
for example. It is preferred that the former polynucleotide further
has the RNA polymerase recognition region on the 5'-end side
thereof and the 5'-end side primer region on the 3' side thereof,
for example. In amplification of RNAs using each of obtained
amplification products of DNAs as a template, nucleic acid
amplification such as a PCR is conducted using each of the DNA
amplification products as a template, a 5'-end side primer region
and the 3'-end side primer region in each of the DNAs, and a
DNA-dependent DNA polymerase. In this case, for example, it is
preferred that a polynucleotide containing the 5'-end side primer
region and a polynucleotide containing a strand complementary to
the 3'-end side primer region are used as primers. It is preferred
that the former polynucleotide has the RNA polymerase recognition
region on the 5'-end side thereof and the 5'-end side primer region
on the 3' side thereof, for example. Then, a transcription reaction
in vitro is conducted using each of obtained amplification products
as a template, the RNA polymerase recognition region in each of the
amplification products, and the DNA-dependent RNA polymerase. Thus,
nucleic acid amplification of the RNA pool involved in formation of
the composite can be conducted. In each of the amplification
products, a DNA of an antisense strand has an RNA polymerase
recognition region on the 3'-end side thereof, for example.
Therefore, the DNA-dependent RNA polymerase is bound to this
region, and each of the RNAs can be synthesized using the antisense
strand as a template. The RNA-dependent DNA polymerase used in the
reverse transcription reaction is not particularly limited, and a
reverse transcriptase derived from avian myeloblastosis virus (AMV
Reverse Transcriptase) can be used, for example.
[0057] The method for amplifying nucleic acid is not particularly
limited, and for example, any of a PCR method and various
isothermal amplification methods can be employed. The conditions
thereof are also not particularly limited.
[0058] As described above, a nucleic acid pool forming a composite
with a target substance is collected. Further, as mentioned above,
formation of composite using a target substance, collection of the
composite, separation of a nucleic acid pool involved in formation
of the composite, amplification of the separated nucleic acid pool,
and the like are repeated. Thus, nucleic acid aptamers having
binding properties to the target substance can be eventually
obtained.
[0059] <Form of Nucleic Acid Element>
[0060] Examples of the nucleic acid element in the present
embodiment include the following two forms (1) and (2):
(1) a nucleic acid element in which the second nucleic acid portion
can bind to a labeling substance when a detection object binds to
the first nucleic acid portion, and the second nucleic acid portion
cannot bind to the labeling substance when a detection object binds
to the first nucleic acid portion; and (2) a nucleic acid element
in which the second nucleic acid portion can cause an enzyme
reaction, the enzyme reaction with the second nucleic acid portion
is inhibited when a detection object does not bind to the first
nuclei acid portion, and the enzyme reaction is released from being
inhibited when a detection object binds to the first nucleic acid
portion.
[0061] In the form (1), the "labeling substance" is an optional
component of the nucleic acid element of the present
embodiment.
[0062] The form (1) may be the one in which the secondary structure
of the second nucleic acid portion changes by binding of the
detection object and the first nucleic acid portion, and by the
change in the secondary structure, the labeling substance binding
to the second nucleic acid portion is released from the second
nucleic acid portion. In this case, the form may be the one in
which the labeling substance is an enzyme, and an enzyme reaction
with the enzyme is inhibited when the enzyme binds to the second
nucleic acid portion, and the enzyme reaction is released from
being inhibited when the enzyme is released from the second nucleic
acid portion.
Embodiment 1-1a
[0063] The present embodiment is an example of a detection
instrument including a nucleic acid element in the form (1). FIGS.
5A and 5B schematically shows a configuration of the nucleic acid
element of the present embodiment. As shown in FIGS. 5A and 5B,
this nucleic acid element 36 includes a first nucleic acid portion
(binding portion) 32 and a second nucleic acid portion (labeling
portion) 33, and the first nucleic acid portion 32 and the second
nucleic acid portion 33 are integrated by linking to each other. As
shown in FIG. 5A, when a detection object 31 does not bind to the
first nucleic acid portion 32, a labeling substance (e.g., enzyme)
34 binds to the second nucleic acid portion 33. Then, as shown in
FIG. 5B, when the detection object 31 is present in a sample and
binds to the first nucleic acid portion 32, the structure of the
second nucleic acid portion 33 changes, resulting in releasing the
labeling substance 34 from the second nucleic acid portion 33. In
this case, the labeling substance 34 is an enzyme, the enzyme
reaction inhibited by binding the enzyme to the second nucleic acid
portion 33 is released from being inhibited by releasing the enzyme
from the second nucleic acid portion 33. At that time, when a
substrate is present, an enzyme reaction occurs. Thus, the
detection object can be detected by detecting this enzyme
reaction.
[0064] In the first nucleic acid portion and the second nucleic
acid portion, the length of each nucleic acid is not particularly
limited. The lower limit of the length in each of the first nucleic
acid portion and the second nucleic acid portion is not
particularly limited and is, for example, 7 bases. The upper limit
of the same is not particularly limited and is, for example, 120
bases, preferably 80 bases, more preferably 35 bases, yet more
preferably 20 bases, and the shorter the length the more
preferable. The range of the length in the first nucleic acid
portion is, for example, from 7 to 120 bases, preferably from 7 to
80 bases, more preferably from 7 to 35 bases, yet more preferably
from 7 to 20 bases. The upper limit of the length of nucleic acid
in the second nucleic acid portion is not particularly limited and
is, for example, 120 bases, preferably 80 bases, more preferably 60
bases, yet more preferably 40 bases, particularly preferably 35
bases, and the shorter the length the more preferable. The range of
the length in the second nucleic acid portion is, for example, from
7 to 120 bases, preferably from 7 to 80 bases, more preferably from
7 to 60 bases, yet more preferably from 7 to 40 bases, particularly
preferably from 7 to 35 bases. The length of nucleic acid in the
entire nucleic acid element is not particularly limited, and the
range of the length is, for example, from 14 to 240 bases,
preferably from 14 to 200 bases, more preferably from 14 to 160
bases, yet more preferably from 14 to 140 bases, particularly
preferably from 14 to 75 bases, more particularly preferably from
14 to 55 bases. The length in the first nucleic acid portion may be
identical to or different from that in the second nucleic acid
portion.
[0065] The substrate is not particularly limited, and is, for
example, preferably a chromogenic substrate that develops a color
by an enzyme reaction because it allows detection to be carried out
easily. In this case, examples of the enzyme include oxidoreductase
and phosphatase. The oxidoreductase can be, for example,
peroxidase. The phosphatase can be, for example, alkaline
phosphatase. When the enzyme is peroxidase, examples of the
chromogenic substrate includes 3,3',5,5'-tetramethylbenzidine
(TMB), 1,2-phenylenediamine (OPD), 2,2'-azinobis
(3-ethylbenzothiazoline-6-sulfonic acid ammonium salt (ABTS),
3,3'-diaminobenzidine (DAB), 3,3'-diaminobenzidine
tetrahydrochloride hydrate (DAB4HCl), 3-amino-9-ethylcarbazole
(AEC), 4-chloro-1-naphthol (4ClN), 2,4,6-tribromo-3-hydroxybenzoic
acid, 2,4-dichlorophenol, 4-aminoantipyrine, and 4-aminoantipyrine
hydrochloride. When the enzyme is alkaline phosphatase, examples of
the chromogenic substrate include
5-bromo-4-chloro-3-indolylphosphate/nitrotetrazolium blue
(Nitro-TB), and nitro-blue tetrazolium chloride (NBT).
[0066] Next, FIGS. 6A and 6B show an example of a nucleic acid
element 36 in which aptamers are employed as a first nucleic acid
portion 32 and a second nucleic acid portion 33. In FIGS. 6A and
6B, the identical parts to those in FIGS. 5A and 5B are denoted by
identical reference numerals. In this nucleic acid element 36, the
aptamer as the first nucleic acid portion 32 and the aptamer as the
second nucleic acid portion 33 bind to each other to form a
single-stranded nucleic acid (RNA). As shown in FIG. 6A, when a
detection object 31 does not bind to the aptamer being the first
nucleic acid portion 32, a labeling substance (e.g., enzyme) 34
binds to the aptamer being the second nucleic acid portion 33.
Then, as shown in FIG. 6B, when the detection object 31 is present
in a sample and binds to the aptamer being the first nucleic acid
portion 32, the structure of the aptamer as the second nucleic acid
portion 33 changes, resulting in releasing the labeling substance
34 from the second nucleic acid portion 33. In this case, the
labeling substance 34 is an enzyme, and an enzyme reaction
inhibited by binding of the enzyme to the second nucleic acid
portion 33 is released from being inhibited by releasing the enzyme
from the second nucleic acid portion 33. At that time, when a
substrate is present, an enzyme reaction occurs. Thus, the
detection object 31 can be detected by detecting this enzyme
reaction. Note here that FIGS. 6A and 6B are a mere example, and as
long as the second nucleic acid portion 33 can bind to the labeling
substance 34 when the detection object 31 does not bind to the
first nucleic acid portion 32, and the second nucleic acid portion
33 cannot bind to the labeling substance 34 when the detection
object 31 binds to the first nucleic acid portion 32, the other
configuration of the nucleic acid element 36 is not particularly
limited.
[0067] The nucleic acid element including aptamers described above
can be produced as follows, for example. First, an aptamer as a
first nucleic acid portion is obtained using a detection object as
a target. An aptamer as a second nucleic acid portion is obtained
using a labeling substance (e.g., enzyme) as a target. The
above-mentioned SELEX method can be employed as a method for
obtaining the aptamers. Then, these two aptamers are caused to bind
to each other. A method for binding these aptamers is not
particularly limited and can be, for example, a method in which a
single-stranded nucleic acid sequence is produced from sequences of
the two aptamers, and a nucleic acid is synthesized based on this
single-stranded nucleic acid sequence. In this case, the secondary
structures of the two aptamers may be predicted using a computer or
the like, and the single-stranded nucleic acid sequence may be
revised, or a sequence may be deleted or added to the
single-stranded nucleic acid sequence.
Embodiment 1-1b
[0068] The present embodiment is an example of a detection
instrument including a detection reagent which contains a nucleic
acid element in the form (2). The nucleic acid element in the form
(2) is characterized in that a second nucleic acid portion itself
can cause an enzyme reaction. FIGS. 7A and 7B schematically shows a
configuration of the nucleic acid element of the present
embodiment. As shown in FIGS. 7A and 7B, this nucleic acid element
46 includes a first nucleic acid portion (binding portion) 42 and a
second nucleic acid portion (labeling portion) 43, and the first
nucleic acid portion 42 and the second nucleic acid portion 43 are
integrated by linking to each other. As shown in FIG. 7A, when a
detection object 31 does not bind to the first nucleic acid portion
42, the second nucleic acid portion 43 is associated with the first
nucleic acid portion 42, resulting in the state of inhibiting an
enzyme reaction. Then, as shown in FIG. 7B, when the detection
object is present in a sample, and the detection object 31 binds to
the first nucleic acid portion 42, the second nucleic acid portion
43 is released from the first nucleic acid portion 42, and thus,
the second nucleic acid portion 43 can cause an enzyme reaction. At
that time, when a substrate is present, an enzyme reaction occurs.
Thus, the detection object 31 can be detected by detecting this
enzyme reaction as in the case of Embodiment 1-1a.
[0069] In the first nucleic acid portion and the second nucleic
acid portion, the length of nucleic acid is not particularly
limited. The length of nucleic acid in each of the first nucleic
acid portion and the second nucleic acid portion is, for example,
the same as that in Embodiment 1-1a. The length of nucleic acid in
the entire nucleic acid element is not particularly limited, and
the range of the length is, for example, from 14 to 240 bases,
preferably from 14 to 200 bases, more preferably from 14 to 160
bases, yet more preferably from 14 to 140 bases, particularly
preferably from 14 to 75 bases, more particularly preferably from
14 to 55 bases.
[0070] The type of the enzyme reaction of the second nucleic acid
portion 43 is not particularly limited and can be, for example, the
same enzyme reaction as in Embodiment 1-1a. A nucleic acid which
can cause an enzyme reaction can be DNA having Hemin peroxidase
activity (Tao et al, Anal. chem. 2009, 81, 2144-2149). The
substrate also is not particularly limited and can be, for example,
the same substrate as in Embodiment 1-1a.
[0071] Next, FIGS. 8A to 8D show an example of a nucleic acid
element in which an aptamer is employed as a first nucleic acid
portion 42, and DNA having peroxidase activity is employed as a
second nucleic acid portion 43. FIG. 8A shows DNA having peroxidase
activity (SEQ ID NO: 1), and FIG. 8B shows an aptamer to adenosine
(SEQ ID NO: 2). FIG. 8C shows a nucleic acid element of a
single-stranded nucleic acid obtained by binding the DNA (A) and
the aptamer (B) (SEQ ID NO: 3). In the nucleic acid element of FIG.
8C, the aptamer (B) does not bind to adenosine, so that the DNA has
a secondary structure which cannot cause an enzyme reaction. Then,
when the adenosine binds to the aptamer (B) in the nucleic acid
element, the secondary structure of the DNA of FIG. 8C changes to
the one which can cause an enzyme reaction as shown in FIG. 8D. At
that time, when a substrate is present, an enzyme reaction occurs.
Thus, the detection object can be detected by detecting this enzyme
reaction. This nucleic acid element can be used in measurement of
viable cells in food. That is, the presence of viable cells in food
means the presence of ATP, and the concentration of ATP is
proportional to the number of viable cells. Therefore, when this
nucleic acid element, a chromogenic substrate, and food are caused
to react, color development in proportion to the concentration of
ATP derived from viable cells can be obtained. Thus, the number of
viable cells in food can be detected by detecting this color
development.
[0072] This nucleic acid element can be produced as follows, for
example. First, an aptamer as a first nucleic acid portion is
obtained using a detection object as a target. The above-mentioned
SELEX method can be employed as a method for obtaining the aptamer.
On the other hand, a sequence of a second nucleic acid portion is
designed according to an enzyme reaction and synthesized. Then the
aptamer as the first nucleic acid portion and the second nucleic
acid portion are caused to bind to each other. A method for binding
them is not particularly limited and can be, for example, a method
in which a single-stranded nucleic acid sequence is produced from a
sequence of the aptamer as the first nucleic acid portion and a
sequence of the second nucleic acid portion, and a nucleic acid is
synthesized based on this single-stranded nucleic acid sequence. In
this case, the secondary structures may be predicted using a
computer or the like, and the single-stranded nucleic acid sequence
may be revised, or a sequence may be deleted or added to the
single-stranded nucleic acid sequence.
Embodiment 1-2)
[0073] The present embodiment is an example of a detection
instrument in which antibodies for capturing a detection object
(antigen) are immobilized, as a detection reagent, on the detection
portion in the case where the detection object in a sample is a
substance which can be an antigen. The antibodies immobilized on
the detection portion are not particularly limited as long as they
bind to the detection object (antigen) in the sample, and can be
any of immunoglobulin (Ig)G, IgA, IgM, IgE, and IgD. These
antibodies may be polyclonal antibodies or monoclonal antibodies.
These antibodies can be produced by a normal method using an animal
such as a mouse, a rat, a goat, or a chicken.
[0074] FIG. 9 shows an example of immobilizing antibodies on the
detection portion. As shown in FIG. 9, antibodies 61 are
immobilized on a detection portion 52. FIG. 9 schematically shows
the example, so that the size and the number of antibodies 61 are
different from the actual size and the number. In FIG. 9, the
numeral 11 indicates a main body of a detection instrument as shown
in FIGS. 1A and 1B.
[0075] Next, a method for using the detection instrument of the
present embodiment is described with reference to figures. The
following method is a mere example, and the present invention is
not limited to this.
[0076] First, a sample containing antibodies which have been
labeled with an enzyme (hereinafter referred to as an
"enzyme-labeled antibodies") and a chromogenic substrate
corresponding to the enzyme of the enzyme-labeled antibodies are
brought into contact with a detection portion 52.
[0077] The enzyme-labeled antibodies are not particularly limited
as long as they bind to a detection object (antigen) in the sample
and may be any of immunoglobulin (Ig)G, IgA, IgM, IgE, and IgD.
These antibodies may be polyclonal antibodies or monoclonal
antibodies. These antibodies can be produced by a normal method
using an animal such as a mouse, a rat, a goat, or a chicken. The
enzyme of the enzyme-labeled antibodies and the chromogenic
substrate are not particularly limited and can be, for example, the
same enzyme and the same chromogenic substrate as in Embodiment
1-1a.
[0078] When the sample contains the detection object (antigen), the
detection object (antigen) and the enzyme-labeled antibodies are
bound to each other by an antigen-antibody reaction.
[0079] FIG. 10 schematically shows an antigen-antibody reaction in
the present embodiment. In FIG. 10, the identical parts to those in
FIG. 9 are denoted by identical reference numerals. As shown in
FIG. 10, the detection object (antigen) 71 in a sample binds to
antibodies 61 immobilized on the detection portion 52 as well as
the enzyme-labeled antibodies 64, thus forming a complex. In FIG.
10, the numeral 62 indicates antibodies of the enzyme-labeled
antibodies 64, and the numeral 63 indicates an enzyme of the
enzyme-labeled antibodies 64. FIG. 10 schematically shows the
antigen-antibody reaction, the sizes of the antibodies and a label
and the like are different from the actual sizes and the like.
[0080] The detection object can be detected by detecting color
development caused by a reaction of the enzyme of the
enzyme-labeled antibodies and the chromogenic substrate in the
detection portion 52 after predetermined time (not particularly
limited and is, for example, from 10 to 20 minutes).
[0081] In the detection instrument of the present embodiment, a
detection object in a sample may be an antibody, and an antigen for
capturing the detection object (antibody) may be immobilized, as a
detection reagent, on the detection portion. In this case, a sample
containing antigen which has been labeled with an enzyme
(hereinafter referred to as an "enzyme-labeled antigen") and a
chromogenic substrate corresponding to the enzyme of the
enzyme-labeled antigen is brought into contact with the detection
portion 52.
[0082] In order to improve reliability of detection, it is
preferred that a positive control which can be read optically is
placed in the detection portion of the detection instrument
according to the present invention.
Embodiment 2
[0083] FIG. 11 shows an example of a configuration of a detection
system according to the present invention. In FIG. 11, the
identical parts to those in FIGS. 1A to 3D are denoted by identical
reference numerals. As shown in FIG. 11, this detection system
includes the detection instrument according to the present
invention shown in FIG. 1A and an optical reader (bar-code reader)
21. The optical reader (bar-code reader) 21 is connected to a
personal computer (PC) 22. The PC 22 is connectable to a server or
a storage medium on a cloud system (hereinafter collectively
referred to as a "server") 23 via a communication network or a
cloud system (hereinafter collectively referred to as
"communication network") 24. Examples of the communication network
24 include the Internet, and LAN (Local Area Network). The PC and
the server are optional components, and the detection system
according to the present invention is not required to have
them.
[0084] In the present invention, the optical reader is not limited
to a bar-code reader as long as it can read color development of
the detection reagent in the detection instrument according to the
present invention and can be, for example, a camera. For example, a
camera integrated in a mobile phone can be used as the optical
reader.
[0085] The method for using the detection system shown in FIG. 11
is as follows, for example. The following method, however, is a
mere example, and the present invention is not limited by this.
[0086] First, a sample is brought into contact with a detection
portion (bar code) 12. At that time, when a detection object is
present in the sample, the detection reagent specifically reacts
with the detection object and develops a color.
[0087] Then, the color development of the detection reagent in the
detection instrument is read by an optical reader (bar-code reader)
21. This read result is displayed on the PC 22. That is, when color
development of the detection reagent is read, a read result showing
the presence of the detection object in the sample is displayed on
the PC 22. On the other hand, when color development of the
detection reagent is not read, a read result showing the absence of
the detection object in the sample is displayed on the PC 22.
[0088] The read result may be stored in the server 23 via the
communication network 24, and a next detection may be performed
with reference to the read result. For example, in the case of
using the detection system of the present embodiment in the
production field of fermented food, when information on the
presence or absence of color development and the extent of color
development (color intensity) in the stages of preparation, the
middle of fermentation, the end of fermentation is stored in the
server 23, production of fermented food can be managed using the
detection system of the present embodiment.
Embodiment 3
[0089] FIG. 12 shows another example of a configuration of the
detection system according to the present invention. In FIG. 12,
the identical parts to those in FIGS. 1A to 3D and 11 are denoted
by identical reference numerals. As shown in FIG. 12, this
detection system is the same as the detection system of Embodiment
2 shown in FIG. 11 except for including a printer (e.g., an ink jet
recording device or a laser printer) 25 connected to a PC 22.
[0090] A method for using the detection system shown in FIG. 12 is
as follows, for example. The following method, however, is a mere
example, and the present invention is not limited by this.
[0091] First, a detection portion (bar code) 12 is formed in a main
body 11 of a detection instrument with the printer 25. A specific
method for forming the detection portion (bar code) 12 is the same
as described in Embodiment 1. In the detection system of the
present embodiment, when information has been already stored in the
server 23, the detection portion (bar code) 12 may be formed with
reference to the information stored in the server 23. Specifically,
for example, in the case of using the detection system of the
present embodiment in restaurants, hospitals, and the like, if
information on seasonal increase and decrease of the number of
microorganisms is stored in the server 23, the kind of the
detection reagent may be changed according to the information in
order to set microorganisms increased in the season of the
detection as the detection object. Furthermore, for example, in the
case where the detection system of the present embodiment is used
in inspection of imported food, if information on a residual
pesticide which has been detected from imported food in the past is
stored in the server 23, the kind of the detection reagent may be
selected according to the information in order to set the residual
pesticide as the detection objects. The detection portion may be
formed with reference to information on the Internet or a cloud
system as substitute for the information stored in the server 23,
for example.
[0092] Subsequently the detection object can be detected in the
same manner as in Embodiment 2.
Embodiment 4
[0093] The present embodiment is an example of a detection
instrument configured so that a detection portion is formed in a
main body. FIG. 13A shows a configuration of the detection
instrument of the present embodiment. As shown in FIG. 13A, as the
main body 11, a transparent and colorless plastic container is
used. Moreover, a bar code is formed in the detection portion 12,
and eight detection reagents 1a to 8a are placed as parts of the
bar code. As the eight detection reagents 1a to 8a, reagents which
develop a color by contact with the respective eight kinds of
bacteria and virus are used. The detection reagents 1a to 8a are
placed so as to be adjacent to the respective bars 1b to 8b in the
detection portion (bar code) 12. Whether or not the eight kinds of
bacteria and virus are present in the sample can be detected by
reading whether or not the thicknesses of the bars 1b to 8b are
increased after the contact.
Embodiment 5
[0094] The present embodiment is an example of a detection
instrument configured so that a detection reagent contains a
nucleic acid element. As the nucleic acid element, a nucleic acid
element containing a first nucleic acid portion and a second
nucleic acid portion which is DNA (DNA enzyme) which can cause an
enzyme reaction is used. FIG. 13B shows a configuration of the
detection instrument of the present embodiment. As shown in FIG.
13B, the detection reagent (nucleic acid elements) containing a
chromogenic substrate which develops a color by an enzyme reaction
with the second nucleic acid portion is placed at 21 positions of a
detection portion 12. The presence or absence of a detection object
which releases the enzyme reaction from being inhibited by binding
to the first nucleic acid portion in the sample can be detected
from the presence or absence of color development of the detection
reagent (nucleic acid element) at 21 positions caused by the
contact of the sample with the detection portion 12.
Embodiment 6
[0095] The present embodiment is an example of an immune
chromatogram detection instrument configured so that a detection
portion is formed in a main body thereof, and antibodies for
capturing a detection object (antigens) are immobilized on the
detection portion. FIG. 13C shows a configuration of a detection
instrument of the present embodiment. As shown in FIG. 13C, a plate
for development exerting capillary action is used as a main body
11. Moreover, a bar code is formed in the detection portion 12, and
as parts of the bar code, four detection reagents 1a to 4a are
placed in the same. As the four detection reagents 1a to 4a, the
respective antibodies corresponding to the respective four kinds of
bacteria and virus are used. The detection reagents 1a to 4a are
placed so as to be adjacent to the respective bars 1b to 4b in the
detection portion (bar code) 12. A part of the bar code 12 in which
the detection reagents 1a to 4a are not placed includes information
9a on the detection reagents such as detection reagent numbers, and
information 9b on a site at which detection is performed. Moreover,
a sample supply portion 81 is formed on one side of the detection
portion (bar code) 12 in the main body 11, and an antibody portion
82 for immobilization is formed on the other side of the same.
Antibodies corresponding to any of the four kinds of bacteria and
virus are immobilized on the antibody portion 82 for
immobilization. A sample containing enzyme-labeled antibodies and a
chromogenic substance corresponding to an enzyme of the
enzyme-labeled antibodies are brought into contact with the sample
supply portion 81, which is then stood still until color
development of the antibody portion 82 for immobilization is
detected. Thereafter, whether or not the thicknesses of the bars 1b
to 4b are increased after the contact with the sample is read by a
bar-code reader. Thus, the presence or absence of the four kinds of
bacteria and virus in the sample can be detected with reference to
information on the detection reagents such as detection reagent
numbers and information on a site at which detection is
performed.
INDUSTRIAL APPLICABILITY
[0096] The present invention provides a detection instrument and a
detection system, by which everyone can easily detect an intended
detection object without any skilled technique. The uses of the
detection instrument and the detection system are not limited, and
they are applicable in various fields such as food, medical cares,
agriculture, and environment.
EXPLANATION OF REFERENCE NUMERALS
[0097] 1 detection instrument [0098] 1a, 2a, 3a, 4a, 5a, 6a, 7a, 8a
detection reagent [0099] 11 main body [0100] 12, 52 detection
portion (bar code) [0101] 21 optical reader (bar-code reader)
[0102] 22 PC [0103] 23 server [0104] 24 communication network
[0105] 25 printer [0106] 31, 71 detection object [0107] 32, 42
first nucleic acid portion [0108] 33, 43 second nucleic acid
portion [0109] 34 labeling substance [0110] 36, 46 nucleic acid
element [0111] 61, 62 antibody [0112] 63 enzyme [0113] 64
enzyme-labeled antibody [0114] 81 sample supply portion [0115] 82
antibody portion for immobilization
Sequence CWU 1
1
3117DNAArtificialDNA with Peroxidase Activity 1gggtagggcg ggttggg
17233DNAArtificialDNA with Affinity to Adenosine 2ccttacgggg
ggagtattgc ggaggacgta agg 33350DNAArtificialExample of Nucleic Acid
3gggtagggcg ggttgggcct tacgggggga gtattgcgga ggacgtaagg 50
* * * * *