KR101724617B1 - Diagnostic Kit for Detecting Cancer including Discoloration Colormatrix - Google Patents

Diagnostic Kit for Detecting Cancer including Discoloration Colormatrix Download PDF

Info

Publication number
KR101724617B1
KR101724617B1 KR1020160078660A KR20160078660A KR101724617B1 KR 101724617 B1 KR101724617 B1 KR 101724617B1 KR 1020160078660 A KR1020160078660 A KR 1020160078660A KR 20160078660 A KR20160078660 A KR 20160078660A KR 101724617 B1 KR101724617 B1 KR 101724617B1
Authority
KR
South Korea
Prior art keywords
sample
color
leu
specimen
chamber
Prior art date
Application number
KR1020160078660A
Other languages
Korean (ko)
Inventor
신동진
Original Assignee
(주)큐브바이오
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by (주)큐브바이오 filed Critical (주)큐브바이오
Priority to KR1020160078660A priority Critical patent/KR101724617B1/en
Application granted granted Critical
Publication of KR101724617B1 publication Critical patent/KR101724617B1/en

Links

Images

Classifications

    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/26—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving oxidoreductase
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
    • C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
    • C12Q1/533—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving isomerase
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y114/00—Oxidoreductases acting on paired donors, with incorporation or reduction of molecular oxygen (1.14)
    • C12Y114/18—Oxidoreductases acting on paired donors, with incorporation or reduction of molecular oxygen (1.14) with another compound as one donor, and incorporation of one atom of oxygen (1.14.18)
    • C12Y114/18001—Tyrosinase (1.14.18.1)
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12Y—ENZYMES
    • C12Y503/00—Intramolecular oxidoreductases (5.3)
    • C12Y503/02—Intramolecular oxidoreductases (5.3) interconverting keto- and enol-groups (5.3.2)
    • C12Y503/02001—Phenylpyruvate tautomerase (5.3.2.1)
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
    • G01N21/17—Systems in which incident light is modified in accordance with the properties of the material investigated
    • G01N21/25—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands
    • G01N21/29—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands using visual detection
    • G01N21/293—Colour; Spectral properties, i.e. comparison of effect of material on the light at two or more different wavelengths or wavelength bands using visual detection with colour charts, graduated scales or turrets
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/52—Use of compounds or compositions for colorimetric, spectrophotometric or fluorometric investigation, e.g. use of reagent paper and including single- and multilayer analytical elements
    • G—PHYSICS
    • G01—MEASURING; TESTING
    • G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
    • G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
    • G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
    • G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
    • G01N33/68—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving proteins, peptides or amino acids
    • G01N33/6803—General methods of protein analysis not limited to specific proteins or families of proteins
    • G01N33/6806—Determination of free amino acids
    • G01N33/6812—Assays for specific amino acids
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B5/00—Optical elements other than lenses
    • G02B5/30—Polarising elements
    • G02B5/3025—Polarisers, i.e. arrangements capable of producing a definite output polarisation state from an unpolarised input state
    • G02B5/3033—Polarisers, i.e. arrangements capable of producing a definite output polarisation state from an unpolarised input state in the form of a thin sheet or foil, e.g. Polaroid
    • G02B5/3041—Polarisers, i.e. arrangements capable of producing a definite output polarisation state from an unpolarised input state in the form of a thin sheet or foil, e.g. Polaroid comprising multiple thin layers, e.g. multilayer stacks
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B6/00—Light guides; Structural details of arrangements comprising light guides and other optical elements, e.g. couplings
    • G02B6/0001—Light guides; Structural details of arrangements comprising light guides and other optical elements, e.g. couplings specially adapted for lighting devices or systems
    • G02B6/0011—Light guides; Structural details of arrangements comprising light guides and other optical elements, e.g. couplings specially adapted for lighting devices or systems the light guides being planar or of plate-like form
    • G02B6/0013—Means for improving the coupling-in of light from the light source into the light guide
    • G02B6/0015—Means for improving the coupling-in of light from the light source into the light guide provided on the surface of the light guide or in the bulk of it
    • G02B6/0016—Grooves, prisms, gratings, scattering particles or rough surfaces
    • G—PHYSICS
    • G02—OPTICS
    • G02B—OPTICAL ELEMENTS, SYSTEMS OR APPARATUS
    • G02B6/00—Light guides; Structural details of arrangements comprising light guides and other optical elements, e.g. couplings
    • G02B6/0001—Light guides; Structural details of arrangements comprising light guides and other optical elements, e.g. couplings specially adapted for lighting devices or systems
    • G02B6/0011—Light guides; Structural details of arrangements comprising light guides and other optical elements, e.g. couplings specially adapted for lighting devices or systems the light guides being planar or of plate-like form
    • G02B6/0013—Means for improving the coupling-in of light from the light source into the light guide
    • G02B6/0023—Means for improving the coupling-in of light from the light source into the light guide provided by one optical element, or plurality thereof, placed between the light guide and the light source, or around the light source
    • G02B6/0031—Reflecting element, sheet or layer

Landscapes

  • Life Sciences & Earth Sciences (AREA)
  • Health & Medical Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Physics & Mathematics (AREA)
  • Engineering & Computer Science (AREA)
  • Organic Chemistry (AREA)
  • Immunology (AREA)
  • Molecular Biology (AREA)
  • General Health & Medical Sciences (AREA)
  • Biochemistry (AREA)
  • General Physics & Mathematics (AREA)
  • Wood Science & Technology (AREA)
  • Zoology (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Hematology (AREA)
  • Analytical Chemistry (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Biotechnology (AREA)
  • Urology & Nephrology (AREA)
  • Genetics & Genomics (AREA)
  • Microbiology (AREA)
  • Biomedical Technology (AREA)
  • General Engineering & Computer Science (AREA)
  • Optics & Photonics (AREA)
  • Pathology (AREA)
  • Biophysics (AREA)
  • Cell Biology (AREA)
  • Food Science & Technology (AREA)
  • Medicinal Chemistry (AREA)
  • Bioinformatics & Computational Biology (AREA)
  • Spectroscopy & Molecular Physics (AREA)
  • Investigating Or Analysing Materials By The Use Of Chemical Reactions (AREA)

Abstract

The present invention relates to a cancer diagnostic kit allowing a discoloration degree of a discolored sample to be precisely understood as a color balance sheet used for determining a color change degree of the sample is changed by corresponding to the sample color before discoloration. The cancer diagnostic kit comprises: a housing having a sample chamber and a comparison chamber in which an input sample is separated and accommodated and having a window capable of checking the inside of the sample chamber and the comparison chamber on one side with naked eyes; and an enzyme composition accommodated in the sample chamber, reacting with tyrosine included in the input sample and discoloring the sample. The color balance sheet which becomes opaque from one side is formed on a part corresponding to the comparison chamber of the window.

Description

{Diagnostic Kit for Detecting Cancer including Discoloration Colormatrix}

The present invention relates to a kit for diagnosing the presence or absence of cancer, and more particularly, to a kit for diagnosing cancer of a subject through color change of a specimen, The present invention relates to a cancer diagnostic kit capable of precisely grasping the degree of discoloration of a discolored specimen because the specimen is variable in response to a color.

Cancer markers are substances that occur in the malignant tumor cells themselves or are caused by reactions of normal tissues against cancer and exhibit abnormally high concentrations in the blood, urine or tissues, and the cancer marker concentration in the blood, urine or tissue You can diagnose, screen, and track progress and degeneration.

In general, the measurement or detection of the concentration of the cancer marker is carried out through the blood, but the blood has a problem that it is difficult for the general person to collect it.

There has been developed a method for diagnosing cancer by detecting the concentration of cancer biomarker present in the urine by using urine which is not easy to obtain and which is not a biomaterial such as blood or tissue.

On the other hand, it is known that the tyrosine can be used as the cancer biomarker because the concentration of tyrosine present in the urine is high in cancer patients as compared with the general public.

Korean Patent No. 10-0033547 relates to a mixed reagent for diagnosing cancer by detecting aromatic amine (Tyrosine) commonly present in urine of a cancer patient, wherein the mixed reagent is composed of mercury, nickel, nitric acid and distilled water .

However, the above-mentioned mixed reagent contains mercury and is currently prohibited because it has a great influence on the environment and human body.

Therefore, there is an urgent need to develop a composition for diagnosing cancer by eco-friendly detection of the concentration of cancer biomarker tyrosine (Tyrosine) present in the urine by using urine which is not easy to obtain but is not easy to obtain.

In addition, kits for diagnosing diseases by color change of a conventional sample have been judged by comparing the color of the pre-printed color chart with the color of the discolored specimen. For example, FIG. 1 is a color chart prepared to diagnose cancer by color change of urine. The colors arranged in the horizontal direction are intended to diagnose cancer to the extent that urine is discolored, but the colors listed in the vertical direction are merely urine According to the subject, even for the same subject, since the color of the urine differs depending on the period of urine collection, various control groups are required.

As described above, there is a difficulty in providing a plurality of color tables as shown in FIG. 1 because the color of the sample is fixed by any one of urine, blood or the like.

Also, since the color table composed of a plurality of colors is physically large, there is a problem that it is not possible to manufacture a color table in a human body in a small cancer diagnosis kit.

In addition, the color chart for diagnosing the disease by discoloration of the specimen inherently does not take into consideration the color of the specimen, thereby posing a risk of misdiagnosis. For example, if the color of the urine is muddy due to other health problems of the subject, even if the color change of the urine is insignificant by the cancer diagnosis composition, in the absolute reference color table, the urine is a color suspected of being a disease I was misinformed. In other words, there is a problem that the reliability of the diagnostic result is not high when comparing the absolute reference color table with the discolored specimen.

Korean Patent No. 10-0033547

The present invention solves the problem of such prior art diagnostic kits and includes the enzymes that are characterized in converting L-Tyrosine into Eumelanin, so that the concentration of tyrosine present in the urine The present invention provides a cancer diagnostic kit comprising an enzyme composition for detecting cancer biomarkers capable of detecting the presence or absence of cancer and metabolic diseases.

In addition, in diagnosing cancer of a subject through color change of a specimen, the present invention changes the degree of discoloration of a discolored specimen such that the color chart of the specimen changes in accordance with the color of the specimen before discoloration A cancer diagnosis kit, and a cancer diagnosis kit.

In addition, the present invention provides a cancer diagnosis kit that can easily diagnose the presence of cancer and metabolic diseases in the home without inconveniencing users to visit a specialized institution such as a hospital by using urine which can be collected easily, The purpose is to provide.

In addition, the present invention provides a self-light source in the diagnostic kit so that the degree of color change due to the reaction can be accurately determined by using the light of the self light source irradiated from the rear side of the reaction area without depending on external natural light or external light A cancer diagnostic kit is provided.

It is another object of the present invention to provide a cancer diagnostic kit having an optical film layer for reducing a contrast ratio by reflection of external light and preventing an external shadow from projecting on a window surface.

Further, the present invention aims to provide a cancer diagnostic kit which can be immediately used for diagnosis using not only urine but also other samples such as blood because the color chart is changed to match the color of the specimen.

The objects of the present invention are not limited to the above-mentioned objects, and other objects not mentioned can be clearly understood by those skilled in the art from the following description.

In order to attain the above object, a cancer diagnosis kit including a color chart that is discolored corresponding to a sample color according to the present invention includes a sample chamber and a control chamber in which a sample is separated and accommodated, A housing having a window through which the inside of the chamber can be visually confirmed; And an enzyme composition which is accommodated in the sample chamber and reacts with tyrosine contained in the sample to change the color of the specimen, and a color matching table which gradually becomes opaque from one side is formed at a portion corresponding to the control chamber of the window .

The specimen may be one of body fluids derived from a human body, and the body fluids may include blood, urine, serum, plasma, lymph fluid, tissue fluid, and secretion.

In addition, the enzyme composition may further comprise tyrosine (L-Tyrosine), tyrosinase (DCT) and tyrosinase-related protein 1 (Tyrp1) Is converted into melanin. The present invention also provides an enzyme composition for detecting tyrosine.

The enzymatic composition may be prepared by adding 0.5 to 2 parts by weight of a dopa chromium mutant enzyme (DCT) to 1 part by weight of the tyrosinase and tyrosinase-related protein 1 (Tyrp1) to 40 parts by weight of tyrosinase, 0.5 to 2 by weight, wherein L-tyrosine is converted into Eumelanin, and the color of the specimen is changed from its original color to black as the concentration of tyrosine is increased .

The enzyme composition may further comprise phosphate buffer or Tris-HCl buffer solution having a final concentration of 50 to 100 mM, Tween 20 having a final concentration of 0.1 to 0.5%, and distilled water , And a final reaction pH of 7.0 to 8.5, and is contained in the sample chamber.

The apparatus may further include a light guide plate unit installed on an inner wall surface of the housing opposite to the window to irradiate light to the sample chamber and the control chamber.

The light guide plate unit may further include a light guide plate having an incident surface on which the light emitted from the light source is incident and an exit surface on which the light propagated through the incident surface is deflected, A diffusing sheet for adjusting the diffusion and propagation direction of light passing through the light guide plate; a prism sheet for refracting and diffusing light emitted from the diffusion sheet to increase the brightness; And a reflective sheet for reflecting light is stacked.

In addition, the apparatus includes a housing provided at one side of the housing for introducing the specimen, and an inlet for introducing the introduced specimen into the specimen chamber and the control chamber. The inlet is provided at an upper portion of the housing to prevent leakage of urine And a sample accommodating portion for allowing the sample chamber and the control chamber to separate and flow into the sample chamber and the control chamber in a state in which the inserted sample is held, wherein the sample chamber and the air in the sample chamber, And a discharge path is formed.

The cancer diagnosis kit including the color charts corresponding to the color of the sample according to the present invention has the following effects.

First, it is possible to provide a cancer diagnostic kit using an enzyme composition for detecting cancer biomarkers that can detect the presence of tyrosine present in urine in an environmentally friendly manner to detect the presence of cancer and metabolic diseases.

Secondly, the user can easily diagnose the presence of cancer and metabolic diseases in the home without the inconvenience of visiting the specialist institution such as the hospital and receiving the cancer diagnosis by using the urine which can be collected easily.

Third, as the color chart is changed in accordance with the color of the sample before discoloration, the discoloration degree of the discolored specimen can be grasped accurately.

Fourth, since the self-light source is provided in the diagnostic kit, the degree of color change due to the reaction can be accurately determined by using the light of the self-light source irradiated from the rear side of the reaction area without depending on external natural light or illumination.

Fifth, the present invention can be used immediately for diagnosis using not only urine but also other samples such as blood because the color chart is variable to match the color of the specimen.

1 is a color chart for measuring the color of the produced eumelanin in a white light environment
FIG. 2 is a perspective view illustrating a cancer diagnostic kit including a color chart that is discolored corresponding to a sample color according to an embodiment of the present invention. FIG.
FIG. 3 is a cross-sectional view of a cancer diagnosis kit including a color chart which is discolored corresponding to a sample color according to an embodiment of the present invention,
FIG. 4A is a front view showing a state before a specimen is injected into a cancer diagnostic kit including a color chart which is discolored corresponding to a specimen color according to an embodiment of the present invention. FIG.
FIG. 4B is a front view showing a state in which a specimen is put in a cancer diagnosis kit including a color chart that is discolored corresponding to a specimen color according to an embodiment of the present invention,
FIG. 4c is a front view showing a state in which a specimen is put in a cancer diagnosis kit including a color chart that is discolored corresponding to a specimen color according to an embodiment of the present invention, and a specimen in the specimen chamber is discolored
5 is a schematic view showing an example of a light guide plate unit according to an embodiment of the present invention.
6A to 6D are diagrams showing an example of an optical film according to an embodiment of the present invention
7 is a photograph comparing the colors of melanin produced in a white light environment

Hereinafter, a preferred embodiment of a cancer diagnostic kit including a color chart which is discolored corresponding to the sample color according to the present invention will be described in detail.

The features and advantages of a cancer diagnostic kit including a color chart that is discolored corresponding to the sample color according to the present invention will be apparent from the following detailed description of each embodiment.

Referring to FIGS. 2 and 3, in order to solve the conventional problem, the cancer diagnostic kit including the color chart 124 that discolors according to the color of a specimen according to the present invention includes a sample chamber 130 A sample chamber 130 having a sample chamber 130 and a control chamber 140 formed on one side thereof and a window 123 through which a sample chamber 130 and a control chamber 140 can be visually confirmed, And an enzymatic composition 132 which accepts and reacts with tyrosine contained in the sample to discolor the specimen and a color chart 124 which gradually becomes opaque from one side is formed at a portion corresponding to the control chamber 140 of the window 123 Is formed.

In the following description, the enzyme composition 132 used as a reaction material is exemplified by the enzyme composition 132 for tyrosine detection and urine is exemplified as the sample used. However, the kind of the enzyme composition 132 and the type of the sample It is natural that it can be different.

For example, the specimen applicable to the present invention is any one of body fluids derived from a human body, and the body fluids may include blood, urine, serum, plasma, lymph fluid, tissue fluid, secretion and other body fluids, Lt; / RTI > group.

In particular, the enzyme composition 132 reacts with tyrosine contained in the specimen to discolor the specimen. The greater the content of tyrosine contained in the specimen, the greater the degree of discoloration of the specimen. However, even if the kinds of body fluids derived from the human body are the same, the color differs depending on the person, depending on the collected time, depending on the collected method. Therefore, there is a problem that it is difficult to accurately determine the discoloration degree of the specimen unless the original color of the specimen and the enzyme composition 132 react with each other to directly compare the discolored specimen. Since the sample chamber 130 in which the sample reacts with the enzyme composition 132 and the control chamber 140 in which the original sample is contained are formed in the housing 100 according to the embodiment of the present invention, 130 can be clearly determined by comparing the degree of discoloration of the specimen with the color of the specimen of the control chamber 140.

As shown in the figure, the sample chamber 130 and the control chamber 140 are preferably arranged side by side so that the colors of the specimens contained in each chamber can be easily compared.

The color matching table 124 is used to determine the possibility of cancer of the subject by the degree of discoloration of the specimen, and it is possible to compare how much the specimen has discolored in the original color and consequently to deduce the specific gravity or content of the tyrosine contained in the specimen It is an index that enables the examinee to judge the likelihood of cancer.

Since the color table 124 is formed in the form of a gradient from a transparent color to an opaque color on a portion of the transparent window 123 where the inside of the control chamber 140 can be identified, when the sample enters the control chamber 140 The overall color is visually changed corresponding to the original color of the specimen.

Particularly, when the color matching sheet 124 is formed to become gradually opaque including the transparent region, it becomes easier to compare the original color of the specimen with the discolored specimen through the transparent region.

 The color look-up table 124 may be printed directly on the window 123, or the printed color sheet 124 may be attached to the window 123, or the like.

The inlet 110, which is provided at one side of the housing 100 and into which the sample is introduced, introduces the introduced sample into the sample chamber 130 and the control chamber 140.

Specifically, the inlet 110 is provided at an upper portion of the inlet 110 to prevent leakage of the introduced urine to the outside, a blocking layer 111 made of an elastic material, a sample chamber 130, And a sample accommodating portion 112 for separating the sample into the control chamber 140.

As shown in the figure, when a specimen is received in a tool such as the eyedropper 200 and then inserted into the shielding film 111 to insert the specimen, the specimen is temporarily stored in the specimen accommodating portion 112 formed to become narrower toward the lower side . A sample inlet 113 communicating with the sample chamber 130 and a control inlet 114 communicating with the control chamber 140 are formed in a lower portion of the sample receiver 112 formed to be narrowed toward the bottom, 112 are separately introduced into the sample chamber 130 and the control chamber 140, respectively.

The sample inlet 130 and the control chamber 140 may have a structure in which the sample inlet 134 and the control inlet 144 are connected to the sample receiver 112 so that the sample can be introduced into the sample chamber 130 and the control chamber 140 .

The air that has filled the conventional sample chamber 130 and the control chamber 140 is discharged to the sample inlet 113 and the control inlet 114 as the sample is injected through the inlet 110 and the sample may not flow smoothly . In order to prevent such a problem, an air discharge path 116 is formed in the inlet 110 to allow the sample chamber 130 and the inside of the control chamber 140 to be discharged to the sample storage unit 112.

As shown in the figure, the air discharge path 116 is formed on the upper part of the sample storage part 112, not on the lower part of the sample storage part 112 where the specimen is concentrated. In order to prevent the specimen from flowing into the air discharge path 116, And a portion communicating with the sample accommodating portion 112 is formed with a hole downward.

When the specimen flows into the specimen chamber 130 and the control chamber 140 from the specimen accommodating portion 112 by the air discharge path 116, the air filling the existing specimen chamber 130 and the control chamber 140 is discharged So that the sample can flow smoothly.

In addition, since the air discharge path 116 is connected to the sample accommodating portion 112, it is possible to prevent odors or minute liquid droplets that may occur in the specimen from leaking to the outside.

In addition, the present invention can accurately determine the degree of color change due to the reaction using the light of the self-light source irradiated from the backside of the sample chamber 130 and the control chamber 140, without depending on external natural light or external light It may have its own light source inside the diagnostic kit.

The sample chamber 130 and the wall of the control chamber 140 inside the housing 100 opposed to the window 123 area are uniformly irradiated with light through its own light source in order to increase the accuracy of color change through the window 123. [ A light guide plate unit is provided.

A battery 172 for supplying power to the self light source is provided in the housing 100 and a switch 170 for controlling power supply is installed on one side of the housing 100.

The position and shape of the switch 170 are not limited as shown in the drawings, but may vary depending on the shape of the housing 100.

In the embodiment of the present invention, considering that the color of the sample changes due to the color of the enzyme composition 132 itself, the coloring solution 142 of the same color as the enzyme composition 132 is added to the control chamber 140 Lt; / RTI > In the case of the liquid enzyme composition 132, the sample may be discolored by mixing with the sample before the reaction by tyrosine occurs when the sample is introduced. For example, even when the enzyme composition 132 is a colorless transparent liquid, when urine of yellow color or blood of red color is added, the enzyme composition 132 is mixed with the enzyme composition 132 and the color of the original sample becomes less clear. Therefore, when the coloring liquid 142 having the same color as that of the enzyme composition 132 is accommodated in the control chamber 140, the hue of the sample introduced into the control chamber 140 is also detected in the sample chamber 130 ) Becomes the same as the color of the lightened sample.

A method of using the cancer diagnostic kit including the color chart 124 corresponding to the sample color according to an embodiment of the present invention will be described with reference to the drawings.

4A, when the specimen is not supplied, only the enzyme composition 132 is filled in the sample chamber 130, and the control chamber 140 is empty or the same amount of the colorant solution 132 as the enzyme composition 132 142 are filled.

At this time, the color matching table 124 does not have a color corresponding to the specimen.

Referring to FIG. 4B, when a specimen is injected through the inlet 110, the specimen is separated into the specimen chamber 130 and the control chamber 140, and then flows into the specimen chamber 130 and the control chamber 140. The sample introduced into the sample chamber 130 is filled with the sample composition 130 in a state of being mixed with the enzyme composition 132 and the sample introduced into the control chamber 140 is mixed with the small- Thereby filling the chamber 140.

When a specimen is introduced into the control chamber 140, the color table 124, which is composed of a transparent color and gradually becomes opaque, changes color visually corresponding to the color of the specimen.

In particular, the enzyme composition 132 in this example is characterized in that the tyrosine is converted to melanin to change the sample to black. The color control portion gradually changes from transparent color to black in response to the characteristics of the enzyme composition 132 It is preferable to be formed to become opaque.

The color of the sample contained in the sample chamber 130 corresponds to the transparent color of the color matching table 124 because the sample tyrosine starts to react with the enzyme composition 132 immediately after the sample is mixed with the enzyme composition 132 Color.

Referring to FIG. 4C, after a predetermined period of time, the color of the specimen contained in the sample chamber 130 gradually changes due to the reaction of the enzyme composition 132 and tyrosine contained in the specimen.

When the color of the specimen contained in the specimen chamber 130 and the color of the specimen contained in the control chamber 140 are compared with each other after the sufficient time has elapsed to complete the color change of the specimen, So that it can be confirmed clearly. At the same time, by comparing the color of the specimen discolored in the specimen chamber 130 with the color chart 124 that has been discolored corresponding to the original color of the specimen, it is possible to determine the cancer risk of the subject.

The light guide plate unit of this embodiment will be described in more detail. The light guide plate unit has a light incident surface on which the light emitted from the light source 62a is incident, and a light guide plate unit in which the light traveling through the incident surface diffuses, reflects, A diffusing sheet 81 for adjusting the direction of diffusion and progress of the light passing through the light guide plate 80 and a light guide plate 80 for refracting and diffusing the light from the diffusing sheet 81, The light guide plate unit 62 is disposed on the lower side of the light guide plate 80 and has a structure in which the light source 62a and the reflection sheet 83 for reflecting the light beam directly under the light guide plate 80 are laminated. And the structure of the light guide plate unit 62 is not limited thereto.

The light guide plate is preferably made of a transparent material such as plastic or resin such as polycarbonate (PC) or polymethylmethacrylate (PMMA), and can be manufactured by injection molding.

The cancer diagnostic kit including the color table 124, which has discolored color corresponding to the color of the sample according to the present invention having the window 123 structure, does not depend on external natural light or illumination upon color discrimination by reaction after urine injection The degree of color change due to the reaction can be accurately determined by using the light of the self light source irradiated from the rear side of the reaction region.

6A to 6D, on the outer surface of the window 123 formed on one side of the housing 100, a contrast ratio is reduced by reflection of external light, and an external shadow is projected on the surface of the window 123 An optical film layer is provided on the surface of the window 123 to prevent color distortion caused by external light in order to prevent color distortion appearing in the window 123. [

The optical film to be applied to the cancer diagnostic kit including the color chart 124 that is discolored corresponding to the sample color according to the present invention may be an anti-reflection film, an anti-glare film or a polarizing film (polarizer), or may be a multilayer structure in which these layers are laminated.

The structure of the optical film described below is an example of an optical film applied to a cancer diagnostic kit including a color chart 124 in which the color of the sample is discolored corresponding to the color of the sample according to the present invention.

In addition, the kind of the pressure-sensitive adhesive on the surface of the window 123 can be applied to the present invention without limiting the material and the optical film used in the production of the optical film.

6A, the antireflection film 12a is formed by depositing a multilayer thin film 12c made of a material having a different refractive index on a base film 12b such as polyethylene terephthalate (PET) Which is formed on the surface of the film by the method of the present invention so as to have an antireflection effect.

For example, silicon oxide (SiO2) and titanium oxide (TiO2) can be alternately laminated.

6B, the anti-glare film 13a includes an irregular surface 13c on the surface of a base film 13b, such as polyethylene terephthalate (PET), to diffuse external light.

Such irregular surface 13c can be formed by spraying silicon particles.

6C, the linear polarizing film 14a is made of triacetyl-cellulose (TAC) with a polymer polarizing medium 14c such as polyvinyl alcohol (PVA) for polarizing incident light as a center. 14d may be located on both sides of the polymeric polarization medium 14c.

6D, a circular polarizing film 15a is formed on a retardation film 15b for compensating for retardation by using a polymer polarizing medium (for example, polyvinyl alcohol (PVA)) for polarizing incident light A support 15c such as triacetyl-cellulose (TAC) 15e may be positioned on both sides of the polymeric polarizing medium 15d with respect to the support 15d.

The optical film applied to the present invention is adhered to the surface of the window 123, and the diagnosis kit is not judged by the naked eye, and may be desorbed if necessary in the case of using a separate analyzer.

In order to adhere and detach the optical film, it is important to control the peeling force of the adhesive.

In the present invention, the pressure-sensitive adhesive used for controlling the peeling force may be a material comprising an acrylic resin, a polyester resin or a silicone resin composition.

Such a resin composition may include an oligomer, a monomer, a photoinitiator, an adhesion promoter, a leveling agent, and other additives.

The diagnostic mechanism using the cancer diagnostic kit including the color check sheet 124 that has discolored in accordance with the color of the sample according to the present invention having such a structure will be described in detail as follows.

The present invention relates to a pharmaceutical composition comprising tyrosinase, Dopachrome tautomerase (DCT) and tyrosinase-related protein 1 (Tyrp1) for detecting the concentration of the cancer biomarker tyrosine present in the urine An enzyme composition (132) for detecting cancer biomarker tyrosine is used as a diagnostic substance.

More specifically, the enzyme composition for detecting tyrosine 132 comprises an enzyme composition in which tyrosinase is supplemented with a Dopachrome tautomerase (DCT) and a tyrosinase-related protein 1 (Tyrp1) enzyme (132). Cancer can be diagnosed by converting the cancer biomarker tyrosine (L-Tyrosine) present in the urine into melanin and measuring the concentration through the color of the melanin.

In particular, in the present invention, the enzyme composition for detecting tyrosine 132 is used to convert L-tyrosine into melanin, wherein the melanin is eumelanin.

Since the enzyme composition 132 for detecting tyrosine according to an embodiment of the present invention is for diagnosing cancer using urine, it is preferable to use the enzyme composition 132 for detecting tyrosine when the color of urine is considered, such as pheomelanin (brown) or mixed melanin Mixed melanins may be difficult to measure because they are similar to the color of the urine. Accordingly, it is preferable that the enzyme composition 132 for detecting tyrosine according to the present invention is measured for its concentration through black colored eumelanin.

The enzyme composition 132 for detecting tyrosine not only accelerates the pathway of eumelanin biosynthesis but also induces the color of the produced uromelanin to a more black path by adding DCT and Tyrp1 enzyme to tyrosinase have.

Since the concentration of tyrosine present in the urine can be measured through the color of the eumelanin produced by the reaction with the tyrosine detecting enzyme composition 132, when the color of urine produced by the produced uromelanin is close to black, It can be seen that the amount of tyrosine is high.

The tyrosine detection reaction by the tyrosine detection enzyme composition 132 may be performed using a phosphate buffer or a Tris-HCl buffer solution having a final concentration of 50 to 100 mM and a final concentration of 0.1% to 0.5 % Tween 20, and the final reaction pH is 7.0 to 8.5.

For example, if the phosphoric acid buffer solution contained in the tyrosine detecting enzyme composition 132 is 200 mM, and the total volume of 500 μl of the enzyme composition 132 and 500 μl of the urine sample is 1000 μl, 50 mM. ≪ / RTI >

In one embodiment of the present invention, the tyrosine detecting enzyme composition 132 comprises tyrosinase, DCT and Tyrp1, and is diluted with a phosphate buffer or a Tris-HCl buffer solution, Tween 20, And adding distilled water.

Preferably, the enzyme composition for detecting tyrosine 132 comprises 40 units of tyrosinase, 1 part by weight of the tyrosinase, 0.5 to 2 parts by weight of DCT, and 0.5 to 2 parts by weight of Tyrp1.

If the above content is not satisfied, it may be difficult to measure the concentration of tyrosine because the color of the produced melanin may appear to be a yellowish brown color and be mixed with the urine color.

As described above, the enzyme composition for detecting tyrosine 132 accelerates the pathway of eumelanin biosynthesis by tyrosinase by DCT and Tyrp1 enzyme, and induces the color of the resulting eumelanin to a more black-colored pathway .

Referring to Reaction Scheme 1 below, L-tyrosine is oxidized by tyrosinase to form 3,4-dihydroxyphenylalanine (DOPA) and then oxidized by tyrosinase to form DOPA quinone , And Leucodopachrome to form Dopachrome.

The dopachrome is a melanin precursor and is tautomerized by DCT to form 5,6-dihydroxyindole-2-carboxylic acid (DHICA).

The 5,6-dihydroxyindole-2-carboxylic acid is oxidized by Tyrp 1 to form indole-5,6-quinonecarboxylic acid and the indole-5,6-quinonecarboxylic acid is polymerized to form uamelanin .

[Reaction Scheme 1]

Figure 112016060845981-pat00001

The tyrosinase is a tyrosinase hydroxylase and a copper-containing enzyme having catalytic activity of dopa oxidase, which can be found in microorganisms and plant and organism tissues. In particular, the tyrosinase promotes the production of melanin and other pigments by oxidation of phenols such as tyrosine.

The Dopachrome tautomerase (DCT), also known as tyrosinase-related protein 2 (Tyrp 2), is a protein that is characterized by the binding of tyrosinase and Tyrp1 to convert L-tyrosine to melanin in melanocytes It is a melanin cell specific enzyme.

The tyrosinase-related protein 1 (Tyrp1) plays a role in stabilizing tyrosinase.

The enzyme composition for detecting tyrosine 132 according to the present invention is prepared by dissolving 5,6-dihydroxyindole in indole-5,6-quinone (Indole-5 , 6-quinone) to form melanin near the brown color (see the following reaction formula 2), but is tautomerized by DCT and oxidized by Tyrp 1 to form melanin close to black, By separating them from color, it is possible to perform density measurement effectively.

[Reaction Scheme 2]

Figure 112016060845981-pat00002

In one embodiment of the present invention, when measuring the concentration of tyrosine present in the urine through the color of the produced eumelanin, the color of the produced uromelanin is compared with the color chart 124 to confirm the concentration of tyrosine in the urine have. This makes it possible to easily diagnose the occurrence of cancer and metabolic diseases.

Hereinafter, the tyrosine detection enzyme composition 132 of the present invention will be described in more detail by way of examples. It is to be understood by those skilled in the art that these embodiments are for illustrative purpose only and that the scope of the present invention is not limited to these embodiments.

Preparation Example 1: Enzyme composition (132) Preparation of each component

The tyrosinase enzyme was purchased from Sigma.

DCT and Tyrp1 were obtained from mouse blood by reference to the following steps and purified after protein expression in E. coli.

(i) RNA extraction from mouse blood

RiboEx RNA extraction solution kit (Jinol Biotechnology, Korea) was used in this embodiment. The process is as follows.

a. 300 μl of mouse blood and 600 μl of RiboEx RNA extraction solution (Jeolol Biotechnology, Korea) were mixed from the mouse blood, mixed strongly and left at room temperature for 5 ~ 10 minutes to dissolve the cell membrane.

b. 200 μl of chloroform was added and mixed vigorously. The mixture was centrifuged at 13,000 rpm at 4 ° C for 10 minutes using a centrifuge to separate the upper and lower layers.

c. After collecting only the upper layer, the same amount of the column-binding buffer was added and mixed well. Then, the solution was added to the column provided in the kit and spin-down to bind only RNA to the column.

d. The wash buffer provided in the kit was added to the column, and then the material other than RNA was removed by spin-down. The remaining wash buffer was removed by centrifugation for an additional 1 minute.

e. The elution buffer was added to recover the RNA bound to the column, and the RNA was recovered by spinning 5 minutes later.

(ii) preparing cDNA from the obtained RNA

 1 μg RNA, 5 units of reverse transcriptase (reverse transcriptase), oligo dT-50 pmol, 0.1 M DTT- 2 μl, 10x reaction buffer - 2 μl, water - about 20 μl; The mixture was prepared in the above composition and reacted at 45 ° C for 10 minutes and at 38 ° C for 3 hours or longer in a PCR instrument.

(iii) securing the gene (DCT, Tyrp1) from cDNA

a. In order to amplify the gene from the cDNA generated in the above step, the mixture was prepared with the following composition, and the gene amplification reaction (PCR) was performed under the following conditions.

Mixture composition: 2 μl of cDNA solution, 5 units of pfu DNA amplification enzyme, 2 μl of 10 mM dNTP, 4 μl of 10 × reaction buffer, 2 pmol of each primer pair, and water - about 40 μl.

Amplification reaction conditions: 95 ° C 4 min - 1 time; 95 ° C for 20 seconds, 60 ° C for 20 seconds, 72 ° C for 1 minute and 50 seconds - 35 times; 72 占 폚 10 min - 1 time; Continue at 4 ℃.

The information of the primer pair used in the above reaction is shown in Table 1 below.

division Base sequence Primer for DCT amplification Forward primer Atgggccttgtgggatggg (SEQ ID NO: 1) Reverse primer Ctaggcttcctccgtgtatctcttgc (SEQ ID NO: 2) Tyrp1 primer for amplification Forward primer Atgaaatcttacaacgtcctccccct (SEQ ID NO: 3) Reverse primer Tcagaccatggagtggttaggattcg (SEQ ID NO: 4)

The DCT and Tyrp1 enzymes were prepared by inserting the genes obtained by the above method into the pET28a vector and constructing a fusion protein in which 6x His was attached to the N-terminus and transformed into BL21 (DE3) Escherichia coli. The process is as follows.

(i) the step of culturing the transformed E. coli

a. NdeI of the pET28a vector, and DCT or Tyrp1 gene into the XhoI restriction enzyme site, thereby preparing a recombinant DNA which produces 6x His ~ protein.

b. E. coli transformed with BL21 (DE3) was prepared by heat shock transformation.

c. The E. coli strain was cultured in 3 L of Luria broth at 37 DEG C and 200 rpm at OD600 ~ 0.7, followed by treatment with 0.1M IPTG to induce protein production.

d. After further culturing at 30 DEG C for 4 hours, E. coli was precipitated by centrifugation.

(ii) Escherichia coli disruption step

a. The precipitated E. coli was resuspended in 200 ml of 1x PBS and washed by centrifugation.

b. The precipitated E. coli was suspended in 40 ml of 0.5 M NaCl, 5 mM imidazole, 20 mM Tris-HCl, pH 7.9; Lt; / RTI >

c. Escherichia coli was disrupted in an ultrasonic cell disruptor under the condition of Energy 38% max, total cell breaking time 4 min (2 second sonic treatment / 4 sec pause). The bottle containing the E. coli sample remained ice-filled during the cell disruption process.

d. The shredded E. coli was centrifuged at 13,000 rpm, 30 minutes, and 4 ° C.

e. Only the supernatant was collected in a clean conical tube.

(iii) Purification of his tag-fusion protein using Ni-NTA agarose resin

a. The water-soluble protein solution obtained in the above step was mixed with Ni-NTA agarose resin and shaken for 30 minutes at 4 ° C to induce binding of the his tag-fusion protein to the Ni-NTA agarose resin.

b. The protein solution and the resin mixture were put into the column, and the column cock was opened to extract the liquid.

c. 400 ml of washing buffer (0.5 M NaCl, 60 mM imidazole, 20 mM Tris-HCl, pH 7.9) was passed through to remove proteins other than impurities and fusion proteins.

d. 3 ml of elution buffer (0.25 M NaCl, 500 mM imidazole, 20 mM Tris-HCl, pH 7.9) was added and allowed to stand for 10 minutes.

e. d process was repeated twice.

(iii) Protein purification using FPLC-size exclusion method

a. The recovered proteins were loaded onto FPLC equipped with a superdex S200 (GE healthcare) column and the proteins were separated by size.

b. Among the obtained fraction samples, fractions corresponding to his tag fusion proteins were identified by polyacrylamide gel electrophoresis and finally the desired his tag fusion protein was obtained.

c. Abs 280 values were measured, and protein concentrations were converted from the obtained values, and they are shown in Table 2 below.

division Protein concentration (OD 280 nm) 1 mg DCT / ml 1.65 1 mg Tyrp1 / ml 1.42

Example 1

40 [micro] l of DCT having an activity of 1000 units / ml prepared in Preparation Example 1 and 40 [micro] l of Tyrp1 having an activity of 1000 units / ml were added to 40 [micro] l of tyrosinase enzyme having activity of 1000 unit / ml, 250 μl of 200 mM Tris buffer pH 8, 25 μl of 10% Tween 20, and 105 μl of distilled water were added to prepare an enzyme composition 132 (stock solution).

Stock solution Volumn (500 μl) 200 mM Tris buffer pH8 250 μl 10% Tween 20 25 μl Tyrosinase (> = 1000 unit / ml) 40 μl DCT (> = 1000 unit / ml) 40 μl TYRP1 (> = 1000 unit / ml) 40 μl D.W 105 μl

Next, urine samples were collected from normal and cancer patients. The method of collecting the urine samples was done by taking 30 ml of sample from the urine without taking the first urine from the urethra in the first urine collection after the morning wake.

In order to measure the concentration of tyrosine present in a certain amount, the same amount of 500 μl of each of the urine collected from the normal person and the cancer patient was reacted with the enzyme composition for detecting tyrosine 132 of Example 1 (total 1000 μl each).

Then, the color of the urine produced by each produced eumelanin is compared with the control color table of FIG. 1, and is shown in Table 4 below.

Color of urine by eumelanin Urine of a normal person Yellow to tan Urine of cancer patient black

As shown in Table 4, when the urine of the normal person is reacted with the enzyme composition 132 of the present invention, the urine color produced by the eumelanin is yellow to yellowish brown, while the urine of the cancer patient reacts with the enzyme composition 132 It was confirmed that the color of urine produced by the produced eumelanin was black.

This shows that tyrosine is present in large amounts in urine of cancer patients.

Example 2

Compared with the conventional enzyme composition 132 containing only tyrosinase, when the enzyme composition 132 of the present invention containing DCT and Tyrp1 in addition to tyrosinase is used, the color of the urine produced by the produced uromelanin is further increased In order to find out that it can appear black, the following experiment was carried out.

(A) obtained by mixing 500 μl of normal human urine collected in the same manner as in Example 1 with the enzyme composition (132) of the present invention comprising 40 units of tyrosinase, 20 units of DCT and 20 units of Tyrp1, (B) in which 500 μl of the patient's urine was mixed with a conventional enzyme composition (132) containing only 40 units of tyrosinase, and 500 μl of urine of the cancer patient were divided into 40 units of tyrosinase, 20 units of DCT and 20 units of Tyrp1 (C) mixed with the enzyme composition (132) of the present invention were reacted at 37 占 폚 for 30 minutes, respectively.

7, the mixture (A) in which the enzyme composition (132) according to the present invention and the urine of the normal human being are reacted shows a yellow color, while the mixture (B) and (C) Yellowish brown or black.

Especially, as a result of urine detection of a cancer patient, it was confirmed that the color of the mixture (C) containing DCT and Tyrp 1 is more black than the mixture (B) containing only tyrosinase.

Therefore, by adding the DCT and Tyrp1 enzyme to the tyrosinase, the enzyme composition 132 according to the present invention not only accelerates the eumelanin biosynthetic pathway, but also induces the color of the produced uromelanin to a more black path .

As described above, it will be understood that the present invention is implemented in a modified form without departing from the essential characteristics of the present invention.

It is therefore to be understood that the specified embodiments are to be considered in an illustrative rather than a restrictive sense and that the scope of the invention is indicated by the appended claims rather than by the foregoing description and that all such differences falling within the scope of equivalents thereof are intended to be embraced therein It should be interpreted.

100: housing 110: inlet
111: blocking membrane 112: sample holding unit
113: sample inlet 114: control inlet
116: air discharge path 123: window
124: Color chart 130: Sample chamber
132: Enzyme composition 134: Sample inlet
140: Control chamber 142: Color small amount
144: control inflow path 170: switch
172: Battery 200:

<110> CubeBio <120> Enzyme Compositions for Detecting Cancer Biomarker Tyrosine <130> DP18030 <160> 7 <170> KoPatentin 3.0 <210> 1 <211> 19 <212> RNA <213> Artificial Sequence <220> <223> primer <400> 1 atgggccttg tgggatggg 19 <210> 2 <211> 26 <212> RNA <213> Artificial Sequence <220> <223> primer <400> 2 ctaggcttcc tccgtgtatc tcttgc 26 <210> 3 <211> 26 <212> RNA <213> Artificial Sequence <220> <223> primer <400> 3 atgaaatctt acaacgtcct ccccct 26 <210> 4 <211> 26 <212> RNA <213> Artificial Sequence <220> <223> primer <400> 4 tcagaccatg gagtggttag gattcg 26 <210> 5 <211> 556 <212> PRT <213> Agaricus bisporus <400> 5 Met Ser Leu Ile Ala Thr Val Gly Pro Thr Gly Gly Val Lys Asn Arg   1 5 10 15 Leu Asn Ile Val Asp Phe Val Lys Asn Glu Lys Phe Phe Thr Leu Tyr              20 25 30 Val Arg Ser Leu Glu Leu Leu Gln Ala Lys Glu Gln His Asp Tyr Ser          35 40 45 Ser Phe Phe Gln Leu Ala Gly Ile His Gly Leu Pro Phe Thr Glu Trp      50 55 60 Ala Lys Glu Arg Pro Ser Met Asn Leu Tyr Lys Ala Gly Tyr Cys Thr  65 70 75 80 His Gly Gln Val Leu Phe Pro Thr Trp His Arg Thr Tyr Leu Ser Val                  85 90 95 Phe Glu Gln Ile Leu Gln Gly Ala Ala Ile Glu Val Ala Asn Lys Phe             100 105 110 Thr Ser Asn Gln Thr Asp Trp Ile Gln Ala Ala Gln Asp Leu Arg Gln         115 120 125 Pro Tyr Trp Asp Trp Gly Phe Glu Leu Met Pro Pro Asp Glu Val Ile     130 135 140 Lys Asn Glu Glu Val Asn Ile Thr Asn Tyr Asp Gly Lys Lys Ile Ser 145 150 155 160 Val Lys Asn Pro Ile Leu Arg Tyr His Phe His Pro Ile Asp Pro Ser                 165 170 175 Phe Lys Pro Tyr Gly Asp Phe Ala Thr Trp Arg Thr Thr Val Arg Asn             180 185 190 Pro Asp Arg Asn Arg Arg Glu Asp Ile Pro Gly Leu Ile Lys Lys Met         195 200 205 Arg Leu Glu Glu Gly Gln Ile Arg Glu Lys Thr Tyr Asn Met Leu Lys     210 215 220 Phe Asn Asp Ala Trp Glu Arg Phe Ser Asn His Gly Ile Ser Asp Asp 225 230 235 240 Gln His Ala Asn Ser Leu Glu Ser Val His Asp Asp Ile His Val Met                 245 250 255 Val Gly Tyr Gly Lys Ile Glu Gly His Met Asp His Pro Phe Phe Ala             260 265 270 Ala Phe Asp Pro Ile Phe Trp Leu His His Thr Asn Val Asp Arg Leu         275 280 285 Leu Ser Leu Trp Lys Ala Ile Asn Pro Asp Val Trp Val Thr Ser Gly     290 295 300 Arg Asn Arg Asp Gly Thr Met Gly Ile Ala Pro Asn Ala Gln Ile Asn 305 310 315 320 Asp Glu Thr Pro Leu Glu Pro Phe Tyr Gln Ser Glu Asp Lys Val Trp                 325 330 335 Thr Ser Ala Ser Leu Ala Asp Thr Ala Arg Leu Gly Tyr Ser Tyr Pro             340 345 350 Asp Phe Asp Lys Leu Val Gly Gly Thr Lys Glu Leu Ile Arg Asp Ala         355 360 365 Ile Asp Asp Leu Ile Asp Glu Arg Tyr Gly Ser Lys Pro Ser Ser Gly     370 375 380 Ala Arg Asn Thr Ala Phe Asp Leu Leu Ala Asp Phe Lys Gly Ile Thr 385 390 395 400 Lys Glu His Lys Glu Asp Leu Lys Met Tyr Asp Trp Thr Ile His Val                 405 410 415 Ala Phe Lys Lys Phe Glu Leu Lys Glu Ser Phe Ser Leu Leu Phe Tyr             420 425 430 Phe Ala Ser Asp Gly Gly Asp Tyr Asp Gln Glu Asn Cys Phe Val Gly         435 440 445 Ser Ile Asn Ala Phe Arg Gly Thr Thr Pro Glu Thr Cys Ala Asn Cys     450 455 460 Gln Asp Asn Glu Asn Leu Ile Gln Glu Gly Phe Ile His Leu Asn His 465 470 475 480 Tyr Leu Ala Arg Asp Leu Glu Ser Phe Glu Pro Gln Asp Val His Lys                 485 490 495 Phe Leu Lys Glu Lys Gly Leu Ser Tyr Lys Leu Tyr Ser Arg Glu Asp             500 505 510 Lys Ser Leu Thr Ser Leu Ser Val Lys Ile Glu Gly Arg Pro Leu His         515 520 525 Leu Pro Pro Gly Glu His Arg Pro Lys Tyr Asp His Thr Gln Asp Arg     530 535 540 Val Val Phe Asp Asp Val Ala Val His Val Ile Asn 545 550 555 <210> 6 <211> 517 <212> PRT <213> Mus musculus <400> 6 Met Gly Leu Val Gly Trp Gly Leu Leu Leu Gly Cys Leu Gly Cys Gly   1 5 10 15 Ile Leu Leu Arg Ala Arg Ala Gln Phe Pro Arg Val Cys Met Thr Leu              20 25 30 Asp Gly Val Leu Asn Lys Glu Cys Cys Pro Pro Leu Gly Pro Glu Ala          35 40 45 Thr Asn Ile Cys Gly Phe Leu Glu Gly Arg Gly Gln Cys Ala Glu Val      50 55 60 Gln Thr Asp Thr Arg Pro Trp Ser Gly Pro Tyr Ile Leu Arg Asn Gln  65 70 75 80 Asp Asp Arg Glu Gln Trp Pro Arg Lys Phe Phe Asn Arg Thr Cys Lys                  85 90 95 Cys Thr Gly Asn Phe Ala Gly Tyr Asn Cys Gly Gly Cys Lys Phe Gly             100 105 110 Trp Thr Gly Pro Asp Cys Asn Arg Lys Lys Pro Ala Ile Leu Arg Arg         115 120 125 Asn Ile His Ser Leu Thr Ala Gln Glu Arg Glu Gln Phe Leu Gly Ala     130 135 140 Leu Asp Leu Ala Lys Lys Ser Ile His Pro Asp Tyr Val Ile Thr Thr 145 150 155 160 Gln His Trp Leu Gly Leu Leu Gly Pro Asn Gly Thr Gln Pro Gln Ile                 165 170 175 Ala Asn Cys Ser Val Tyr Asp Phe Phe Val Trp Leu His Tyr Tyr Ser             180 185 190 Val Arg Asp Thr Leu Leu Gly Pro Gly Arg Pro Tyr Lys Ala Ile Asp         195 200 205 Phe Ser His Gln Gly Pro Ala Phe Val Thr Trp His Arg Tyr His Leu     210 215 220 Leu Trp Leu Glu Arg Glu Leu Gln Arg Leu Thr Gly Asn Glu Ser Phe 225 230 235 240 Ala Leu Pro Tyr Trp Asn Phe Ala Thr Gly Lys Asn Glu Cys Asp Val                 245 250 255 Cys Thr Asp Glu Leu Leu Gly Ala Ala Arg Gln Asp Asp Pro Thr Leu             260 265 270 Ile Ser Arg Asn Ser Arg Phe Ser Thr Trp Glu Ile Val Cys Asp Ser         275 280 285 Leu Asp Asp Tyr Asn Arg Arg Val Thr Leu Cys Asn Gly Thr Tyr Glu     290 295 300 Gly Leu Leu Arg Arg Asn Lys Val Gly Arg Asn Asn Glu Lys Leu Pro 305 310 315 320 Thr Leu Lys Asn Val Gln Asp Cys Leu Ser Leu Gln Lys Phe Asp Ser                 325 330 335 Pro Pro Phe Phe Gln Asn Ser Thr Phe Ser Phe Arg Asn Ala Leu Glu             340 345 350 Gly Phe Asp Lys Ala Asp Gly Thr Leu Asp Ser Gln Val Met Asn Leu         355 360 365 His Asn Leu Ala His Ser Phe Leu Asn Gly Thr Asn Ala Leu Pro His     370 375 380 Ser Ala Asn Asp Pro Val Phe Val Val Leu His Ser Phe Thr Asp 385 390 395 400 Ala Ile Phe Asp Glu Trp Leu Lys Arg Asn Asn Pro Ser Thr Asp Ala                 405 410 415 Trp Pro Gln Glu Leu Ala Pro Ile Gly His Asn Arg Met Tyr Asn Met             420 425 430 Val Pro Phe Pro Pro Val Thr Asn Glu Glu Leu Phe Leu Thr Ala         435 440 445 Glu Gln Leu Gly Tyr Asn Tyr Ala Val Asp Leu Ser Glu Glu Glu Ala     450 455 460 Pro Val Trp Ser Thr Thr Leu Ser Val Val Ile Gly Ile Leu Gly Ala 465 470 475 480 Phe Val Leu Leu Leu Gly Leu Leu Ala Phe Leu Gln Tyr Arg Arg Leu                 485 490 495 Arg Lys Gly Tyr Ala Pro Leu Met Glu Thr Gly Leu Ser Ser Lys Arg             500 505 510 Tyr Thr Glu Glu Ala         515 <210> 7 <211> 537 <212> PRT <213> Mus musculus <400> 7 Met Lys Ser Tyr Asn Val Leu Pro Leu Ala Tyr Ile Ser Leu Phe Leu   1 5 10 15 Met Leu Phe Tyr Gln Val Trp Ala Gln Phe Pro Arg Glu Cys Ala Asn              20 25 30 Ile Glu Ala Leu Arg Arg Gly Val Cys Cys Pro Asp Leu Leu Pro Ser          35 40 45 Ser Gly Pro Gly Thr Asp Pro Cys Gly Ser Ser Ser Gly Arg Gly Arg      50 55 60 Cys Val Ala Val Ile Ala Asp Ser Arg Pro His Ser Arg His Tyr Pro  65 70 75 80 His Asp Gly Lys Asp Asp Arg Glu Ala Trp Pro Leu Arg Phe Phe Asn                  85 90 95 Arg Thr Cys Gln Cys Asn Asp Asn Phe Ser Gly His Asn Cys Gly Thr             100 105 110 Cys Arg Pro Gly Trp Arg Gly Ala Ala Cys Asn Gln Lys Ile Leu Thr         115 120 125 Val Arg Arg Asn Leu Leu Asp Leu Ser Pro Glu Glu Lys Ser His Phe     130 135 140 Val Arg Ala Leu Asp Met Ala Lys Arg Thr Thr His Pro Gln Phe Val 145 150 155 160 Ile Ala Thr Arg Arg Leu Glu Asp Ile Leu Gly Pro Asp Gly Asn Thr                 165 170 175 Pro Gln Phe Glu Asn Ile Ser Val Tyr Asn Tyr Phe Val Trp Thr His             180 185 190 Tyr Tyr Ser Val Lys Lys Thr Phe Leu Gly Thr Gly Gln Glu Ser Phe         195 200 205 Gly Asp Val Asp Phe Ser His Glu Gly Pro Ala Phe Leu Thr Trp His     210 215 220 Arg Tyr His Leu Leu Gln Leu Glu Arg Asp Met Gln Glu Met Leu Gln 225 230 235 240 Glu Pro Ser Phe Ser Leu Pro Tyr Trp Asn Phe Ala Thr Gly Lys Asn                 245 250 255 Val Cys Asp Val Cys Thr Asp Asp Leu Met Gly Ser Arg Ser Asn Phe             260 265 270 Asp Ser Thr Leu Ile Ser Pro Asn Ser Val Phe Ser Gln Trp Arg Val         275 280 285 Val Cys Glu Ser Leu Glu Glu Tyr Asp Thr Leu Gly Thr Leu Cys Asn     290 295 300 Ser Thr Glu Gly Gly Pro Ile Arg Arg Asn Pro Ala Gly Asn Val Gly 305 310 315 320 Arg Pro Ala Val Gln Arg Leu Pro Glu Pro Gln Asp Val Thr Gln Cys                 325 330 335 Leu Glu Val Arg Phe Asp Thr Pro Pro Phe Tyr Ser Asn Ser Thr             340 345 350 Asp Ser Phe Arg Asn Thr Val Glu Gly Tyr Ser Ala Pro Thr Gly Lys         355 360 365 Tyr Asp Pro Ala Val Arg Ser Leu His Asn Leu Ala His Leu Phe Leu     370 375 380 Asn Gly Thr Gly Gly Gln Thr His Leu Ser Pro Asn Asp Pro Ile Phe 385 390 395 400 Val Leu Leu His Thr Phe Thr Asp Ala Val Phe Asp Glu Trp Leu Arg                 405 410 415 Arg Tyr Asn Ala Asp Ile Ser Thr Phe Pro Leu Glu Asn Ala Pro Ile             420 425 430 Gly His Asn Arg Gln Tyr Asn Met Val Pro Phe Trp Pro Pro Val Thr         435 440 445 Asn Thr Glu Met Phe Val Thr Ala Pro Asp Asn Leu Gly Tyr Ala Tyr     450 455 460 Glu Val Gln Trp Pro Gly Gln Glu Phe Thr Val Ser Glu Ile Ile Thr 465 470 475 480 Ile Ala Val Ala Ala Leu Leu Ala Val Ala Ile Phe Gly Val                 485 490 495 Ala Ser Cys Leu Ile Arg Ser Ser Ser Thr Lys Asn Glu Ala Asn Gln             500 505 510 Pro Leu Leu Thr Asp His Tyr Gln Arg Tyr Ala Glu Asp Tyr Glu Glu         515 520 525 Leu Pro Asn Pro Asn His Ser Met Val     530 535

Claims (8)

A housing having a sample chamber and a control chamber in which the injected sample is separated and accommodated, and a window for observing the sample chamber and the inside of the control chamber on one side;
And an enzyme composition which is accommodated in the sample chamber and reacts with tyrosine contained in the sample to discolor the sample,
Wherein a color table which becomes opaque gradually from one side is formed in a portion of the window corresponding to the control chamber,
Wherein the specimen is any one of body fluids derived from a human body and the body fluids include blood, urine, serum, plasma, lymph, tissue fluid, secretion,
The enzyme composition comprises tyrosine (L-Tyrosine) present in a sample, including tyrosinase, Dopachrome tautomerase (DCT) and tyrosinase-related protein 1 (Tyrp1) Wherein the enzyme is an enzyme composition for tyrosine detection, wherein the color of the sample is discolored corresponding to the sample color. delete delete The method according to claim 1,
(DCT) and tyrosinase-related protein 1 (Tyrp1) in a ratio of 0.5 to 2 parts by weight based on 1 part by weight of the tyrosinase, and 40 parts by weight of tyrosinase, , L-Tyrosine into Eumelanin, and the color of the specimen is changed from its original color to black as the concentration of tyrosine increases. &Lt; / RTI &gt; 5. The pharmaceutical composition according to claim 4,
Further comprising a phosphate buffer or Tris-HCl buffer solution having a final concentration of 50 to 100 mM, a Tween 20 having a final concentration of 0.1 to 0.5%, and distilled water, And the sample is accommodated in the sample chamber, wherein the sample is color-coded in correspondence with the sample color. The method according to claim 1,
Further comprising a light guide plate unit installed on an inner wall surface of the housing opposite to the window to irradiate light to the sample chamber and the control chamber. . The light guide plate according to claim 6,
A light guide plate having an incident surface on which light irradiated from a light source is incident and an exit surface on which the light propagated through the incident surface is deflected,
A diffusion sheet for adjusting the diffusion and propagation direction of the light transmitted through the light guide plate,
A prism sheet for refracting and diffusing light emitted from the diffusion sheet to increase brightness,
And a reflection sheet for reflecting the light beam directly below the light source or the light guide plate is laminated on the lower side of the light guide plate. The method according to claim 1,
And a sample inlet provided at one side of the housing for introducing the sample into the sample chamber and the control chamber,
Wherein the inlet has an elastic material blocking membrane installed on the upper portion to prevent leakage of urine to the outside, and a specimen accommodating unit for separating the introduced urine into the specimen chamber and the control chamber while the specimen is loaded, And a sample chamber, and an air discharge path through which the sample of the sample chamber is discharged to the specimen accommodating portion.
KR1020160078660A 2016-06-23 2016-06-23 Diagnostic Kit for Detecting Cancer including Discoloration Colormatrix KR101724617B1 (en)

Priority Applications (1)

Application Number Priority Date Filing Date Title
KR1020160078660A KR101724617B1 (en) 2016-06-23 2016-06-23 Diagnostic Kit for Detecting Cancer including Discoloration Colormatrix

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
KR1020160078660A KR101724617B1 (en) 2016-06-23 2016-06-23 Diagnostic Kit for Detecting Cancer including Discoloration Colormatrix

Publications (1)

Publication Number Publication Date
KR101724617B1 true KR101724617B1 (en) 2017-04-10

Family

ID=58581248

Family Applications (1)

Application Number Title Priority Date Filing Date
KR1020160078660A KR101724617B1 (en) 2016-06-23 2016-06-23 Diagnostic Kit for Detecting Cancer including Discoloration Colormatrix

Country Status (1)

Country Link
KR (1) KR101724617B1 (en)

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0332444B1 (en) * 1988-03-11 1992-07-01 SAM I1 PHARMACEUTICAL MANUFACTURING CO., Ltd Reagent for diagnosis of cancer
KR20130083618A (en) * 2012-01-13 2013-07-23 주식회사 아이센스 Catridge for sensing elements in a sample
KR20150068755A (en) * 2013-12-12 2015-06-22 염상철 Apparatus for analyzing sample automatically by using defined substrate technology

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
EP0332444B1 (en) * 1988-03-11 1992-07-01 SAM I1 PHARMACEUTICAL MANUFACTURING CO., Ltd Reagent for diagnosis of cancer
KR20130083618A (en) * 2012-01-13 2013-07-23 주식회사 아이센스 Catridge for sensing elements in a sample
KR20150068755A (en) * 2013-12-12 2015-06-22 염상철 Apparatus for analyzing sample automatically by using defined substrate technology

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
Journal of Investigative Dermatology Vol.100(2):126-131 *
Journal of Investigative Dermatology Vol.100(2):126-131*

Similar Documents

Publication Publication Date Title
AU608510B2 (en) Method and apparatus for monitoring glucose
WO2009021039A1 (en) Device for detection of molecules of interest
AU2001253079A1 (en) Methods, compositions and kits for the detection and monitoring of breast cancer
KR101705185B1 (en) Self-detectable Diagnostic Kit for Detecting Cancer Existence Using Enzyme Composition And Method for Diagnosing Cancer Existence
JP2005230521A (en) Humor component detecting device and humor component detecting system
ES2325923T3 (en) COLORIMETRIC SUBSTRATES, COLORIMETRIC SENSORS AND USE PROCEDURES.
KR101702596B1 (en) Self-detectable Diagnostic Kit for Detecting Cancer Existence Using Enzyme Composition
EP1767636A3 (en) Compounds for immunotherapy and diagnosis of colon cancer and methods for their use
Nikawa et al. Effects of dietary sugars and saliva and serum on Candida bioflim formation on acrylic surfaces
CN104975104B (en) Alzheimer disease diagnosis and treatment marker and its application
Su et al. Ratiometric glucose sensing based on fluorescent oxygen films and glucose oxidase
KR101724617B1 (en) Diagnostic Kit for Detecting Cancer including Discoloration Colormatrix
DK2687847T3 (en) PROBE TO ANALYZE BIOLOGICAL Tissue AND PROCEDURE FOR USING SAME
KR101749629B1 (en) System and Method for Determining Color of Cancer Diagnostic Kit using Smart Devices
CN110272996A (en) Biomarker relevant to glaucoma occurrence and development and its application
KR101685091B1 (en) High Resolution Window Diagnostic Kit for Detecting Cancer Existence
KR101666957B1 (en) High Resolution Window Diagnostic Kit with Own Light Source for Detecting Cancer Existence
KR101707122B1 (en) System for Determining Color of Cancer Diagnostic Kit using Smart Devices
KR101694322B1 (en) Flat-plate type cancer diagnostic kit with scanner
KR101666954B1 (en) Module type High Resolution Window Diagnostic Kit with Own Light Source for Detecting Cancer Existence
KR101725837B1 (en) Module type Decision Condition Control Window Diagnostic Kit with Own Light Source for Detecting Cancer Existence
KR101702601B1 (en) Portable Diagnostic Kit for Detecting Cancer Existence Using Enzyme Composition
KR101694341B1 (en) Decision Condition Control Window Diagnostic Kit with Own Light Source for Detecting Cancer Existence
KR101666952B1 (en) Flat-plate type cancer diagnostic kit with high resolution window
KR101749627B1 (en) System and Method for Calculate the Color Change Amount of Enzyme Compositions for Tyrosine

Legal Events

Date Code Title Description
GRNT Written decision to grant