WO2011132978A2 - Appareil d'analyse de polarisation à fluorescence incluant une double source de lumière - Google Patents

Appareil d'analyse de polarisation à fluorescence incluant une double source de lumière Download PDF

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Publication number
WO2011132978A2
WO2011132978A2 PCT/KR2011/002912 KR2011002912W WO2011132978A2 WO 2011132978 A2 WO2011132978 A2 WO 2011132978A2 KR 2011002912 W KR2011002912 W KR 2011002912W WO 2011132978 A2 WO2011132978 A2 WO 2011132978A2
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light
light source
sample chamber
vertical
disposed
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PCT/KR2011/002912
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English (en)
Korean (ko)
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WO2011132978A3 (fr
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김성우
김덕중
류호선
신수정
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나노바이오시스(주)
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Publication of WO2011132978A3 publication Critical patent/WO2011132978A3/fr

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    • 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
    • G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
    • G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
    • G01N21/64—Fluorescence; Phosphorescence
    • G01N21/6445—Measuring fluorescence polarisation

Definitions

  • the present invention relates to a fluorescence polarization analyzer for analyzing a sample solution containing a target material.
  • Fluorescence Polarization (FP) measurement which measures the degree of fluorescence polarization generated when a fluorescent substance is irradiated with polarized light in a sample solution, was devised by Perrin in 1926. Thereafter, the measurement method is very useful for quantitative and qualitative analysis of materials present in a sample solution by detecting a change in fluorescence signal according to a change in material properties. In particular, it can be usefully applied to the detection of chemical or biological or biochemical materials, and is widely used for analysis and diagnostic purposes in the fields of life sciences, genetic engineering and medicine, environment, food, military, marine, and livestock.
  • the present invention is to provide a fluorescence polarization analyzer that can exert an excellent performance in analyzing the target material contained in the sample solution.
  • One embodiment of the present invention comprises a sample chamber of a light transparent material for placing a sample solution containing a target material; A first light source and a second light source disposed separately from the sample chamber and disposed to emit light toward the sample chamber; A first vertical polarizer disposed between the sample chamber and the first light source and polarizing the vertical light of the light emitted from the first light source; A first horizontal polarizer disposed between the sample chamber and the second light source and polarizing horizontal light of light emitted from the second light source; A light detector spaced apart from the sample chamber; And a second vertical or horizontal polarizer disposed between the sample chamber and the light detector and polarizing the vertical or horizontal light of the light emitted from the sample chamber.
  • the target material may be selected from the group consisting of chemicals, sugars, peptides, nucleic acids, proteins, viruses, cells and organelles.
  • the target material may be a compound capable of forming a fluorescent material and a target material-fluorescent material complex.
  • the target substance-fluorescent substance complex may be an antigen-antibody complex.
  • the light transparent material is a plastic selected from the group consisting of polymethyl methacrylate (polymethylmethacrylate, PMMA), polycarbonate (PC), cycloolefin copolymer (COC) It may be a material or a glass material.
  • the sample chamber may further include a disposable sample container including the sample solution, which is removable within the sample chamber and has a light transparent material.
  • the first light source and the second light source may be a light emitting diode (LED) or a laser light source.
  • the first light source and the second light source may be spaced apart from the sample chamber at the same distance.
  • the first and second light sources may be spaced apart from each other to be symmetrical about the sample chamber.
  • the light detector may be spaced apart from the sample chamber perpendicular to the connection axis of the first light source and the second light source.
  • the light source may further include a first optical lens disposed between the first light source and the first vertical polarizer and configured to collect light emitted from the first light source. It may further include a second optical lens disposed between the horizontal polarizer, for collecting the light emitted from the second light source.
  • the light is disposed between the first optical lens and the first vertical polarizer and selects light having a predetermined wavelength with respect to the first light source from the light passing through the first optical lens.
  • a first light filter disposed between the second optical lens and the first horizontal polarizer, for selecting light of a predetermined wavelength with respect to the second light source from light passing through the second optical lens. It may further comprise a second light filter.
  • the light emitting device may further include a third optical lens disposed between the sample chamber and the second vertical or horizontal polarizer and collecting light emitted from the sample chamber.
  • a predetermined wavelength is disposed between the third optical lens and the second vertical or horizontal polarizer and is related to the first light source or the second light source from the light passing through the third optical lens. It may further include a third light filter for selecting the light.
  • the light emitting device may further include a fourth optical lens disposed between the second vertical or horizontal polarizers and collecting light emitted from the second vertical or horizontal polarizers.
  • FIG. 1 illustrates a fluorescence polarization analyzer according to an embodiment of the present invention.
  • FIG. 2 illustrates detailed modules included in a fluorescence polarization analyzer according to an embodiment of the present invention.
  • FIG. 3 is a comparison of the results of analyzing a target material included in a sample solution using another fluorescence polarization analyzer commercially available with a fluorescence polarization analyzer according to an embodiment of the present invention.
  • FIG. 1 illustrates a fluorescence polarization analyzer according to an embodiment of the present invention.
  • the fluorescence polarization analyzer 1 includes a sample chamber 100 made of a light transparent material for disposing a sample solution containing a target material; A first light source (200) and a second light source (300) disposed separately from the sample chamber (100) and disposed to emit light toward the sample chamber; A first vertical polarizer 400 disposed between the sample chamber 100 and the first light source 200 to polarize the vertical light of the light emitted from the first light source 200; A first horizontal polarizer (500) disposed between the sample chamber (100) and the second light source (300) and polarizing horizontal light of light emitted from the second light source (300); A light detector 600 spaced apart from the sample chamber 100; And a second vertical or horizontal polarizer 700 disposed between the sample chamber 100 and the light detector 100 and polarizing the vertical or horizontal light of the light emitted from the sample chamber 100.
  • the fluorescence polarization analyzer 1 includes a sample chamber 100 of a light transparent material for placing a sample solution (not shown) containing a target material (not shown).
  • the fluorescence polarization generally emits light having the same directionality as the received light when the fluorescent material receives light having a specific directionality, in which case the molecular weight of the fluorescent material and the viscosity of the solution containing the target material Or a phenomenon in which the directionality of the emitted light is different from the directionality of the received light according to a property including the density of the fluorescent material and the like.
  • the fluorescence polarization analyzer 1 refers to a device for measuring and analyzing a target substance included in a sample solution based on the principle of fluorescence polarization.
  • the fluorescence polarization analyzer 1 irradiates light (polarization) having a specific directionality to a sample solution containing a fluorescent material, and measures the degree of change in the directionality of light emitted from the sample solution (polarization degree) and An apparatus for analyzing to measure and analyze the target substance in the sample solution. Therefore, the fluorescence polarization analysis apparatus 1 according to an embodiment of the present invention is an apparatus including modules for performing the fluorescence polarization analysis reaction, and the detailed modules not described herein perform the fluorescence polarization analysis reaction. It is assumed on the premise that all of them are disclosed in the prior art for the following and provided in the obvious range.
  • the sample solution includes a target material and a fluorescent material.
  • the target material refers to any material to be measured and analyzed in the sample solution, and specifically, the target material includes a chemical or biological or biochemical target material.
  • the target material may include one selected from the group consisting of chemicals, sugars, peptides, nucleic acids, proteins, viruses, cells, and organelles.
  • the fluorescent material includes all materials that emit light having a specific direction according to the fluorescence polarization principle when light having a specific direction is irradiated.
  • the fluorescent material may be introduced into the sample solution to bind with the target material, form a target material-fluorescent material complex, and the target material-fluorescent material complex may be an antigen-antibody complex through an antigen-antibody reaction. Therefore, when light having a specific directionality is irradiated, the degree of polarization is changed by the change in the properties of the target material-fluorescent material composite, and the degree of polarization can be measured.
  • the sample chamber 100 may be made of a light transparent material or may include a light transparent material as a whole.
  • the light transparent material may be a plastic material or a glass material selected from the group consisting of polymethylmethacrylate (PMMA), polycarbonate (PC), and cycloolefin copolymer (COC). Can be.
  • the sample chamber 100 may directly receive a sample solution containing the target material.
  • the sample chamber 100 may further include a disposable sample container (not shown) including the sample solution, which is removable in the sample chamber 100 and has a light transparent material.
  • the disposable sample container may be made of a light transparent material or may include a light transparent material as a whole.
  • the light transparent material is a light transparent plastic, for example, a plastic material selected from the group consisting of polymethylmethacrylate (PMMA), polycarbonate (PC) or cycloolefin copolymer (COC) or It may be a glass material, but is not limited thereto.
  • the disposable sample container has an outer wall designed to be detachable from the sample chamber 100, and includes a receiving portion for accommodating the sample solution.
  • the outer wall is designed to minimize reflection or scattering of vertical light or horizontal light, and may have, for example, a cylindrical shape such as a cylindrical, triangular, or square pillar.
  • the disposable sample container may include a single or a plurality of receivers to contain a single or a plurality of sample solutions by injection molding or glass manufacturing method.
  • the fluorescence polarization analyzer 1 may be spaced apart from the sample chamber 100, and the first light source 200 and the first light source 200 disposed to emit light toward the sample chamber 100. 2 includes a light source 300.
  • the first light source 200 and the second light source 300 includes all light sources capable of emitting light.
  • the first light source and the second light source may be a light emitting diode (LED) or a laser light source. Can be.
  • LED light emitting diode
  • the fluorescence polarization analyzer 1 is disposed between the sample chamber 100 and the first light source 200 and polarizes vertical light of light emitted from the first light source 200.
  • a first horizontal polarizer 400 and a first horizontal polarizer disposed between the sample chamber 100 and the second light source 300 and polarizing horizontal light of light emitted from the second light source 300 ( 500).
  • the polarizers 400 and 500 polarize the light emitted from the light sources 200 and 300 to have a specific direction.
  • the first polarizer 400 polarizes vertical light of light emitted from the first light source 200
  • the second polarizer 500 polarizes horizontal light of light from the second light source 300.
  • a fluorescence polarization analyzer includes a single polarizer that polarizes light emitted from a single light source. Therefore, there is a need for a separate drive device for adjusting a single polarizer in a vertically polarized state or a horizontally polarized state in order to obtain light having a specific directionality, for example, vertical light or horizontal light from a single light source.
  • the first vertical polarizer 400 and the second horizontal polarizer 500 are the sample chamber 100 and the first light source 200.
  • the fluorescence polarization analyzer 1 By being disposed between and between the sample chamber 100 and the second light source 300, there is no need to adjust the position of the single polarizer, so that a separate polarizer for adjusting the single polarizer to the vertical polarization state or the horizontal polarization state is therefore required. Since no driving device is required, the fluorescence polarization analyzer can be miniaturized.
  • the first vertical polarizer 400 and the second horizontal polarizer 500 are the sample chamber 100 and the first light source 200. Between and between the sample chamber 100 and the second light source 300, respectively, the vertical light of the light emitted from the first light source 200 and the horizontal light of the light emitted from the second light source 300 Since the light is directly irradiated to the sample chamber 100, the light emitted from the target material in the sample chamber 100 may be individually measured.
  • the fluorescence polarization analyzer 1 is disposed between the photo detector 600 and the sample chamber 100 and the photo detector 100 spaced apart from the sample chamber 100, And a second vertical or horizontal polarizer 700 for polarizing the vertical or horizontal light of the light emitted from the sample chamber 100.
  • the light detector 600 measures and analyzes light emitted from the sample chamber 100.
  • the second vertical or horizontal polarizer 700 is disposed between the sample chamber 100 and the light detector 100.
  • the light emitted from the first light source 200 is vertical by the first vertical polarizer 400. Only the vertical light is polarized with light to be irradiated to the sample chamber 100 and thus has various directional light emitted from the sample chamber 100, for example, vertical light and horizontal light.
  • the light emitted from the second light source 300 is irradiated to the light detector 600 by a vertical polarizer, and is polarized into horizontal light by the first horizontal polarizer 500 and irradiated to the sample chamber 100.
  • the light detector 100 has two kinds of directional light irradiated to the sample chamber 100, that is, light having a single directional light emitted from a fluorescent material coupled to a target material from vertical light and horizontal light. That is, vertical light can be received as a detection signal.
  • the second horizontal polarizer 700 is disposed between the sample chamber 100 and the light detector 100, the light emitted from the first light source 200 is transmitted to the first vertical polarizer 400.
  • Polarized light into vertical light and irradiated to the sample chamber 100 and thus, only horizontal light among light having various directions, for example, vertical light and horizontal light emitted from the sample chamber 100, is
  • the light emitted from the second light source 300 is irradiated to the light detector 600 by a second horizontal polarizer, and the light is polarized into horizontal light by the first horizontal polarizer 500 and irradiated to the sample chamber 100.
  • the light detector 100 has two kinds of directional light irradiated to the sample chamber 100, that is, light having a single directional light emitted from a fluorescent material coupled to a target material from vertical light and horizontal light. That is, horizontal light can be received as a detection signal.
  • the light detector 600 receives an optical signal polarized by the second vertical or horizontal polarizer and measures and analyzes a target material included in the sample solution. Specifically, when the target solution is included in the sample solution included in the sample chamber 100, the material properties of the target material-fluorescent material composite are changed by the combination of the target material and the fluorescent material, and thus the sample chamber ( The direction of light emitted from the sample chamber 100 is changed relative to the direction of light irradiated to 100, and the light detector 600 measures and analyzes a change in polarization degree accordingly.
  • the polarization degree P of light emitted from the fluorescent material is represented by the following equation (1).
  • I H refers to the intensity of emitted light having the same polarization as incident light to the sample solution
  • I V refers to the intensity of emitted light having polarization perpendicular to the incident light to the sample solution. That is, the I H and I V are divided based on the polarization measurement direction of the light detector 600.
  • the sample chamber 100 Based on the first light source 200 passed through the second vertical polarizer 700 from the sample chamber 100 having the same polarization as the incident light (vertical light) passing through the first vertical polarizer 400.
  • the intensity of the emitted light (vertical light) is defined as I H and is perpendicular to the incident light (horizontal light) passing from the second light source 300 through the second horizontal polarizer 400 with respect to the sample chamber 100.
  • the intensity of the emitted light (vertical light) passing through the second vertical polarizer 700 from the sample chamber 100 having one polarization is defined as I V.
  • the first vertical light source 200 may be moved from the first light source 200 based on the sample chamber 100.
  • the intensity of the emitted light (horizontal light) passing through the second horizontal polarizer 700 from the sample chamber 100 having polarization perpendicular to the incident light (vertical light) passing through the polarizer 400 is defined as I V.
  • the intensity of the emitted light (horizontal light) passing through the polarizer 700 is defined as I H.
  • the polarization degree P has a specific value depending on the presence of the target substance in the sample solution, and the specific range according to the amount of the target substance in the sample solution, for example, the concentration. Will have the value of. For example, as the concentration of the target material in the sample solution increases, the degree of polarization P decreases.
  • FIG. 2 shows detailed modules included in the fluorescence polarization analyzer 1 according to an embodiment of the present invention.
  • the first light source 200 and the second light source 300 may be spaced apart from the sample chamber 100 by various distances, but are preferably spaced apart by the same distance, due to the characteristics of the optical system.
  • the first light source 200 and the second light source 300 may be spaced apart from each other so as to be symmetrical about the sample chamber 100.
  • the light detector 600 may be spaced apart from the sample chamber 100 at various distances, but in view of the characteristics of the optical system, preferably, the connection axis of the first light source 200 and the second light source 300 is Spaced vertically relative to.
  • the fluorescence polarization analyzer 1 may include first to fourth optical lenses 810, 820, 830, and 840.
  • the first to fourth optical lenses 810, 820, 830, and 840 collect the incident light and increase the intensity of the emitted light.
  • the first optical lens 810 is disposed between the first light source 200 and the first vertical polarizer 400, collects the light emitted from the first light source 200
  • the second optical lens ( 820 is disposed between the second light source 300 and the first horizontal polarizer 500, and collects the light emitted from the second light source 300
  • the third optical lens 830 is the sample chamber.
  • the fourth optical lens 840 is connected to the second vertical or horizontal polarizer 700.
  • the light is disposed between the light detectors 600 and collects light passing through the second vertical or horizontal polarizer 700.
  • the fluorescence polarization analyzer 1 may include first to third light filters 910, 920, and 930.
  • the first to third light filters 910, 920, and 930 select and emit light having a specific wavelength from incident light having various wavelength bands, and the light to the first to third light filters 910, 920, and 930. It may be variously selected depending on the light source to provide.
  • the first light filter 910 is disposed between the first optical lens 810 and the first vertical polarizer 400, and from the light passing through the first optical lens 810, the first light source 200. Select light with a predetermined wavelength with respect to For example, only light of the 488 nm wavelength band of light emitted from the first light source 200 may be passed.
  • the second light filter 920 is disposed between the second optical lens 820 and the first horizontal polarizer 500, and from the light passing through the second optical lens 820, the second light source ( Select light of a predetermined wavelength with respect to 300). For example, only light of the 488 nm wavelength band of light emitted from the second light source 300 may be passed.
  • the third light filter 930 is disposed between the third optical lens 830 and the second vertical or horizontal polarizer 700, and the third optical filter 930 is disposed from the light passing through the third optical lens 830.
  • the light of the predetermined wavelength with respect to the first light source 200 or the second light source 300 is selected. For example, only light having a wavelength of 533 nm may pass through light passing through the third optical lens 830.
  • FIG. 3 is a comparison of the results of analyzing a target material included in a sample solution by using a fluorescence polarization analyzer of another company commercialized with a fluorescence polarization analyzer according to an embodiment of the present invention.
  • the fluorescence polarization analyzer of another company used Sentry 100 of Diachemix was used for the fluorescence polarization reaction.
  • an Alftoxin kit (Kit) containing a control solution, an antibody solution, and an apratoxin FP tracer (including a flatoxin fluorescence polarization tracer) was used for the fluorescence polarization reaction.
  • the fluorescence polarization analysis reaction was performed by the fluorescence polarization analyzer according to an embodiment of the present invention.
  • About 200 ⁇ l of a solution containing about 20 ppb of Aflatoxin as a target substance about 100 ⁇ l of a solution containing about 10 ppb of apratoxin, and about 1 apratoxin in about 1 ml of the antibody solution.
  • About 10 ⁇ l of the solution containing ppb was added to prepare a control solution for each concentration of the target material. Thereafter, each of the control solutions was introduced into the sample chamber of the fluorescence polarization analyzer according to one embodiment of the present invention by about 300 ⁇ l, and each polarization degree (mP) was measured.
  • mP polarization degree
  • the fluorescence polarization analysis reaction by the fluorescence polarization analyzer of the other company was performed.
  • About 200 ⁇ l of a solution containing about 20 ppb of Aflatoxin as a target substance about 100 ⁇ l of a solution containing about 10 ppb of apratoxin, and about 1 apratoxin in about 1 ml of the antibody solution.
  • About 10 ⁇ l of the solution containing ppb was added to prepare a control solution for each concentration of the target material. Thereafter, 1.2 ml of each of the control solutions was introduced into the sample chamber of the third-party fluorescence polarization analyzer, and each polarization degree (mP) was measured.
  • mP polarization degree
  • the polarization value is lowered as the concentration of the target material increases. That is, the slopes of the first-order curves of equations (1) and (2) both have negative numbers.
  • the polarization degree value of the same concentration of the target material is higher by the fluorescence polarization analyzer according to an embodiment of the present invention. That is, when compared with the results of the fluorescence polarization analyzer of the other company, it can be seen that the fluorescence polarization analyzer according to an embodiment of the present invention is more sensitive to the same concentration of the target material.

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Abstract

La présente invention concerne un appareil d'analyse de polarisation à fluorescence incluant une double source de lumière. L'appareil d'analyse de polarisation à fluorescence selon la présente invention permet de réaliser une analyse efficace d'un matériau cible contenu dans un échantillon de solution.
PCT/KR2011/002912 2010-04-23 2011-04-22 Appareil d'analyse de polarisation à fluorescence incluant une double source de lumière WO2011132978A2 (fr)

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KR1020100037961A KR101037790B1 (ko) 2010-04-23 2010-04-23 듀얼 광원을 포함하는 형광 편광 분석 장치
KR10-2010-0037961 2010-04-23

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WO2011132978A3 WO2011132978A3 (fr) 2012-03-08

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WO2014176136A1 (fr) * 2013-04-21 2014-10-30 Mobileoct, Ltd. Appareil d'imagerie polariseur et ses procédés de séparation d'une lumière d'une surface d'un échantillon de ses couches diffuses plus profondes

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KR101776776B1 (ko) 2011-05-31 2017-09-11 삼성전자주식회사 형광 검출 광학계 및 이를 포함하는 다채널 형광 검출 장치
KR101680451B1 (ko) 2015-03-03 2016-11-28 충북대학교 산학협력단 형광 비등방성 측정시스템
KR101674018B1 (ko) * 2016-05-16 2016-11-08 (주)글로리바이오텍 Cd4, cd8 세포 정보를 이용한 hiv 진단 방법
PL233602B1 (pl) * 2017-10-16 2019-11-29 Inst Biologii Doswiadczalnej Im M Nenckiego Polskiej Akademii Nauk Urzadzenie do obrazowania przezroczystych obiektow

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