WO2009157695A2 - Thermal cycling reaction block and continuous real-time monitoring apparatus using the same - Google Patents
Thermal cycling reaction block and continuous real-time monitoring apparatus using the same Download PDFInfo
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- WO2009157695A2 WO2009157695A2 PCT/KR2009/003376 KR2009003376W WO2009157695A2 WO 2009157695 A2 WO2009157695 A2 WO 2009157695A2 KR 2009003376 W KR2009003376 W KR 2009003376W WO 2009157695 A2 WO2009157695 A2 WO 2009157695A2
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- Prior art keywords
- fluorescence
- capillary tube
- excitation light
- band pass
- beam splitter
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- 238000006243 chemical reaction Methods 0.000 title claims abstract description 45
- 238000005382 thermal cycling Methods 0.000 title claims abstract description 32
- 238000012544 monitoring process Methods 0.000 title claims abstract description 30
- 238000010438 heat treatment Methods 0.000 claims abstract description 56
- 230000005284 excitation Effects 0.000 claims abstract description 47
- 238000012546 transfer Methods 0.000 claims abstract description 12
- 230000001678 irradiating effect Effects 0.000 claims description 11
- 238000003780 insertion Methods 0.000 claims description 2
- 230000037431 insertion Effects 0.000 claims description 2
- 238000003752 polymerase chain reaction Methods 0.000 description 35
- 238000003753 real-time PCR Methods 0.000 description 8
- 238000000034 method Methods 0.000 description 6
- 238000004519 manufacturing process Methods 0.000 description 4
- 238000006116 polymerization reaction Methods 0.000 description 4
- 230000035484 reaction time Effects 0.000 description 4
- 230000004544 DNA amplification Effects 0.000 description 3
- 238000000137 annealing Methods 0.000 description 3
- 230000007257 malfunction Effects 0.000 description 3
- 238000005192 partition Methods 0.000 description 3
- RYGMFSIKBFXOCR-UHFFFAOYSA-N Copper Chemical compound [Cu] RYGMFSIKBFXOCR-UHFFFAOYSA-N 0.000 description 2
- 230000003321 amplification Effects 0.000 description 2
- 239000007795 chemical reaction product Substances 0.000 description 2
- 238000001816 cooling Methods 0.000 description 2
- 229910052802 copper Inorganic materials 0.000 description 2
- 239000010949 copper Substances 0.000 description 2
- 238000004925 denaturation Methods 0.000 description 2
- 230000036425 denaturation Effects 0.000 description 2
- 238000001962 electrophoresis Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 229910052736 halogen Inorganic materials 0.000 description 2
- 238000003199 nucleic acid amplification method Methods 0.000 description 2
- 230000003287 optical effect Effects 0.000 description 2
- 229910052721 tungsten Inorganic materials 0.000 description 2
- 239000010937 tungsten Substances 0.000 description 2
- -1 tungsten halogen Chemical class 0.000 description 2
- 229910052724 xenon Inorganic materials 0.000 description 2
- FHNFHKCVQCLJFQ-UHFFFAOYSA-N xenon atom Chemical compound [Xe] FHNFHKCVQCLJFQ-UHFFFAOYSA-N 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000010367 cloning Methods 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 238000001917 fluorescence detection Methods 0.000 description 1
- 239000007850 fluorescent dye Substances 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000002844 melting Methods 0.000 description 1
- 230000008018 melting Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 108090000623 proteins and genes Proteins 0.000 description 1
- 238000004445 quantitative analysis Methods 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
Images
Classifications
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- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L7/00—Heating or cooling apparatus; Heat insulating devices
- B01L7/52—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples
- B01L7/525—Heating or cooling apparatus; Heat insulating devices with provision for submitting samples to a predetermined sequence of different temperatures, e.g. for treating nucleic acid samples with physical movement of samples between temperature zones
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12M—APPARATUS FOR ENZYMOLOGY OR MICROBIOLOGY; APPARATUS FOR CULTURING MICROORGANISMS FOR PRODUCING BIOMASS, FOR GROWING CELLS OR FOR OBTAINING FERMENTATION OR METABOLIC PRODUCTS, i.e. BIOREACTORS OR FERMENTERS
- C12M1/00—Apparatus for enzymology or microbiology
- C12M1/36—Apparatus for enzymology or microbiology including condition or time responsive control, e.g. automatically controlled fermentors
- C12M1/38—Temperature-responsive control
-
- 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
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2300/00—Additional constructional details
- B01L2300/08—Geometry, shape and general structure
- B01L2300/0832—Geometry, shape and general structure cylindrical, tube shaped
- B01L2300/0838—Capillaries
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2300/00—Additional constructional details
- B01L2300/08—Geometry, shape and general structure
- B01L2300/0861—Configuration of multiple channels and/or chambers in a single devices
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L2300/00—Additional constructional details
- B01L2300/18—Means for temperature control
- B01L2300/1805—Conductive heating, heat from thermostatted solids is conducted to receptacles, e.g. heating plates, blocks
-
- 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
- G01N21/645—Specially adapted constructive features of fluorimeters
- G01N2021/6463—Optics
- G01N2021/6467—Axial flow and illumination
Definitions
- the present invention relates to a thermal cycling reaction block and a continuous real-time monitoring apparatus using the same, and more particularly, to a thermal cycling reaction block which is capable of heating or cooling samples at different temperatures so as to generate a polymerase chain reaction (PCR) and allow the PCR to be monitored in real-time, and a continuous real-time monitoring apparatus using the same.
- PCR polymerase chain reaction
- a polymerase chain reaction is a method of amplifying DNA by multiple synthesis of a selected region of the DNA, thereby producing a large amount of DNAby cloning a very small amount of the DNA.
- the PCR generally includes denaturation, primer annealing and DNA polymerization processes.
- the real-time PCR is a technology which allows monitoring of a reaction state in real-time by measuring an intensity of fluorescence showing the level of DNA amplification at every cyclein a status that a reaction product in a gel is not separated by electrophoresis. Therefore, in the real-time PCR, there are some advantages in that precise quantitative analysis is allowed, and it is possible to simply and rapidly perform the analysis without the electrophoresis, and also there is less risk of contamination.
- a real-time PCR apparatus includes a thermal cycler for PCR and a fluorometer for detecting fluorescence of a reaction product.
- a conventional real-time PCR apparatus is comprised of a thermoelectric element, a thermal block for transferring heat to a reaction tube in which a sample is received, a light source for irradiating excitation light to the sample in the tube, and a light receiving part for receiving the fluorescence generated from the sample.
- cooling and heating cycles are repeatedly performed by using the thermoelectric element so as to react the sample, and the excitation light is irradiated to the sample using the light source and the light receiving part at every end of each cycle, and then an amount of the fluorescence generated from the sample is measured so as to display the progress of the PCR in real-time.
- a heat transfer block includes four constant temperature blocks, and samples and reagents are supplied to or removed from the capillary tube using a solution supplying unit.
- the PCR is performed byrotating the heat transfer block and changing temperature transferred to the capillary tube using the above-mentioned apparatus.
- the problem in this type apparatus is that the heat transfer block should be rotated to perform the PCR, and also the reproducibility of the PCR is deteriorated since the PCR may be changed depend on a contacting level between the capillary tube and the heat transfer block.
- the capillary tube of 3.5 meters in length is wound 33 times on a copper block of 30mm in diameter, which is divided into melting, annealing and extension temperature regions.
- a reaction mixture flowed in the capillary tube is circulated once around the heating block formed of copper, each cycle of the PCR is performed.
- the capillary tube through which the PCR sample is flowed is wound on the heating block, and the capillary tube is scanned by a scanning unit having a fluorescence detector.
- the scanning unit is a means for irradiating light to the capillary tube wound on the heating block using a light irradiating unit and measuring an amount of fluorescence generated in the capillary tube.
- the light irradiating unit for irradiating light to the capillary tube wound on the heating block and a sensor for measuring the fluorescence generated from the capillary tube are installed at a moving stage, so that the scanning unit is linearly driven and the light is irradiated, in turn, to the capillary tube according to movement of the scanning unit. Then, the fluorescence generated from the capillary tube is measured, in turn, according to the movement of the scanning unit.
- the scanning unit in which a fluorescence detecting sensor and a light source for generating a light beam having a desired wavelength are moved at a constant speed above the heating block on which the capillary tube is wound.
- the light source and the fluorescence detecting sensor installed at the scanning are moved in an axial direction that is parallel with a central axis of the heating block on which the capillary is wound or that is cross the central axis thereof, so as to irradiate the light to the capillary tube or measure the fluorescence.
- An object of the present invention is to provide a thermal cycling reaction block which provides a simple monitoring apparatus so as to continuously monitor a polymerase chain reaction (PCR) in real-time, and facilely detects a PCR, and enhances detecting accuracy of the apparatus.
- PCR polymerase chain reaction
- the present invention provides a thermal cycling reaction block including a doughnut-shaped heating blocks 10a and 10b which are formed of a hollow part 11 at a central portion thereof and divided by an insulating layer so as to respectively provide different temperatures and a capillary tube 20 through which a sample is flowed in and/or out and which is wound on the heating blocks 10a and 10b at regular intervals to pass through the hollow part 11,so that the different temperatures are transferred and thus reaction cycle is repeatedly performed.
- the heating blocks 10a and 10b further include an additional heating block 13 which surrounds the outer heating block 10b, on which the capillary tube 20 is wound, so as to be coupled with an outer side of the outer heating block 10b.
- an inserting groove 12 which has a regular size and a regular interval for partial insertion of the capillary tube 20, is formed in an outer surface of the heating blocks 10a and 10b so as to increase a contacting surface area between the heating blocks 10a and 10b and the capillary tube 20.
- the present invention provides an real-time monitoring apparatus including a thermal cycling reaction block100; a light source 110 for irradiating excitation light; a band pass filter 130 for passing the excitation light having only a desired wavelength irradiated from the light source 110; a condensing lens 140 for condensing the excitation light a beam splitter 120 which reflects the excitation light and passes fluorescence generated from a sample in a capillary tube 20; a reflecting mirror 150 which is rotatably connected with a motor 160 so as to transfer the excitation light reflected from the beam splitter 120 to the capillary tube 20 and reflect the fluorescence generated from the sample in the capillary tube 20; and a fluorescence detecting part 170 for detecting the fluorescence that is reflected by the reflecting mirror 150 and then passed through the beam splitter 120.
- a thermal cycling reaction block100 including a thermal cycling reaction block100; a light source 110 for irradiating excitation light; a band pass filter 130 for passing the excitation light having only
- the fluorescence detecting part 170 includes a fluorescence condensing lens 171 for condensing the fluorescence passing through the beam splitter 120; a fluorescence band pass filter 172 for passing the condensed fluorescence havingonly a desired wavelength; and a fluorescence detecting sensor 173 for detecting the fluorescence having the desired wavelength passing through the fluorescence band pass filter 172.
- the fluorescence detecting part 170 further includes one or more fluorescence condensing lenses 171, fluorescence band pass filters 172 and fluorescence beam splitters 174 according to a wavelength region of the fluorescence.
- the present invention provides an real-time monitoring apparatus including a thermal cycling reaction block 100 a light source 110 for irradiating excitation light; a band pass filter 130 for passing the excitation light having only a desired wavelength irradiated from the light source 110; a condensing lens 140 for condensing the excitation light; a beam splitter 120 which reflects the excitation light and passes fluorescence generated from a sample in a capillary tube 20; a reflecting mirror 150 which is rotatably connected with a motor 160a so as to transfer the excitation light reflected from the beam splitter 120 to the capillary tube 20 and reflect the fluorescence generated from the sample in the capillary tube 20; a condensing lens 141 which is positioned between the reflecting mirror 150 and the thermal cycling reaction block100 so as to condense the excitation light reflected from the reflecting mirror 150 and the fluorescence generated from a sample in a capillary tube 20; and a fluorescence detecting part 170 for detecting the fluorescence that is
- the fluorescence detecting part 170 includes a fluorescence condensing lens 171 for condensing the fluorescence passing through the beam splitter 120; a fluorescence band pass filter fixing part 175 which has one or more fluorescence band pass filters 172 for passing the fluorescence havingdifferent desired wavelengths from the condensed fluorescence; a motor 160b for rotating the fluorescence band pass filter fixing part 175; and a fluorescence detecting sensor 173 for detecting the fluorescence having the desiredwavelength passing through the fluorescence band pass filters 172.
- a fluorescence condensing lens 171 for condensing the fluorescence passing through the beam splitter 120
- a fluorescence band pass filter fixing part 175 which has one or more fluorescence band pass filters 172 for passing the fluorescence havingdifferent desired wavelengths from the condensed fluorescence
- a motor 160b for rotating the fluorescence band pass filter fixing part 175
- a fluorescence detecting sensor 173 for detecting the fluorescence having
- the real-time monitoring apparatus further includes a polarizer or a polarizer film 131 between the light source 110 and the condensing lens 140 and at a fluorescence measuring part.
- the motors 160a and 160b are a constant rotation motor for rotating at a constant speed.
- the continuous real-time monitoring apparatus of the present invention uses the fixed light source and fixed fluorescence detecting part so as to be controlled by only the motor without the movement of the light source andthe fluorescence detecting part, so that the real-time monitoring is performed at a fixed position. Therefore, it is facile to detect the amplification of the sample, and it is possible to enhance the detecting accuracy and reduce the manufacturing cost andeffort, and it is also possible to reduce the malfunction and size thereof.
- Fig. 1 is a perspective view of a heating block in accordance with an embodiment of the present invention.
- Fig. 2 is a cross-sectional perspective view of the heating block in accordance with the embodiment of the present invention.
- Fig. 3 is a cross-sectional view of the heating block in accordance with another embodiment of the present invention.
- Fig. 4 is a perspective view of a thermal cycling reaction block in accordance with the embodiment of the present invention.
- Figs. 5 and 6 are perspective views showing a schematic structure of a real-time monitoring apparatus using the thermal cycling reaction block in accordance with an embodiment of the present invention.
- Figs. 7 and 8 are schematic views showing the real-time monitoring apparatus using the thermal cycling reaction block in accordance with another embodiment of the present invention.
- heating block 11 hollow part
- capillary tube 30 insulating layer
- Fig. 1 is a perspective view of a heating block in accordance with an embodiment of the present invention
- Fig. 2 is a cross-sectional perspective view of the heating block in accordance with the embodiment of the present invention
- Fig. 3 is a cross-sectional view of the heat block in accordance with another embodiment of the present invention
- Fig. 4 is a perspective view of a thermal cycling reaction block in accordance with the embodiment of the present invention.
- a thermal cycling reaction block 100 has a hollow part 11 at a central portion thereof, and also includes heating blocks 10a and 10bwhich are divided by an insulating layer so as to respectively provide different temperatures and a capillary tube 20 through which a sample is flowed in and/or out and which is wound on the heating blocks 10a and 10b at regular intervals to pass through the hollow part 11, so that the different temperatures are transferred and thus reaction cycle is repeatedly performed.
- Atypical polymerase chain reaction includes a denaturation process at 94°C an annealing process at 45 ⁇ 67°C and a polymerization process at 72°C
- PCR Atypical polymerase chain reaction
- a real-time PCR has a tendency to remove the reaction time of the polymerization process in order to reduce time.
- the two-divided heating blocks 10a and 10b are described in the drawings of the present invention, the heating block may be further divided.
- the insulting layer 30 is formed of a material having a very low heat transferring rate to facilely maintain the different temperature in each partition of the heating blocks 10a and 10b.
- the heating blocks 10a and 10b are formed into a long hole shape, it may have various shapes such as a circular shape, an elliptical shape, a polygonal shape and a rectangular shape.
- the thermal cycling reaction block 100 of the present invention may have an additional heating block 13 which surrounds the outer heating block 10b, on which the capillary tube 20 is wound, so as to be coupled with an outer side of the outer heating block 10b.
- This is a heat transfer method from an outer side of the outer heating block 10b to an inside direction thereof when transferring the heat to the capillary tube 20 through the additional heating block 13.
- the heat is further efficiently transferred to the capillary tube 20.
- the sample is flowed in and/or out through the capillary tube 20.
- the capillary tube 20 is passed through the hollow part 11 and spirally wound on the heating blocks 10a and 10b at regular intervals so that the different temperatures are transferred to each of the heating blocks 10a and 10b and thus the reaction cycle is repeatedly performed. Therefore, while the capillary tube 20 is serially and repeatedly contacted with the heating blocks 10a and 10b having the different temperature, the PCR is performed so as to amplify gene(DNA etc).
- the reason why the capillary tube 20 is wound at regular intervals is to uniformly maintain the PCR and thus to facilely rotate at a constant angle a reflecting mirror to be described later.
- An inserting groove 12 in which a part of the capillary tube 20 is inserted may be formed in an outer surface of the heating blocks 10a and 10bso as to increase a contacting surface area between the heating blocks 10a and 10b and the capillary tube 20.
- the contacting surface area between the heating blocks 10a and 10b and the capillary tube 20 is associated with the reaction time of the PCR, the contacting surface area may be changed according to the reaction time condition.
- the reaction time may be controlled by changing a radial width of the partition of the heating blocks 10a and 10b.
- a position of the insulating layer 30 in the heating blocks 10a and 10b may be also changed according to the radial width of the heating blocks 10a and 10b.
- the thermal cycling reaction block 100 as described above is used in an real-time monitoring apparatus for measuring DNA amplification in real-time.
- Figs. 5 and 6 are perspective views showing a schematic structure of a real-time monitoring apparatus using the thermal cycling reaction block in accordance with an embodiment of the present invention.
- the real-time monitoring apparatus using the thermal cycling reaction block100 in accordance with an embodiment of the present invention includes a light source 110 for irradiating excitation light; a band pass filter 130 for passing the excitation light having only a desired wavelength irradiated from the light source 110; a condensing lens 140 for condensing the excitation light; a beam splitter 120 which reflects the excitation light and passes fluorescence generated from a sample in a capillary tube 20; a reflecting mirror 150 which is rotatably connected with a motor 160 so as totransfer the excitation light reflected from the beam splitter 120 to the capillary tube 20 and reflect the fluorescence generated from the sample in the capillary tube 20; and a fluorescencedetecting part 170 for detecting the fluorescence that is reflected by the reflecting mirror 150 and then passed through the beam splitter 120.
- a light source 110 for irradiating excitation light
- a band pass filter 130 for passing the excitation light having only a desired wavelength irradiated
- the light source 110 functions to generate the excitation light and includes a white light source such as a tungsten halogen lamp and a xenon discharge lamp, and a single-colored light source such as LED and laser. But the light source 110 is not limited to them.
- the band pass filter 130 functions to pass the excitation light having only a desired wavelength irradiated from the light source 110.
- the condensing lens 140 functions to condense the excitation light irradiated from the light source 110.
- the condensing lens 140 includes any lens which condenses the excitation light, preferably, a double convex lens.
- the beam splitter 120 functions to reflect the excitation light irradiated from the light source 110 and pass fluorescence generated from the sample in the capillary tube 20.
- the beam splitter 120 is a dichroic beam splitter.
- the excitation light reflected by the beam splitter 120 is transferred to the reflecting mirror 150, and the fluorescence passing through the beam splitter 120 is transferred to the fluorescence detecting part 170.
- the reflecting mirror 150 that the excitation light reflected by the beam splitter 120 is transferred is disposed at the hollow part 11 of the thermal cycling reaction block 100.
- the reflecting mirror 150 functions to transfer the excitation light reflected from the beam splitter 120 to the capillary tube 20 that is spirally wound in the thermal cycling reaction block 100 and also functions toreflect the fluorescence generated from the sample in the capillary tube 20 to the beam splitter 120.
- the fluorescence reflected from the reflecting mirror 150 is passed through the beam splitter 120 and then transferred to the fluorescence detecting part 170.
- the reflecting mirror 150 is connected with the motor 160 for rotating the reflecting mirror 150.
- the motor 160 functions to rotate the reflecting mirror 150 so that the excitation light is reflected to the sample in the capillary tube 20 by the reflecting mirror 150 and the fluorescence generated from the sample is reflected to the fluorescence detecting part 170.
- the motor 160 is a constant rotation motor for rotating the reflecting mirror 150 at a constant speed.
- the fluorescence detecting part 170 functions to detect the fluorescence that is reflected by the reflecting mirror 150 and then passed through the beam splitter 120, thereby estimating the DNA amplification.
- the fluorescencedetecting part 170 may include a fluorescence condensing lens 171 for condensing the fluorescence passing through the beam splitter 120, a fluorescence band pass filter 172 for passing the condensed fluorescence having only a desired wavelength, and a fluorescence detecting sensor 173 for detecting the fluorescence having the desired wavelength passing through the fluorescence band pass filter 172.
- Fig. 5 shows a status of detecting the fluorescence having one wavelength.
- the fluorescencedetecting part 170 may further include one or more fluorescence condensing lenses 171, fluorescence band pass filters 172 and fluorescence beam splitters 174 according to a wavelength region of the fluorescence.
- the fluorescence beam splitters 174a and 174b are equipped differently from each other according to a wavelength of the fluorescence to be detected.
- the fluorescence condensing lenses 171a, 171b and 171c are equipped differently from each other according to a distance between the capillary tube 20 and the fluorescence detecting sensors 173a, 173b and 173c.
- the fluorescence band pass filters 172a, 172b and 172c are also equipped differently from each other according to a wavelength of the fluorescence to be detected.
- the fluorescence beam splitters 174a and 174b are the dichroic beam splitters by which a long wavelength is passed and a short wavelength is reflected on the basis of a desired wavelength.
- the desired wavelength is changed according to fluorescent dyes.
- Figs. 7 and 8 are schematic views showing the real-time monitoring apparatus using the thermal cycling reaction block in accordance with another embodiment of the present invention.
- the real-time monitoring apparatus using the thermal cycling reaction block 100 in accordance with another embodiment of the present invention includes a light source 110 for irradiating excitation light; a band pass filter 130 for passing the excitation light having only a desired wavelength irradiated from the light source 110; a condensing lens 140 for condensing the excitation light; a beam splitter 120 which reflects the excitation light and passes fluorescence generated from a sample in a capillary tube 20; a reflecting mirror 150 which is rotatably connected with a motor 160a so as to transfer the excitation light reflected from the beam splitter 120 to the capillary tube 20 and reflect the fluorescence generated from the sample in the capillary tube 20; a condensing lens 141 which is positioned between the reflecting mirror 150 and the thermal cycling reaction block 100 so as to condense the excitation light reflected from the reflecting mirror 150 and the fluorescence generated from a sample in a capillary tube 20; and a fluorescencedetecting part 170 for detecting the fluor
- the light source 110 may include a white light source such as a tungsten halogen lamp and a xenon discharge lamp, and a single-colored light source such as LED and laser, but the light source 110 is not limited to them.
- a neutral density (ND) filter 132 may be further provided to control an intensity of the laser.
- the condensing lens 141 which is positioned between the reflecting mirror 150 and the thermal cycling reaction block 100 so as to condense the excitation light reflected from the reflecting mirror 150 and the fluorescence generated from a sample in a capillary tube 20 is an aspheric lens .
- the fluorescencedetecting part 170 may include a fluorescence condensing lens 171 for condensing the fluorescence passing through the beam splitter 120 a fluorescence band pass filter fixing part 175 which has one or more fluorescence band pass filters 172for passing the fluorescence having different desired wavelengths from the condensed fluorescence; a motor 160b for rotating the fluorescence band pass filter fixing part 175; and a fluorescence detecting sensor 173 for detecting the fluorescence having the desiredwavelength passing through the fluorescence band pass filters 172.
- the fluorescence detecting part170 has one or more fluorescence band pass filters 172 provided on the fluorescence band pass filter fixing part 175.
- the fluorescence band pass filter fixing part 175 is connected with the motor 160b to be rotated, so that the fluorescence is passed through each fluorescence band pass filter 172 provided on the fluorescence band pass filter fixing part 175 and then detected, thereby enhancing fluorescence detection and space efficiency.
- the motor 160b is a constant rotation motor for rotating the fluorescence band pass filter fixing part 175 including the fluorescence band pass filters 172 at a constant speed.
- the fluorescence detecting sensor 173 is a photo multiplier tube, and the fluorescence condensing lens 171 is an aspheric lens, but the fluorescence detecting sensor 173 and the fluorescence condensing lens 171 are not limited to them.
- the real-time monitoring apparatus of the present invention may further include a polarizer or a polarizer film 131 between the light source 110 and the condensing lens 140 and at the fluorescence measuring part.
- the real-time monitoring apparatus uses the fixed light source and fixed fluorescence detecting sensor so as to be fixed at a positioned controlled by rotationof the motor, it is facile to detect the amplification of the sample, and it is possible to enhance the detecting accuracy and reduce the manufacturing cost andeffort, and it is also possible to reduce the malfunction and size thereof.
Abstract
Description
Claims (11)
- A thermal cycling reaction block, comprising:a doughnut-shaped heating blocks 10a and 10b which are formed of a hollow part 11 at a central portion thereof and divided by an insulating layer so as to respectively provide different temperatures anda capillary tube 20 through which a sample is flowed in and/or out and which is wound on the heating blocks 10a and 10b at regular intervals to pass through the hollow part 11, so that the different temperatures are transferred and thus reaction cycle is repeatedly performed.
- The thermal cycling reaction block of claim 1, further comprising an additional heating block 13 which surrounds the outer heating block 10b, on which the capillary tube 20 is wound, so as to be coupled with an outer side of the outer heating block 10b.
- The thermal cycling reaction block of claim 1, wherein an inserting groove 12, which has a regular size and a regular interval for partial insertion of the capillary tube 20, is formed in an outer surface of the heating blocks 10a and 10b so as to increase a contacting surface area between the heating blocks 10a and 10b and the capillary tube 20.
- A real-time monitoring apparatus comprising:a thermal cycling reaction block 100 according to any one of claims 1 to 3;a light source 110 for irradiating excitation light;a band pass filter 130 for passing the excitation light having only a desired wavelength irradiated from the light source 110;a condensing lens 140 for condensing the excitation light;a beam splitter 120 which reflects the excitation light and passes fluorescence generated from a sample in a capillary tube 20;a reflecting mirror 150 which is rotatably connected with a motor 160 so as to transfer the excitation light reflected from the beam splitter 120 to the capillary tube 20 and reflect the fluorescence generated from the sample in the capillary tube 20; anda fluorescence detecting part 170 for detecting the fluorescence that is reflected by the reflecting mirror 150 and then passes through the beam splitter 120.
- The apparatus of claim 4, wherein the fluorescence detecting part 170 comprises:a fluorescence condensing lens 171 for condensing the fluorescence passing through the beam splitter 120;a fluorescence band pass filter 172 for passing only the condensed fluorescence having a desired wavelength; anda fluorescence detecting sensor 173 for detecting the fluorescence having the desired wavelength passing through the fluorescence band pass filter 172.
- The apparatus of claim 5, wherein the fluorescence detecting part 170 further comprisesone or more fluorescence condensing lenses 171, fluorescence band pass filters 172 and fluorescence beam splitters 174 according to a wavelength region of the fluorescence.
- The apparatus of claim 4, wherein the motor 160 is a constant rotation motor for rotation at a constant speed.
- A real-time monitoring apparatus comprising:a thermal cycling reaction block according to any one claims 1 to 3;a light source 110 for irradiating excitation light;a band pass filter 130 for passing only the excitation light having a desired wavelength irradiated from the light source 110;a condensing lens 140 for condensing the excitation light;a beam splitter 120 which reflects the excitation light and passes fluorescence generated from a sample in a capillary tube 20;a reflecting mirror 150 which is rotatably connected with a motor 160a so as to transfer the excitation light reflected from the beam splitter 120 to the capillary tube 20 and reflect the fluorescence generated from the sample in the capillary tube 20;a condensing lens 141 which is positioned between the reflecting mirror 150 and the thermal cycling reaction block 100 so as to condense the excitation light reflected from the reflecting mirror 150 and the fluorescence generated from a sample in a capillary tube 20; anda fluorescence detecting part 170 for detecting the fluorescence that is reflected by the reflecting mirror 150 and then passes through the beam splitter 120.
- The apparatus of claim 8, wherein the fluorescence detecting part 170 comprises:a fluorescence condensing lens 171 for condensing the fluorescence passing through the beam splitter 120;a fluorescence band pass filter fixing part 175 which has one or more fluorescence band pass filters 172 for passing the fluorescence having different desired wavelengths from the condensed fluorescence;a motor 160b for rotating the fluorescence band pass filter fixing part 175; anda fluorescence detecting sensor 173 for detecting the fluorescence having the desired wavelength passing through the fluorescence band pass filters 172.
- The apparatus of claim 8, further comprising a polarizer or a polarizer film 131 between the light source 110 and the condensing lens 140 and at a fluorescence measuring part.
- The apparatus of claim 8, wherein the motors 160a and 160b are constant rotation motors for rotation at a constant speed.
Priority Applications (7)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CA2728203A CA2728203C (en) | 2008-06-23 | 2009-06-23 | Thermal cycling reaction block and continuous real-time monitoring apparatus using the same |
JP2011516122A JP5284468B2 (en) | 2008-06-23 | 2009-06-23 | Real-time monitoring device |
US12/999,696 US9205425B2 (en) | 2008-06-23 | 2009-06-23 | Thermal cycling reaction block and continuous real-time monitoring apparatus using the same |
EP09770367.2A EP2297294B1 (en) | 2008-06-23 | 2009-06-23 | Thermal cycling reaction block and continuous real-time monitoring apparatus using the same |
BRPI0915218-0A BRPI0915218B1 (en) | 2008-06-23 | 2009-06-23 | BLOCK OF THERMAL CYCLIC REACTION AND CONTINUOUS MONITORING DEVICES IN REAL TIME AND USE OF THE SAME |
AU2009263158A AU2009263158B2 (en) | 2008-06-23 | 2009-06-23 | Thermal cycling reaction block and continuous real-time monitoring apparatus using the same |
CN2009801238103A CN102083956B (en) | 2008-06-23 | 2009-06-23 | Thermal cycling reaction block and continuous real-time monitoring apparatus using the same |
Applications Claiming Priority (4)
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KR20080059211 | 2008-06-23 | ||
KR10-2008-0059211 | 2008-06-23 | ||
KR10-2009-0053677 | 2009-06-17 | ||
KR1020090053677A KR101390250B1 (en) | 2008-06-23 | 2009-06-17 | Thermal block and Continuous Real-time Monitoring Apparatus using it |
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WO2009157695A3 WO2009157695A3 (en) | 2010-03-25 |
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PCT/KR2009/003376 WO2009157695A2 (en) | 2008-06-23 | 2009-06-23 | Thermal cycling reaction block and continuous real-time monitoring apparatus using the same |
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US (1) | US9205425B2 (en) |
EP (1) | EP2297294B1 (en) |
JP (1) | JP5284468B2 (en) |
KR (1) | KR101390250B1 (en) |
CN (1) | CN102083956B (en) |
AU (1) | AU2009263158B2 (en) |
BR (1) | BRPI0915218B1 (en) |
CA (1) | CA2728203C (en) |
WO (1) | WO2009157695A2 (en) |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN103154232A (en) * | 2010-09-17 | 2013-06-12 | 韩国食品研究院 | Non-contact heating type of gene amplification system |
CN106978328A (en) * | 2016-01-15 | 2017-07-25 | 北京酷搏科技有限公司 | Thermal cycle reaction component and the real-time detection apparatus with it |
US11207691B2 (en) | 2015-09-04 | 2021-12-28 | Life Technologies Corporation | Thermal isolation of reaction sites on a substrate |
Families Citing this family (9)
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WO2012015165A2 (en) * | 2010-07-30 | 2012-02-02 | 나노바이오시스(주) | Pcr apparatus including an optically transmissive heat block |
KR101253455B1 (en) * | 2012-06-05 | 2013-04-11 | 주식회사 진시스템 | Polymerase chain reaction apparatus |
WO2014035124A1 (en) * | 2012-08-30 | 2014-03-06 | (주) 메디센서 | Rotary pcr device and pcr chip |
CN104263634B (en) * | 2014-09-24 | 2016-08-17 | 中国科学技术大学 | A kind of streaming aggregate polymerase chain reaction circulating-heating instrument based on capillary tube and heating means |
CN104730265B (en) * | 2015-03-27 | 2016-05-11 | 华南师范大学 | Hand-held POCT streaming gene alaysis system |
CN107446811A (en) * | 2017-07-25 | 2017-12-08 | 新疆昆泰锐生物技术有限公司 | A kind of reaction instrument for PCR tubular type temperature regulating device and comprising the device |
KR102133633B1 (en) * | 2018-04-17 | 2020-07-13 | (주)로고스바이오시스템스 | A device for real-time detecting nucleic acids amplification products |
CN108642158A (en) * | 2018-06-19 | 2018-10-12 | 苏州雅睿生物技术有限公司 | A kind of PCR real-time fluorescence detection systems of multichannel point detection |
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Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2007087690A1 (en) | 2006-02-02 | 2007-08-09 | Corbett Life Science Pty Ltd | Thermocycler and sample port |
Family Cites Families (19)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US5270183A (en) | 1991-02-08 | 1993-12-14 | Beckman Research Institute Of The City Of Hope | Device and method for the automated cycling of solutions between two or more temperatures |
US5207183A (en) * | 1991-12-06 | 1993-05-04 | Martha Praschnik | Flea-rid and grooming apparatus and methods |
ATE208658T1 (en) | 1993-07-28 | 2001-11-15 | Pe Corp Ny | APPARATUS AND METHOD FOR NUCLEIC ACID DUPLICATION |
US5415839A (en) * | 1993-10-21 | 1995-05-16 | Abbott Laboratories | Apparatus and method for amplifying and detecting target nucleic acids |
AU698953B2 (en) * | 1994-04-29 | 1998-11-12 | Applied Biosystems, Llc | System for real time detection of nucleic acid amplification products |
GB9621357D0 (en) | 1996-10-12 | 1996-12-04 | Central Research Lab Ltd | Heating apparatus |
US7410793B2 (en) * | 1999-05-17 | 2008-08-12 | Applera Corporation | Optical instrument including excitation source |
US6900059B1 (en) * | 1999-11-26 | 2005-05-31 | Associates Of Cape Cod, Inc. | Reader for conducting assays |
US6930314B2 (en) * | 2000-10-27 | 2005-08-16 | Molecular Devices Corporation | Light detection device |
AUPR707101A0 (en) * | 2001-08-16 | 2001-09-06 | Corbett Research Pty Ltd | Continuous flow thermal device |
US8293471B2 (en) * | 2004-01-28 | 2012-10-23 | Marshall University Research Corporation | Apparatus and method for a continuous rapid thermal cycle system |
KR100593263B1 (en) | 2004-02-02 | 2006-06-26 | 학교법인 포항공과대학교 | High throughput device for performing continuous-flow reactions |
CA2555081A1 (en) * | 2004-02-03 | 2005-08-18 | Postech Foundation | High throughput device for performing continuous-flow reactions |
JP2005295877A (en) * | 2004-04-09 | 2005-10-27 | Taiyo Yuden Co Ltd | Method for analyzing nucleic acid, analyzer and disk for analysis |
WO2006025703A1 (en) * | 2004-09-02 | 2006-03-09 | Bioneer Corporation | Miniaturized apparatus for real-time monitoring |
CN101194021A (en) * | 2005-05-13 | 2008-06-04 | 阿普里拉股份有限公司 | Low-mass thermal cycling block |
DE102005027555B3 (en) * | 2005-06-14 | 2006-10-05 | Eppendorf Ag | Thermocycler for carrying out polymerase chain reactions, has thermostatically controlled area, in which reaction vessel is placed, lid being placed over this incorporating an optical unit adjusted using pins on base and sleeves on lid |
WO2007010803A1 (en) * | 2005-07-15 | 2007-01-25 | Olympus Corporation | Light measuring instrument |
US8124033B2 (en) * | 2006-02-17 | 2012-02-28 | Agency, Science, Technology and Research | Apparatus for regulating the temperature of a biological and/or chemical sample and method of using the same |
-
2009
- 2009-06-17 KR KR1020090053677A patent/KR101390250B1/en active IP Right Grant
- 2009-06-23 WO PCT/KR2009/003376 patent/WO2009157695A2/en active Application Filing
- 2009-06-23 CN CN2009801238103A patent/CN102083956B/en active Active
- 2009-06-23 EP EP09770367.2A patent/EP2297294B1/en active Active
- 2009-06-23 US US12/999,696 patent/US9205425B2/en not_active Expired - Fee Related
- 2009-06-23 AU AU2009263158A patent/AU2009263158B2/en not_active Ceased
- 2009-06-23 CA CA2728203A patent/CA2728203C/en active Active
- 2009-06-23 JP JP2011516122A patent/JP5284468B2/en not_active Expired - Fee Related
- 2009-06-23 BR BRPI0915218-0A patent/BRPI0915218B1/en active IP Right Grant
Patent Citations (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2007087690A1 (en) | 2006-02-02 | 2007-08-09 | Corbett Life Science Pty Ltd | Thermocycler and sample port |
Non-Patent Citations (1)
Title |
---|
See also references of EP2297294A4 |
Cited By (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN103154232A (en) * | 2010-09-17 | 2013-06-12 | 韩国食品研究院 | Non-contact heating type of gene amplification system |
CN103154232B (en) * | 2010-09-17 | 2014-10-08 | 韩国食品研究院 | Non-contact heating type of gene amplification system |
US11207691B2 (en) | 2015-09-04 | 2021-12-28 | Life Technologies Corporation | Thermal isolation of reaction sites on a substrate |
CN106978328A (en) * | 2016-01-15 | 2017-07-25 | 北京酷搏科技有限公司 | Thermal cycle reaction component and the real-time detection apparatus with it |
Also Published As
Publication number | Publication date |
---|---|
EP2297294A2 (en) | 2011-03-23 |
JP2011525373A (en) | 2011-09-22 |
BRPI0915218A2 (en) | 2019-07-02 |
KR101390250B1 (en) | 2014-05-02 |
CN102083956B (en) | 2013-06-26 |
WO2009157695A3 (en) | 2010-03-25 |
US20110159579A1 (en) | 2011-06-30 |
AU2009263158B2 (en) | 2014-07-24 |
EP2297294A4 (en) | 2016-09-07 |
CA2728203A1 (en) | 2009-12-30 |
KR20090133079A (en) | 2009-12-31 |
US9205425B2 (en) | 2015-12-08 |
CN102083956A (en) | 2011-06-01 |
JP5284468B2 (en) | 2013-09-11 |
BRPI0915218B1 (en) | 2020-10-06 |
CA2728203C (en) | 2016-08-09 |
AU2009263158A1 (en) | 2009-12-30 |
EP2297294B1 (en) | 2021-03-17 |
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