CN114177963B - Nucleic acid analyzer - Google Patents
Nucleic acid analyzer Download PDFInfo
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- CN114177963B CN114177963B CN202210036525.5A CN202210036525A CN114177963B CN 114177963 B CN114177963 B CN 114177963B CN 202210036525 A CN202210036525 A CN 202210036525A CN 114177963 B CN114177963 B CN 114177963B
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- nucleic acid
- extraction
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- 150000007523 nucleic acids Chemical class 0.000 title claims abstract description 52
- 108020004707 nucleic acids Proteins 0.000 title claims abstract description 48
- 102000039446 nucleic acids Human genes 0.000 title claims abstract description 48
- 238000000605 extraction Methods 0.000 claims abstract description 61
- 230000003321 amplification Effects 0.000 claims abstract description 53
- 238000003199 nucleic acid amplification method Methods 0.000 claims abstract description 53
- 230000007246 mechanism Effects 0.000 claims abstract description 51
- 238000001514 detection method Methods 0.000 claims abstract description 28
- 238000004458 analytical method Methods 0.000 claims abstract description 17
- 238000000034 method Methods 0.000 claims abstract description 10
- 230000008569 process Effects 0.000 claims abstract description 10
- 230000003287 optical effect Effects 0.000 claims abstract description 7
- 238000007789 sealing Methods 0.000 claims abstract description 3
- 238000009413 insulation Methods 0.000 claims description 35
- 238000010438 heat treatment Methods 0.000 claims description 26
- 238000012408 PCR amplification Methods 0.000 description 25
- 230000005284 excitation Effects 0.000 description 12
- 238000010586 diagram Methods 0.000 description 9
- 238000001917 fluorescence detection Methods 0.000 description 6
- 239000013307 optical fiber Substances 0.000 description 6
- 239000002470 thermal conductor Substances 0.000 description 6
- 230000005540 biological transmission Effects 0.000 description 5
- 239000004020 conductor Substances 0.000 description 4
- 239000000835 fiber Substances 0.000 description 4
- 238000003752 polymerase chain reaction Methods 0.000 description 4
- 239000007788 liquid Substances 0.000 description 3
- 238000010828 elution Methods 0.000 description 2
- 238000005192 partition Methods 0.000 description 2
- 238000005057 refrigeration Methods 0.000 description 2
- 239000004065 semiconductor Substances 0.000 description 2
- 125000006850 spacer group Chemical group 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 238000000137 annealing Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 238000003745 diagnosis Methods 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 230000007613 environmental effect Effects 0.000 description 1
- 238000001506 fluorescence spectroscopy Methods 0.000 description 1
- 238000002955 isolation Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000002184 metal Substances 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000000149 penetrating effect Effects 0.000 description 1
- 238000002203 pretreatment Methods 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 238000005070 sampling Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
Classifications
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01L—CHEMICAL OR PHYSICAL LABORATORY APPARATUS FOR GENERAL USE
- B01L3/00—Containers or dishes for laboratory use, e.g. laboratory glassware; Droppers
- B01L3/50—Containers for the purpose of retaining a material to be analysed, e.g. test tubes
- B01L3/502—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures
- B01L3/5027—Containers for the purpose of retaining a material to be analysed, e.g. test tubes with fluid transport, e.g. in multi-compartment structures by integrated microfluidic structures, i.e. dimensions of channels and chambers are such that surface tension forces are important, e.g. lab-on-a-chip
-
- 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
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- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Clinical Laboratory Science (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Health & Medical Sciences (AREA)
- Analytical Chemistry (AREA)
- Hematology (AREA)
- Dispersion Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Biochemistry (AREA)
- Molecular Biology (AREA)
- Automatic Analysis And Handling Materials Therefor (AREA)
- Apparatus Associated With Microorganisms And Enzymes (AREA)
Abstract
The present invention relates to a nucleic acid analysis device comprising: the centrifugal driving mechanism is used for driving the micro-fluidic chip to rotate and comprises a rotating shaft, and the rotating shaft is used for being connected with the micro-fluidic chip; the extraction temperature control mechanism comprises a top temperature control unit and a bottom temperature control unit which are positioned at two sides of the microfluidic chip; the amplification temperature control mechanism is positioned beside the extraction temperature control mechanism and comprises an upper temperature control module and a lower temperature control module, wherein the upper temperature control module and the lower temperature control module are used for sealing the amplification cavity and adjusting the temperature of the amplification cavity when the amplification cavity performs amplification reaction; and the detection mechanism forms an optical detection channel with the amplification cavity through the amplification temperature control module. The nucleic acid analysis device controls the temperature of different extraction temperature areas on the microfluidic chip, independently controls the temperature of the amplification cavity, reduces the temperature influence among different areas of the microfluidic chip, and is convenient and quick in detection process.
Description
Technical Field
The present invention relates to the field of biomedical technology, and in particular, to a nucleic acid analysis device.
Background
Nucleic acid detection is one type of molecular diagnostics. It is widely used because of its high accuracy and early diagnostic timeliness. The detection process can be divided into: sampling, nucleic acid extraction, amplification, detection and the like. The nucleic acid extraction process generally requires sequential steps of lysis, washing, and elution to remove substances present in the sample that may affect the amplification step. Each step needs to be correspondingly added with different reagents and is carried out at a certain temperature so as to finally obtain purer nucleic acid for subsequent amplification.
Nucleic acid amplification usually employs the Polymerase Chain Reaction (PCR) to amplify a nucleic acid sequence to be detected, with the purpose of allowing a minute amount of target nucleic acid, which is difficult to detect, to be amplified to a large amount and then easily detected. PCR amplification is generally performed for 40 cycles, and one cycle can be divided into three steps of denaturation, annealing and extension, and theoretically, the number of target nucleic acid fragments can be doubled after one cycle. Wherein, each step is realized by the temperature change of the PCR reaction system, namely, each step corresponds to one temperature. It is known that the key to the realization of PCR amplification is rapid and accurate temperature control. The target nucleic acid fragments are marked by fluorescent groups in a PCR amplification reaction system, and the fluorescence becomes stronger along with the amplification of the target nucleic acid fragments. The optical module is used for collecting fluorescence data in real time, so that an S-shaped amplification curve can be drawn for analysis, and further detection is completed.
Because the nucleic acid detection flow has more steps, in the existing scheme for completing the whole nucleic acid detection, auxiliary equipment such as a mixing instrument, a centrifugal machine, a pipetting gun and the like are needed besides a PCR instrument and a nucleic acid extraction instrument, so that the sample circulation is complicated, the detection flow is complex, and the cost is high.
Disclosure of Invention
Based on this, it is necessary to provide a nucleic acid analyzer against the problem of redundancy of the existing nucleic acid detecting apparatus.
A nucleic acid analysis device for analyzing a sample within a microfluidic chip, the microfluidic chip comprising an extraction region and an amplification chamber, the extraction region comprising a plurality of extraction temperature zones, comprising:
the centrifugal driving mechanism is used for driving the micro-fluidic chip to rotate and comprises a rotating shaft, and the rotating shaft is used for being connected with the micro-fluidic chip;
The extraction temperature control mechanism comprises a top temperature control unit and a bottom temperature control unit which are positioned at two sides of the microfluidic chip and are used for respectively controlling the air temperatures of different extraction temperature areas on the microfluidic chip;
The amplification temperature control mechanism is positioned beside the extraction temperature control mechanism and comprises an upper temperature control module and a lower temperature control module, wherein the upper temperature control module and the lower temperature control module are used for sealing the amplification cavity and adjusting the temperature of the amplification cavity when the amplification cavity performs amplification reaction;
and the detection mechanism forms an optical detection channel with the amplification cavity through the amplification temperature control module.
In one embodiment, the top temperature control unit includes a top thermal insulation cover and a top heating element, the top heating element is located in the top thermal insulation cover, the bottom temperature control unit includes a bottom thermal insulation cover and a bottom heating element, the bottom heating element is located in the bottom thermal insulation cover, and when the microfluidic chip is connected to the rotating shaft, the top thermal insulation cover and the bottom thermal insulation cover are not in contact with the microfluidic chip.
In one embodiment, the top temperature control unit further comprises a top fan blade structure and a top flow guide disc, wherein the top fan blade structure and the top flow guide disc are positioned in the top heat insulation cover, and the top fan blade structure is sleeved on the rotating shaft and positioned on one side of the top flow guide disc; the bottom temperature control unit further comprises a bottom fan blade structure, a bottom guide disc and an idler shaft, wherein the bottom fan blade structure, the bottom guide disc and the idler shaft are arranged in the bottom temperature insulation cover, the bottom fan blade structure is sleeved on the idler shaft and is positioned on one side of the bottom guide disc, and the idler shaft is used for being connected with the rotating shaft to synchronously rotate along with the rotating shaft.
In one embodiment, the top baffle plate and the bottom baffle plate are both provided with a through air inlet and at least one air outlet.
In one embodiment, a through air inlet is formed in the central area of the top flow guiding disc, and a through air outlet is formed in the edge position of the top flow guiding disc; the bottom flow guide disc is provided with a through central air inlet in the central area, and an inner ring air outlet and an outer ring air outlet are arranged on the bottom flow guide disc along the radial direction.
In one embodiment, the bottom baffle includes a first air distribution plate and a second air distribution plate stacked, the first air distribution plate and the second air distribution plate being rotatable relative to each other to allow one of the inner ring air outlet and the outer ring air outlet to communicate with the central air inlet.
In one embodiment, the upper temperature control module or the lower temperature control module is provided with a receiving groove, and the receiving groove is used for receiving the amplification chamber.
In one embodiment, the amplification temperature control mechanism further comprises a driving module, the driving module comprises an upper connecting rod, a lower connecting rod, a horizontal sliding block guide rail and a vertical sliding block guide rail, the horizontal sliding block guide rail and the vertical sliding block guide rail are vertically arranged, the vertical sliding block guide rail comprises an upper sliding block connected with the upper temperature control module and a lower sliding block connected with the lower temperature control module, one end of the upper connecting rod is hinged with the upper sliding block, the other end of the upper connecting rod is hinged with a sliding block on the horizontal sliding block guide rail, one end of the lower connecting rod is hinged with the lower sliding block, and the other end of the lower connecting rod is hinged with the sliding block on the horizontal sliding block guide rail.
In one embodiment, the driving module further comprises a transmission shaft and a cam sleeved on the transmission shaft, the cam is abutted with the upper temperature control module, and the upper temperature control module and the lower temperature control module can be mutually close to or mutually far away from each other in the cam rotation process.
In one embodiment, the amplifying temperature control mechanism further comprises an elastic piece, one end of the elastic piece is connected with the upper temperature control module, the other end of the elastic piece is connected with the lower temperature control module, when the upper temperature control module and the lower temperature control module are far away from each other, the elastic piece is in a deformation state, and when the upper temperature control module and the lower temperature control module are in contact with each other, the elastic piece is in an original state.
According to the nucleic acid analysis device, on one hand, the microfluidic chip is adopted, so that nucleic acid extraction, amplification and analysis can be integrated, and equipment investment is reduced. On the other hand, the temperature control mechanism is used for controlling the temperature of different extraction temperature areas on the microfluidic chip, so that the temperature influence among the extraction temperature areas can be reduced, and the amplification temperature control mechanism is used for independently controlling the temperature of an amplification cavity on the microfluidic chip, so that the temperature required by the amplification cavity can be accurately and rapidly controlled. In still another aspect, an optical detection channel can be formed between the detection mechanism and the amplification chamber, and the detection process is convenient and quick.
Drawings
FIG. 1 is a schematic diagram showing a structure of a nucleic acid analyzer according to an embodiment of the present invention.
Fig. 2 is a schematic structural diagram of a microfluidic chip according to an embodiment of the present invention.
FIG. 3 is a schematic diagram showing a structure of a centrifugal temperature control apparatus in a nucleic acid analyzer according to an embodiment of the present invention.
Fig. 4 is a cross-sectional view of fig. 3.
FIG. 5 is a cross-sectional view showing the configuration of the top fan blade and the top baffle plate in a nucleic acid analyzer according to an embodiment of the present invention.
FIG. 6 is a schematic diagram showing the structure of a bottom baffle in a nucleic acid analyzer according to an embodiment of the present invention.
FIG. 7 is a schematic diagram showing an amplification temperature control mechanism of a nucleic acid analyzer according to an embodiment of the present invention.
FIG. 8 is a schematic diagram of an amplification temperature control mechanism of a nucleic acid analyzer according to an embodiment of the present invention, in which the support frame is omitted.
FIG. 9 is a schematic diagram showing a structure of a nucleic acid analyzer according to an embodiment of the present invention at another angle.
Detailed Description
The present invention may be embodied in many other forms than described herein and similarly modified by those skilled in the art without departing from the spirit of the invention, so that the invention is not limited to the specific embodiments disclosed below.
In the description of the present invention, terms such as "vertical", "horizontal", "upper", "lower", "left", "right", "center", "longitudinal", "lateral", "length", and the like are used to indicate azimuth or positional relationships based on the azimuth or positional relationships shown in the drawings, and are used for convenience of description of the present invention and for simplification of description. A first feature "on" or "under" a second feature may be the first and second features directly contacting each other, or the first and second features may be indirectly contacting each other through an intervening medium. The terms "first," "second," and the like, are used for descriptive purposes only and are not to be construed as indicating or implying relative importance or implicitly indicating the number of technical features indicated. In the description of the present invention, the meaning of "plurality" means at least two, for example, two, three, etc., unless specifically defined otherwise.
In the present invention, the terms "mounted," "connected," "secured," and the like are to be construed broadly, unless otherwise specifically indicated and defined. When an element is referred to as being "fixed" or "disposed on" another element, it can be directly on the other element or intervening elements may also be present. When an element is referred to as being "connected" to another element, it can be directly connected to the other element or intervening elements may also be present.
Referring to fig. 1, fig. 1 shows a schematic structural diagram of a nucleic acid analysis device according to an embodiment of the present invention, and the nucleic acid analysis device according to an embodiment of the present invention can be matched with a microfluidic chip 10 to realize molecular diagnosis, such as nucleic acid detection. The microfluidic chip 10 integrates nucleic acid extraction and PCR amplification functions, and requires pretreatment such as lysis-washing and elution of a sample before performing the PCR amplification function to extract and purify nucleic acid. These pretreatment sections need to be performed at different temperatures, and the temperatures required for the pretreatment sections are often different from those required for PCR amplification. For this, the nucleic acid analysis apparatus includes a centrifugal temperature control device 20, an amplification temperature control mechanism 300, and a detection mechanism 500, the centrifugal temperature control device 20 being used to provide a liquid flow driving force to the microfluidic chip 10 and temperature control during pretreatment. The amplification temperature control mechanism 300 is used for performing temperature control on PCR amplification. The detection mechanism 500 is used for detecting a product in an amplification chamber on the microfluidic chip 10 after the microfluidic chip 10 completes PCR amplification.
Referring to fig. 2, in this embodiment, the microfluidic chip 10 is a centrifugal microfluidic chip, and is in a disc shape, and is provided with an extraction area and a PCR amplification chamber for performing pretreatment, and since the temperatures required in the steps of the extraction area are different, in this embodiment, the portions of the microfluidic chip 10 located in the extraction area are named as a first extraction temperature area a, a second extraction temperature area B, and a third extraction temperature area C according to different functions implemented, and the first extraction temperature area, the second extraction temperature area, and the third extraction temperature area are concentric annular areas. The PCR amplification chamber is denoted by reference numeral P in the figure and is located at the edge of the microfluidic chip 10. The PCR amplification chamber protrudes from the surface of the microfluidic chip 10. A spline hole is provided at the center of the microfluidic chip 10 for connection with the centrifugal temperature control device 20.
Referring to fig. 3, fig. 3 is a schematic diagram showing the structure of a centrifugal temperature control apparatus 20 in a nucleic acid analysis apparatus according to an embodiment of the invention. The centrifugal temperature control apparatus 20 includes a frame 21 and a bracket 23 on the frame 21, an extraction temperature control mechanism 100, a centrifugal drive mechanism 900, and a lifting mechanism 200. The extraction temperature control mechanism 100 includes a top temperature control unit 110 and a bottom temperature control unit 130 that are disposed at intervals in a vertical direction. The top temperature control unit 110 is connected to the frame 21, and the bottom temperature control unit 130 is connected to the lifting mechanism 200. The bracket 23 is used for pushing the microfluidic chip 10 to a set position, the lifting mechanism 200 is used for pushing the microfluidic chip 10 to be connected with the centrifugal driving mechanism 900, and the top temperature control unit 110 and the bottom temperature control unit 130 are respectively located at two sides of the microfluidic chip 10 and used for performing temperature regulation and control on different areas of the microfluidic chip 10. When the microfluidic chip 10 is mounted on the centrifugal driving mechanism 900, the top temperature control unit 110 and the bottom temperature control unit 130 located at both sides of the microfluidic chip 10 are not in contact with the microfluidic chip 10.
In the present embodiment, the top temperature control unit 110 is used to control the air temperature above the first extraction temperature region of the microfluidic chip 10, and the bottom temperature control unit 130 is used to control the air temperature above the second extraction temperature region and the air temperature above the third extraction temperature region of the microfluidic chip 10.
Referring to fig. 3, the centrifugal driving mechanism 900 includes a driving motor 920 and a rotating shaft 930, the driving motor 920 is located at the top end of the frame 21, a power output end of the driving motor 920 is connected to one end of the rotating shaft 930, and the other end of the rotating shaft 930 is used for being connected to the microfluidic chip 10. In one embodiment, a chuck is disposed at one end of the shaft 930 for connection with the microfluidic chip 10, so as to be connected with a splined hole on the microfluidic chip 10 through the chuck.
Referring to fig. 4, the top temperature control unit 110 includes a top thermal insulation cover 111 and a top heating member 120. The top thermal shield 111 is fixed to the side of the frame 21 facing away from the drive motor 920. The top thermal insulation cover 111 is hollow, the top heating element 120 is located in the top thermal insulation cover 111, and the rotating shaft 930 is partially penetrating through the top thermal insulation cover 111. The top heating element 120 is used for generating heat and conducting the heat out to heat the air in the region covered by the heat insulation cover 111, so as to heat the corresponding region on the microfluidic chip 10 connected to the rotating shaft 930. The side of the top thermal insulation cover 111 away from the top end of the frame 21 has a small gap with the microfluidic chip 10.
In this embodiment, the top heating element 120 is a spiral annular resistance wire with a radial dimension corresponding to the first temperature zone.
Referring to fig. 4, the top temperature control unit 110 further includes a top airflow circulation assembly within the top thermal shield 111, the top airflow circulation assembly including a top fan blade structure 113 and a top diaphragm 115. The top diaphragm 115 is connected to the inner wall of the top heat insulating cover 111, and divides the inner space of the top heat insulating cover 111 into an upper chamber and a lower chamber. The top heating element 120 and the top fan blade structure 113 are both located on the side of the top baffle 115 facing away from the microfluidic chip 10, i.e. both located in the upper chamber of the top thermal shield 111. The top heating element 120 surrounds the outside of the top fan blade structure 113. The top fan blade structure 113 is sleeved on the rotating shaft 930, and can synchronously rotate under the rotation of the rotating shaft 930.
For ease of viewing, fig. 5 shows a schematic cross-sectional view of top fan blade structure 113 and top diaphragm 115 in isolation. The top fan blade structure 113 includes a plurality of fan blades circumferentially arranged. The top flow guiding disc 115 is provided with a through air inlet 112 and an air outlet 114, the air inlet 112 is formed in the central area of the top flow guiding disc 115, and the air outlet 114 is formed at the edge position of the top flow guiding disc 115. Wherein the air outlets 114 are provided in a plurality, and are uniformly distributed around the central axis of the top baffle 115. The upper and lower chambers inside the top thermal shield 111 are connected through the air inlet 112 and the air outlet 114. In this embodiment, the top heating element 120 is located at the air outlet 114 of the top diaphragm 115.
Referring to fig. 4 and 5, when the top fan blade structure 113 is driven to rotate by the rotating shaft 930, a negative pressure is generated in the central area of the top fan blade structure 113, so as to cause air flow in the top heat insulation cover 111. Air flows into the air inlet 112 and out of the air outlet 114 to form a flowing air stream, which is repeatedly circulated. The circulating air flow drives the air near the top heating member 120 to flow, so that the temperature of the air in the top insulating cover 111 can be made uniform. Since the air outlet 114 is located at the periphery of the air inlet 112, that is, the air inlet 112 is located in the area surrounded by the air outlet 114, a micro gap exists between the top thermal insulation cover 111 and the microfluidic chip 10, so that a "wind short circuit" is formed between the air outlet 114 and the air inlet 112, that is, flowing gas can be restrained from flowing along the air outlet 114 and the air inlet 112, and can not diffuse, so that the air flow is ensured to circularly flow in the top thermal insulation cover 111.
Referring to fig. 4, the bottom temperature control unit 130 includes a bottom thermal insulation cover 131 and a bottom heating member 140. The bottom insulating cover 131 is hollow inside, and the bottom heating member 140 is located inside the bottom insulating cover 131. There is a small gap between the bottom thermal insulation cover 131 and the microfluidic chip 10 in the vertical direction. The bottom heating element 140 is used for heating air in the area covered by the bottom thermal insulation cover 131, so as to heat the corresponding area of the microfluidic chip 10.
The bottom heating member 140 is a spiral ring-shaped resistance wire.
Referring to fig. 4, similar to the top temperature control unit 110, the bottom temperature control unit 130 further includes a bottom airflow circulation assembly within the bottom thermal shield 131, the bottom airflow circulation assembly including a bottom fan blade structure 133, a bottom baffle 135, and an idler shaft 230. The bottom fan blade structure 133 is similar in size to the top fan blade structure 113, and the central axes of the two are on the same straight line. Bottom diaphragm 135 is structurally and functionally similar to top diaphragm 115. The axis of the idle shaft 230 is on the same line as the axis of the rotating shaft 930. The bottom fan blade structure 133 and the bottom heating element 140 are both located on the side of the bottom baffle 135 facing away from the microfluidic chip 10. The bottom heating element 140 surrounds the outer side of the bottom fan blade structure 133, and the bottom fan blade structure 133 is sleeved on the idler shaft 230. When the microfluidic chip 10 is connected to the rotating shaft 930, the idle shaft 230 is also connected to the rotating shaft 930, so that only a portion of the driving motor 920 is required to drive the idle shaft 230 and the rotating shaft 930 to rotate simultaneously, thereby driving the bottom fan blade structure 133 and the top fan blade structure 113 to rotate. In one embodiment, the idler shaft 230 is also coupled to the spindle 930 via the chuck.
When the bottom fan blade structure 133 rotates, the bottom guide disc 135 and the bottom fan blade structure 133 cooperate with each other to form a circulating air flow in the bottom heat insulation cover 131, so that the heat generated by the bottom heating element 140 is uniformly distributed in the bottom heat insulation cover 131.
When the bottom temperature control unit 130 needs to regulate at least two extraction temperature zones, for example, temperature control is performed on the second extraction temperature zone and the third extraction temperature zone of the microfluidic chip 10, air with a set temperature in the bottom thermal insulation cover 131 is required to be able to pertinently heat different extraction temperature zones. Except for the above points, the bottom diaphragm 135 and the top diaphragm 115 are different in that in the present embodiment, the bottom diaphragm 135 has a central air inlet, an inner annular air outlet and an outer annular air outlet, that is, a plurality of air outlets in the radial direction compared to the top diaphragm 115. The central air inlet is arranged in the central area of the bottom air guide disc 135, and the inner ring air outlet is closer to the center of the bottom air guide disc 135 than the outer ring air outlet. Wherein, the air inlet is normally open, and one of the inner ring air outlet and the outer ring air outlet can be selectively opened or closed. When the bottom fan blade structure 133 rotates, the bottom temperature control unit 130 has two different airflow circulation paths by controlling the opening and closing of the inner ring air outlet and the outer ring air outlet respectively.
Referring to fig. 6, in one embodiment, bottom diaphragm 135 includes first and second equally sized air distribution plates 137, 139. The first air distribution plate 137 is provided with a first air inlet 132, a first inner ring air outlet 134 and a first outer ring air outlet 136 which are communicated. The first air inlet 132 is formed in a central area of the first air distribution plate 137, and the first inner ring air outlet 134 is closer to a central point of the first air distribution plate 137 than the first outer ring air outlet 136. The first inner ring air outlets 134 and the first outer ring air outlets 136 are all provided in plurality and are uniformly provided in the circumferential direction. The second air distribution plate 139 is provided with a second air inlet 142, a second inner ring air outlet 144 and a second outer ring air outlet 146 which are communicated. The second air inlet 142 is formed in a central area of the second air distribution plate 139, and the second inner ring air outlet 144 is closer to a central point of the second air distribution plate 139 than the second outer ring air outlet 146. The second inner ring air outlet 144 and the second outer ring air outlet 146 are all provided in plurality and are uniformly arranged in the circumferential direction.
In this embodiment, the connection line between the first inner ring air outlet 134 and the central point of the first air distribution plate 137 and the connection line between the first outer ring air outlet 136 and the central point of the first air distribution plate 137 are not located on the same radial line. The second inner ring air outlet 144 is connected to the center point of the second air distribution plate 139, and the second outer ring air outlet 146 is located on the same radial line as the second air distribution plate 139.
The first air distribution plate 137 and the second air distribution plate 139 are stacked to form the bottom air guide plate 135. The first air inlet 132 and the second air inlet 142 together form a central air inlet of the bottom baffle 135. When the first air distribution disc 137 and the second air distribution disc 139 rotate relatively, the first inner ring air outlet 134 and the second inner ring air outlet 144 can be communicated, meanwhile, the first outer ring air outlet 136 and the second outer ring air outlet 146 do not have air flow passing through, and the first inner ring air outlet 134 and the second inner ring air outlet 144 communicated at this time jointly form an inner ring air outlet of the bottom guide disc 135; or the first outer ring air outlet 136 and the second outer ring air outlet 146 are communicated, meanwhile, the first inner ring air outlet 134 and the second inner ring air outlet 144 do not pass through air, and at this time, the first outer ring air outlet 136 and the second outer ring air outlet 146 which are communicated together form the outer ring air outlet of the bottom flow guiding disc 135. By rotating the first air distribution disk 137 and the second air distribution disk 139 to a set angle, the central air inlet of the bottom air distribution disk 135 is respectively communicated with the inner ring air outlet or the outer ring air outlet.
The bottom temperature control unit 130 further comprises a partition plate 138 located on the bottom baffle 135, and located on a side of the bottom baffle 135 facing the microfluidic chip 10, in cooperation with different air outlets of the bottom baffle 135. There is a small gap between the spacer plate 138 and the microfluidic chip 10. At least two partition plates 138 are respectively located between the central air inlet and the inner ring air outlet, between the inner ring air outlet and the outer ring air outlet, and between the outer ring air outlet and the side surface of the bottom flow guiding disc 135, so as to enclose different annular spaces to correspond to different extraction temperature areas on the microfluidic chip 10. The provision of different spacer plates 138 prevents the air flow having a predetermined temperature from affecting the adjacent extraction temperature zone.
It should be noted that, if the second extraction temperature region of the microfluidic chip 10 is subjected to the corresponding pre-treatment, the temperature is not affected any more, that is, the final extraction result of the nucleic acid is not affected even if the temperature of the second extraction temperature region is increased again during the heating process of the third extraction temperature region.
Referring to fig. 3, the lifting mechanism 200 includes a lifting motor 250, a lift bar 260, and an idler disk 210. The bottom temperature control unit 130 is located above the idle plate 210, and the top bar 260 is located below the idle plate 210. A sliding rail structure is arranged between the left side and the right side of the idler plate 210 and the frame 21, and the ejector rod 260 can push the idler plate 210 to move in the vertical direction relative to the frame 21 under the drive of the lifting motor 250, so that the idler shaft 230 in the bottom temperature control unit 130 can push the microfluidic chip 10 positioned in the bracket 23 to be connected with the rotating shaft 930.
Referring to FIG. 4, the lifting mechanism 200 further includes an idler shaft support 220 and a resilient assembly 240, the resilient assembly 240 including a shroud 241 and a resilient element 243, the resilient element 243 being located within the shroud 241. One end of the cover shell 241 is connected with the ejector rod 260, and the other end is sleeved on the idle shaft support 220 in a sliding way. The idle shaft supporter 220 has one end abutting against the elastic element 243 in the case 241 and the other end being embedded in the center of the idle disk 210 to be rotatably coupled with the idle shaft 230. The elastic element 243 can provide a cushioning effect therebetween when the cover 241 and the idler shaft mount 220 are relatively moved.
When the carriage 23 conveys the microfluidic chip 10 to the set position, the lifting motor 250 drives the cover shell 241 connected with the ejector rod 260 to move upwards, the idle shaft support 220 moves upwards under the support of the elastic element 243 in the cover shell 241, and thus the idle plate 210 is pushed to move upwards, and the bottom temperature control unit 130 moves upwards along with the idle plate 210. The idle shaft 230 is engaged with the microfluidic chip 10 during the upward movement, and pushes the microfluidic chip 10 to disengage from the bracket 23 until the microfluidic chip 10 is engaged with the rotating shaft 930. At this time, the rotation shaft 930, the microfluidic chip 10, and the idle shaft 230 are sequentially coupled together. When the driving motor 920 drives the rotating shaft 930 to rotate, the microfluidic chip 10 and the idle shaft 230 also synchronously rotate, so that the top fan blade structure 113 sleeved on the rotating shaft 930 and the bottom fan blade structure 133 sleeved on the idle shaft 230 also synchronously rotate. So configured, the driving force of the liquid flow can be provided to the liquid in the microfluidic chip 10 by only driving the motor 920 in one step, and the environmental temperatures in the top temperature control unit 110 and the bottom temperature control unit 130 can be adjusted.
Referring to fig. 7, the amplification temperature control mechanism 300 includes a support 350, and a driving module 370, an upper temperature control module 310 and a lower temperature control module 330 disposed on the support 350, wherein the driving module 370 is used for moving the upper temperature control module 310 and the lower temperature control module 330 towards each other to clamp two sides of a PCR amplification chamber of the microfluidic chip 10 or moving away from the PCR amplification chamber. The upper temperature control module 310 includes a first heat sink 311, a first semiconductor refrigeration sheet 312, and a first thermal conductor 313, which are sequentially stacked and connected, and the lower temperature control module 330 includes a second heat sink 331, a second semiconductor refrigeration sheet 332, and a second thermal conductor 333, which are sequentially stacked and connected. The first heat conductor 313 is provided with a receiving groove (not shown in the figure) for receiving a PCR amplification chamber protruding from the surface of the microfluidic chip. When the temperature of the PCR amplification chamber is controlled, the first thermal conductor 313 and the second thermal conductor 333 are enclosed on two sides of the PCR amplification chamber, and the PCR amplification chamber is located in the accommodating groove, so that the PCR amplification chamber is completely wrapped, thereby realizing rapid temperature adjustment in a targeted manner and saving reaction time. It can be understood that when the PCR amplification chamber protrudes from the other side surface of the microfluidic chip 10, the receiving groove is provided on the second heat conductor 333.
The first heat sink 311 and the second heat sink 331 may include a heat sink and a fan, and the first heat conductor 313 and the second heat conductor 333 may be made of metal.
For ease of viewing, the support bracket 350 of the amplification temperature control mechanism 300 is omitted from fig. 8. The drive module 370 includes a horizontal slider rail 320, a vertical slider rail 340, an upper link 375, and a lower link 376. The vertical slider rail 340 includes a vertical rail 341, an upper slider 342 and a lower slider 343 slidably connected to the vertical rail 341. The vertical guide rail 341 is fixedly arranged on the support frame 350, the upper temperature control module 310 is connected with the upper slider 342, and the lower temperature control module 330 is connected with the lower slider 343. The horizontal slider rail 320 includes a cross rail 321, and a front slider 322 slidably connected to the cross rail 321. The transverse guide rail 321 is fixedly arranged on the support frame 350, and the length direction of the transverse guide rail 321 is perpendicular to the length direction of the vertical guide rail 341. One end of the upper link 375 is hinged to the upper slider 342, the other end is hinged to the front slider 322, one end of the lower link 376 is hinged to the lower slider 343, and the other end is also hinged to the front slider 322. When the front slider 322 slides along the transverse guide rail 321, the upper link 375 is driven to drive the upper slider 342 to slide along the vertical guide rail 341, and the lower link 376 is driven to drive the lower slider 343 to slide along the vertical guide rail 341, so that the upper temperature control module 310 and the lower temperature control module 330 are close to or far from each other.
The movement stroke required by the upper and lower temperature control modules 310, 330 may be accommodated by varying the lengths of the upper and lower links 375, 376.
To smooth the movement of the upper and lower temperature control modules 310 and 330, the vertical slider rails 340 are provided in two symmetrical groups, and the upper and lower links 375 and 376 are also provided in two symmetrical groups, and accordingly, a rear slider 323 is slidably provided on the lateral rail 321 to hinge with the other groups of upper and lower links 375 and 376.
Referring to fig. 8, in the present embodiment, the amplification temperature control mechanism 300 further includes a cam mechanism and an elastic member 360, where the cam mechanism includes a transmission shaft 371 and a cam 373 sleeved on the transmission shaft 371, and the cam 373 abuts against the upper slider 342. One end of the elastic member 360 is connected to the upper temperature control module 310, and the other end is connected to the lower temperature control module 330. When the amplification temperature control mechanism 300 is not in operation, the elastic member 360 is in an undeformed state.
When the cam 373 rotates synchronously with the drive shaft 371, the upper slider 342 slides along the vertical slider rail 340, and at the same time, the upper slider 342 drives the front slider 322 to slide along the transverse rail 321 through the upper link 375, and the lower link 376 connected to the front slider 322 also drives the lower slider 343 to slide along the vertical slider rail 340. In a series of linkage processes, the upper temperature control module 310 and the lower temperature control module 330 are gradually far away or close; the elastic member 360 is also stretched as the upper temperature control module 310 and the lower temperature control module 330 are separated, and gradually recovers deformation as the upper temperature control module 310 and the lower temperature control module 330 are brought closer together.
The elastic member 360 is disposed between the upper temperature control module 310 and the lower temperature control module 330, so as to ensure that the cam 373 is always attached to the upper slider 342. In other embodiments, a roller may be further disposed between the cam 373 and the upper slider 342, and the roller may rotate relative to the upper slider 342 to reduce friction between the cam 373 and the upper slider 342.
Referring to fig. 9, the detection mechanism 500 includes an optical fiber fluorescence detector, where the optical fiber fluorescence detector includes a fixed disk 510 and a rotating disk 530 that are stacked, an excitation light optical fiber 511 and an emission light optical fiber 512 are disposed on the fixed disk 510, and an excitation light source module 531 and a fluorescence detection module 532 are disposed on the rotating disk 530. One end of the excitation light fiber 511 is coupled to the excitation light source module 531, and the other end is connected to the first thermal conductor 313 in the amplification temperature control mechanism 300. The light emitting fiber 512 has one end coupled to the fluorescence detection module 532 and the other end connected to the first thermal conductor 313 in the amplification temperature control mechanism 300. The excitation light source module 531 is used for emitting excitation light, the excitation light is projected to the PCR amplification chamber through the excitation light optical fiber 511, fluorescent materials in the PCR amplification chamber are excited to generate fluorescence, and the generated fluorescence is transmitted to the fluorescence detection module 532 through the emission light optical fiber 512 for processing to obtain a detection result.
In the present embodiment, the excitation light source modules 531 and the fluorescence detection modules 532 are provided in three groups and are uniformly arranged in the circumferential direction of the rotating disk 530. By rotating the rotating disc 530, different groups of excitation light source modules 531 and fluorescence detection modules 532, the excitation light fibers 511 and the emission light fibers 512 can form different optical detection channels, so as to improve the detection efficiency.
In the nucleic acid analysis device, the extraction temperature of the different extraction areas of the microfluidic chip 10 is controlled respectively in the process of extracting nucleic acid, so that the temperature influence between adjacent extraction areas is reduced. The application rotates the top blade structure 113 and the bottom blade structure 133 synchronously with the rotating shaft 930, and can adjust the air temperature above the extraction area of the microfluidic chip 10 by using a driving motor 920. In addition, the air is driven to form dynamic air flow in the rotation process of the top fan blade structure 113, the air flow circulates only in the top heat insulation cover 111, and similarly, the air flow formed in the rotation process of the bottom fan blade structure 133 also circulates only in the bottom heat insulation cover 131, so that the air temperatures above adjacent extraction areas can not be affected each other under the condition that the temperatures in the top heat insulation cover 111 and the bottom heat insulation cover 131 are kept balanced. When the PCR amplification cavity is subjected to amplification reaction, the PCR amplification cavity is completely wrapped by the amplification temperature control mechanism, and the reaction temperature of the PCR amplification cavity can be quickly adjusted due to the large heating area and short temperature transmission path of the PCR amplification cavity. The detection mechanism 500 of the present application is provided with a plurality of groups of optical detection channels, and all detection results can be obtained by rotating the rotating disc 530. Meanwhile, the excitation light source modules 531 and the fluorescence detection modules 532 of corresponding groups can be arranged on the rotating disk 530 according to the need, and the expansion is convenient. The nucleic acid analysis device can automatically control and complete the whole nucleic acid detection process after the microfluidic chip 10 is placed in the bracket 23, and is convenient and quick.
The technical features of the above-described embodiments may be arbitrarily combined, and all possible combinations of the technical features in the above-described embodiments are not described for brevity of description, however, as long as there is no contradiction between the combinations of the technical features, they should be considered as the scope of the description.
The above examples illustrate only a few embodiments of the invention, which are described in detail and are not to be construed as limiting the scope of the invention. It should be noted that it will be apparent to those skilled in the art that several variations and modifications can be made without departing from the spirit of the invention, which are all within the scope of the invention. Accordingly, the scope of protection of the present invention is to be determined by the appended claims.
Claims (9)
1. A nucleic acid analysis device for analyzing a sample in a microfluidic chip, the microfluidic chip comprising an extraction region and an amplification chamber, the extraction region comprising a plurality of extraction temperature zones, characterized in that a portion of the microfluidic chip located in the extraction region comprises a first extraction temperature zone, a second extraction temperature zone, and a third extraction temperature zone, the first extraction temperature zone, the second extraction temperature zone, and the third extraction temperature zone being concentric annular regions; the nucleic acid analysis device comprises:
the centrifugal driving mechanism is used for driving the micro-fluidic chip to rotate and comprises a rotating shaft, and the rotating shaft is used for being connected with the micro-fluidic chip;
The extraction temperature control mechanism comprises a top temperature control unit and a bottom temperature control unit which are positioned at two sides of the microfluidic chip and are used for respectively controlling the air temperatures of different extraction temperature areas on the microfluidic chip; the top temperature control unit is used for controlling the air temperature above the first extraction temperature zone, and the bottom temperature control unit is used for controlling the air temperature above the second extraction temperature zone and the air temperature above the third extraction temperature zone; the top temperature control unit comprises a top heat insulation cover and a top heating piece, the top heating piece is positioned in the top heat insulation cover, the bottom temperature control unit comprises a bottom heat insulation cover and a bottom heating piece, the bottom heating piece is positioned in the bottom heat insulation cover, and when the microfluidic chip is connected to the rotating shaft, the top heat insulation cover and the bottom heat insulation cover are not contacted with the microfluidic chip;
The amplification temperature control mechanism is positioned beside the extraction temperature control mechanism and comprises an upper temperature control module and a lower temperature control module, wherein the upper temperature control module and the lower temperature control module are used for sealing the amplification cavity and adjusting the temperature of the amplification cavity when the amplification cavity performs amplification reaction;
And the detection mechanism and the amplification cavity form an optical detection channel through the amplification temperature control mechanism.
2. The nucleic acid analyzer of claim 1, wherein the top temperature control unit further comprises a top fan blade structure and a top flow guide disk, wherein the top fan blade structure is positioned in the top thermal insulation cover, and the top fan blade structure is sleeved on the rotating shaft and positioned on one side of the top flow guide disk; the bottom temperature control unit further comprises a bottom fan blade structure, a bottom guide disc and an idler shaft, wherein the bottom fan blade structure, the bottom guide disc and the idler shaft are arranged in the bottom temperature insulation cover, the bottom fan blade structure is sleeved on the idler shaft and is positioned on one side of the bottom guide disc, and the idler shaft is used for being connected with the rotating shaft to synchronously rotate along with the rotating shaft.
3. The nucleic acid analysis device of claim 2, wherein the top baffle plate and the bottom baffle plate are each provided with a through air inlet and at least one air outlet.
4. The nucleic acid analyzer of claim 2, wherein the center region of the top baffle has a through air inlet, and the edge of the top baffle has a through air outlet; the bottom flow guide disc is provided with a through central air inlet in the central area, and an inner ring air outlet and an outer ring air outlet are arranged on the bottom flow guide disc along the radial direction.
5. The nucleic acid analysis device of claim 4, wherein the bottom baffle comprises a first and a second stack of air distribution plates, the first and second air distribution plates being rotatable relative to each other to place one of the inner and outer ring air outlets in communication with the central air inlet.
6. The nucleic acid analyzer according to claim 1, wherein the upper temperature control module or the lower temperature control module is provided with a receiving groove for receiving the amplification chamber.
7. The nucleic acid analyzer according to claim 1, wherein the amplification temperature control mechanism further comprises a driving module, the driving module comprises an upper connecting rod, a lower connecting rod, a horizontal slide guide rail and a vertical slide guide rail, the horizontal slide guide rail and the vertical slide guide rail are vertically arranged, the vertical slide guide rail comprises an upper slide block connected with the upper temperature control module and a lower slide block connected with the lower temperature control module, one end of the upper connecting rod is hinged with the upper slide block, the other end of the upper connecting rod is hinged with a slide block on the horizontal slide guide rail, one end of the lower connecting rod is hinged with the lower slide block, and the other end of the lower connecting rod is hinged with a slide block on the horizontal slide guide rail.
8. The nucleic acid analyzer of claim 7, wherein the driving module further comprises a driving shaft and a cam sleeved on the driving shaft, the cam is abutted against the upper temperature control module, and the upper temperature control module and the lower temperature control module can be close to each other or far away from each other in the cam rotation process.
9. The nucleic acid analyzer according to claim 8, wherein the amplification temperature control mechanism further comprises an elastic member, one end of the elastic member is connected to the upper temperature control module, the other end of the elastic member is connected to the lower temperature control module, the elastic member is in a deformed state when the upper temperature control module and the lower temperature control module are away from each other, and the elastic member is in an original state when the upper temperature control module and the lower temperature control module are in contact with each other.
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| CN114509575B (en) * | 2022-04-19 | 2022-06-14 | 天津德祥生物技术有限公司 | Microfluidic detection device |
| CN115340946B (en) * | 2022-10-19 | 2023-02-03 | 天津智善生物科技有限公司 | Amplification device for nucleic acid detection and detection method based on amplification device |
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