CN114225980A - Micro-fluidic chip for molecular cycle adsorption and continuous optical detection and method for detecting multiple nucleic acid samples by using micro-fluidic chip - Google Patents
Micro-fluidic chip for molecular cycle adsorption and continuous optical detection and method for detecting multiple nucleic acid samples by using micro-fluidic chip Download PDFInfo
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- 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
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Abstract
A micro-fluidic chip for molecular cycle adsorption and continuous optical detection comprises a substrate layer and a flow channel layer, wherein the front surface of the substrate layer is provided with an optical detection area, and the back surface of the substrate layer is attached with a temperature control heating sheet and a cooling sheet; the back of the runner layer is provided with an independent circulation runner system, the independent circulation runner system comprises a storage tank, a hot runner, a detection tank, a cooling reflux runner, a cold runner, a sample inlet pipe and a sample outlet pipe, and the storage tank-the hot runner-the detection tank-the cooling reflux runner-the cold runner-the in-chip circulation runner of the storage tank is formed in a chip. The invention can collect the biological molecules in the limited sample in the optical effective detection area fully and concentratedly, simultaneously uses the external detection light beam to irradiate the detection area, and extracts the sample detection information through the property change of the transmission or reflection light beam, thereby not only solving the problem of the adsorption efficiency of the optical detection interface to the target detection object, but also improving the optical detection efficiency and the detection precision.
Description
Technical Field
The invention relates to the field of molecular diagnosis, in particular to a micro-fluidic chip for molecular cycle adsorption and continuous optical detection and a method for detecting a polynucleic acid sample by using the micro-fluidic chip.
Background
The micro-fluidic technology is a comprehensive and crossed application technology of chemistry, materials science and micro-scale hydromechanics. Since the birth of microfluidic technology several decades ago, the application of microfluidic chips in the fields of chemistry, life science and the like is continuously widened and deepened, the development of the microfluidic chips is particularly rapid in nearly ten years, and the microfluidic chip plays a very important role in the front of scientific research and industrial application of rapid detection of biomolecules (such as viruses).
A novel detection method combining an optical detection technology (such as surface plasma cell resonance SPR, surface light enhanced Raman detection SERS and the like) and a microfluidic chip is a leading-edge method in the field of biomolecule detection. Conventionally, such methods usually involve passing a fluid carrying biomolecules through a special sensing interface, irradiating the sensing interface with laser light, heating the sensing interface to a specific temperature (T1) by light and heat to achieve characteristic adsorption of the biomolecules thereon (e.g., hybridization), detecting the biomolecules by a change in a relevant parameter of an incident light beam incident on the sensing interface, and desorbing the biomolecules by heating the sensing interface to a higher temperature (T2) after detection is completed to achieve regeneration of the sensing interface, but the microfluidic chip used in such detection methods has the following disadvantages:
1. the simple microfluidic chip is used for detection, and does not have the function of concentrating and fully adsorbing limited biomolecule samples on a sensing interface, so that the detection sensitivity of the whole system is reduced to a great extent;
2. the chip is used for generating a heating beam of photo-thermal, and the heating beam can interfere with a detection beam to a certain extent, so that the detection sensitivity and the signal-to-noise ratio are influenced;
3. due to the regeneration requirement of the sensing interface, the simple microfluidic chip can not realize continuous sample detection, and the detection efficiency of the microfluidic chip can be influenced to a certain extent in practical application.
Disclosure of Invention
The invention aims to provide a micro-fluidic chip which can separate a heating area from a detection area, can concentrate limited biomolecule samples and fully adsorb the limited biomolecule samples on a sensing interface of an optical detection area and can realize continuous optical detection of the samples and a method for continuously detecting a plurality of nucleic acid samples by using the micro-fluidic chip, so as to overcome the defects of the prior art.
The technical scheme adopted by the invention is as follows: a micro-fluidic chip for molecular cycle adsorption and continuous optical detection comprises a substrate layer and a flow channel layer, wherein the front surface of the substrate layer is provided with an optical detection area, a sensing interface is arranged on the optical detection area, and the back surface is attached with a temperature control heating sheet and a cooling sheet;
the back of the runner layer is provided with an independent circulating runner system which comprises a storage tank, a hot runner, a detection tank, a cooling return runner, a cold runner, a sample inlet pipe and a sample outlet pipe, one end of the storage tank is communicated with one end of the hot runner, the other end of the hot runner is communicated with one end of the detection tank, the other end of the detection tank is communicated with one end of the cooling return runner, the other end of the cooling return runner is communicated with one end of the cold runner, the other end of the cold runner is communicated with the other end of the storage tank, thereby forming an in-chip circulation flow channel of a storage tank, a hot runner, a detection tank, a cooling reflux flow channel, a cold runner and the storage tank, one end of the sample inlet pipe is communicated with the cold runner, the other end of the sample inlet pipe is communicated with the front surface of the runner layer, the sample inlet valve and the external sample inlet pipeline are connected, one end of the sample outlet pipe is communicated with the hot runner, and the other end of the sample outlet pipe is communicated with the front surface of the runner layer, is connected with the sample outlet valve and an external sample outlet pipeline;
the front surface of the basal layer is attached to the back surface of the flow passage layer, the position of the optical detection area is overlapped with the position of the detection groove, the position of the temperature control heating sheet is overlapped with the position of the storage groove, and the position of the cooling sheet is overlapped with the position of the cooling circulation flow passage.
The further technical scheme is as follows: the independent circulating flow channel system is provided with a plurality of sets, the sample inlet pipe is communicated with the middle section of the cold flow channel, the sample outlet pipe is communicated with the middle section of the hot flow channel, and the sample outlet pipe is higher than the sample inlet pipe.
The further technical scheme is as follows: the micro-fluidic chip is placed in an inclined or vertical fixed mode during working.
Further: the bulk of the substrate layer is a crystalline or amorphous, ceramic or glass material, such as quartz glass or the like, that is transparent to a selected wavelength of light.
Further: the optical detection area is provided with a plurality of optical detection areas, each optical detection area is provided with a sensing interface, and the sensing interface is a nanoscale film or a microarray or a film-microarray multilayer composite structure.
Further: the temperature control heating sheets are a plurality of temperature control heating bodies formed by compounding resistance type heating sheets and thermocouples.
Further: one or more of the cooling fins are semiconductor coolers.
Further: the flow channel layer is made of a polymer light-transmitting material, and the material of the flow channel layer is a transparent material which can transmit light beams in a specific wavelength range and is convenient for processing a corresponding flow channel in the modes of machining, forming, 3D printing, hot pressing, etching and the like, such as polymethyl methacrylate (PMMA) and the like.
Further: the independent circulation flow channel system is provided with a left set and a right set, the optical detection area is provided with a left set and a right set, and the temperature control heating plate is provided with a left set and a right set.
Another related technical scheme is as follows: the method for detecting the polynucleic acid sample by utilizing the microfluidic chip for molecular cycle adsorption and continuous optical detection comprises the following specific steps of:
(1) opening all valves, and injecting biological probe reagents aiming at target nucleic acid into the micro flow channels of the left and right flow channel systems respectively;
(2) closing all valves, starting a left temperature control heating sheet and a right temperature control heating sheet and a cooling sheet, controlling the temperature of the temperature control heating sheets to be 40-95 ℃, and fully modifying the biological probe molecules on a sensing interface after keeping for 1-90 minutes;
(3) stopping heating and cooling, opening all valves, respectively injecting pure water into each micro-channel for cleaning, and closing all valves for standby after the cleaning is finished;
(4) respectively preparing all samples to be detected into solutions by using pure water;
(5) opening all valves, injecting the No. 1 sample solution into the left flow channel, and discharging pure water in the left flow channel;
(6) closing all valves, starting a left temperature control heating sheet and a cooling sheet, controlling the heating temperature of the left temperature control heating sheet and the cooling sheet to be 40-70 ℃, keeping for 1-90 minutes, adjusting the front surface of a test beam to irradiate the left optical detection area during the heating process, and obtaining sample detection information through detection and analysis of a transmission beam;
(7) opening all valves, injecting the No. 2 sample solution into the right flow channel, simultaneously injecting a cleaning solution into the left flow channel, and discharging the sample solution in the left flow channel;
(8) closing all valves, starting a right temperature control heating sheet and a cooling sheet, controlling the heating temperature of the right temperature control heating sheet and the cooling sheet to be 40-70 ℃, simultaneously starting a left temperature control heating sheet, controlling the heating temperature of the left temperature control heating sheet to be 75-95 ℃, keeping for 1-90 minutes, adjusting the front side of the test light beam to irradiate the right optical detection area during the period, and obtaining the detection information of the sample 2 through the detection analysis of the transmission beam;
(9) opening all valves, injecting No. 3 sample solution into the left flow channel, discharging the cleaning solution in the left flow channel, simultaneously injecting the cleaning solution into the right flow channel, and discharging the sample solution in the right flow channel;
(10) closing all valves, starting a left temperature control heating sheet and a cooling sheet, controlling the heating temperature of the left temperature control heating sheet to be 40-70 ℃, simultaneously starting a right temperature control heating sheet, controlling the heating temperature to be 75-95 ℃, keeping for 1-90 minutes, adjusting the front of a test light beam to irradiate a left optical detection area during the period, and acquiring the detection information of a sample 3 through the detection and analysis of a transmission beam;
(11) and (5) circularly executing the step (7) to the step (10) until all the sample solution is detected.
Due to the adoption of the technical scheme, the micro-fluidic chip for molecular cycle adsorption and continuous optical detection and the method for detecting the polynucleic acid sample by using the micro-fluidic chip have the following beneficial effects:
1. the invention provides a small micro-fluidic chip integrating biological sample concentration and optical detection, which is characterized in that biological molecules in a plurality of limited biological samples are concentrated and adsorbed in real time and continuously detected optically, so that the biological molecules in the limited samples in an optically effective detection area can be fully and concentratedly collected, meanwhile, an external detection light beam is used for irradiating the detection area, and sample detection information is extracted through the property change of a transmission beam or a reflection beam of the external detection light beam.
2. The micro-fluidic chip is provided with the temperature control heating sheet, and the temperature of fluid flowing through the optical detection interface is controlled by changing different heating temperature changes of the temperature control heating sheet, so that the temperature change control requirements of biomolecules in different processes of absorption, reaction, desorption and the like of the detection interface are met, the cycle of absorption, desorption regeneration and reabsorption of the detection interface on the biomolecules is realized, and the in-chip integration and optical in-situ detection in the absorption, reaction and desorption processes are also realized.
3. Because the micro-fluidic chip is provided with a plurality of sets of independent circulating flow channel systems, and the temperature control heating sheet and the cooling sheet can respectively control the real-time temperature of the micro-fluidic chip through the independent external power supply and control system, the independent control of the interaction between hot fluid and a sensing interface in each set of flow channel system in the chip (the interaction includes but is not limited to the adsorption, conversion, desorption and the like of fluid on the sensing interface) is realized, and the synchronous implementation of the adsorption and desorption processes of biomolecules in a plurality of biological samples can be realized through the switching of the plurality of sets of flow channels in the detection light beam chip and the independent control of the injection switching and the heating temperature of the fluid in the plurality of sets of flow channels in the chip, so that the seamless connection of the concentration adsorption of the plurality of samples on the detection interface and the desorption regeneration of the detection interface is realized, and the detection efficiency is further improved.
4. The micro-fluidic chip can regulate and control the circulating flow rate of fluid in the internal flow channel under the condition of constant heating temperature of the temperature control heating plate by regulating the included angle formed between the micro-fluidic chip and the horizontal plane.
5. The micro-fluidic chip greatly reduces the manual operations of sample pretreatment, detection interface regeneration and the like, improves the speed of multi-sample optical detection, shortens the detection time, has simple structure and high functional integration level, can be continuously multiplexed to carry out seamless multi-sample continuous detection, has controllable and convenient operation detection process, meets the requirement of quick detection of biological samples in various occasions, and has huge practical application prospect.
The technical features of a microfluidic chip for molecular cycling adsorption and continuous optical detection and a method for detecting a polynucleic acid sample using the same according to the present invention will be further described with reference to the accompanying drawings and examples.
Drawings
Fig. 1 is a schematic structural diagram (front view) of a microfluidic chip for molecular cycling adsorption and continuous optical detection according to an embodiment of the present invention;
FIG. 2 is a schematic view of a lamination of a base layer and a runner layer before being attached according to a first embodiment of the present invention;
FIG. 3 is a schematic view of a substrate layer structure according to a first embodiment of the present invention;
fig. 4 is a schematic view of a flow channel layer structure according to a first embodiment of the invention.
In the figure:
1-basal layer, 2-runner layer, 3-left optical detection zone, 4-right optical detection zone, 5-left temperature control heating plate, 6-right temperature control heating plate, 7-cooling plate, 8-left sample outlet pipe, 9-left descending hot runner, 10-left ascending hot runner, 11-left detection groove, 12-left cooling circulation runner, 13-left storage groove, 14-left descending cold runner, 15-left ascending cold runner, 16-left sample inlet pipe, 17-right sample outlet pipe, 18-right descending hot runner, 19-right ascending hot runner, 20-right detection groove, 21-right cooling circulation runner, 22-right storage groove, 23-right descending cold runner, 24-right ascending cold runner and 25-right sample inlet pipe.
Detailed Description
Example one
The utility model provides a micro-fluidic chip for molecule circulation adsorbs and continuous optical detection, also can be used to the continuous real-time concentration detection of many nucleic acid samples of Surface Plasmon Resonance (SPR) optical detection, as shown in figure 1, 2, including stratum basale 1 and runner layer 2, left optics detection zone 3 and right optics detection zone 4 are established to the stratum basale front, all have prepared the sensing interface according to the needs of detection on left optics detection zone 3 and the right optics detection zone 4, and the attached left side accuse temperature heating plate 5 in the stratum basale back, right side accuse temperature heating plate 6 and semiconductor cooling fin 7 (refer to figure 3).
The micro-fluidic chip is fixed in a vertical mode of 90 degrees with the horizontal plane, a left set and a right set of independent circulation flow channel systems are arranged on the back of the flow channel layer and comprise a left storage tank, a right storage tank, a left ascending hot runner, a right ascending hot runner, a left descending hot runner, a right descending hot runner, a left detecting tank, a right detecting tank, a left cooling backflow runner, a right cooling backflow runner, a left ascending cold runner, a right ascending cold runner, a left descending cold runner, a right descending cold runner, a left sample feeding pipe, a right sample feeding pipe and a left sample discharging pipe, wherein the left sample feeding pipe and the right sample discharging pipe are arranged below the left sample feeding pipe and the right sample discharging pipe are arranged above the left sample feeding pipe and the right sample feeding pipe are arranged below the right sample feeding pipe.
As shown in fig. 4, the upper end of the left storage tank 13 is communicated with the lower end of a left ascending hot runner 10, the other end of the left ascending hot runner 10 is communicated with a left descending hot runner 9, the common end of the left ascending hot runner is communicated with the lower end of a left sample outlet pipe 8, the upper end of the left sample outlet pipe 8 is communicated with the front surface of a runner layer, a sample valve is connected and a sample outlet pipeline is connected, the lower end of the left descending hot runner 9 is communicated with the upper end of a left detection tank 11, the lower end of the left detection tank 11 is communicated with the upper end of a serpentine left cooling circulating flow channel 12, the lower end of the left cooling circulating flow channel 12 is communicated with the upper end of a left descending cold channel 14, the other end of the left descending cold channel 14 is communicated with a left ascending cold channel 15, the common end of the left sample inlet pipe 16 is communicated with the upper end of a left sample inlet pipe 16, the lower end of the left sample inlet pipe 16 is communicated with the front surface of the runner layer, a sample inlet valve is connected and an external sample inlet pipeline, and the upper end of the left ascending cold channel 15 is communicated with the lower end of the left storage tank 13, thereby forming a left storage tank 13-left ascending hot runner 10-left ascending hot runner 9-left ascending hot runner 11-left cooling circulating flow channel 12-left descending cold runner 12-left cold runner The runner 14, the left ascending cold runner 15 and the left storage tank 13 are internally provided with an internal circulating runner.
Relatively, the upper end of the right storage tank 22 is communicated with the lower end of a right uplink hot runner 19, the other end of the right uplink hot runner 19 is communicated with a right downlink hot runner 18, the common end of the right downlink hot runner 18 is communicated with the lower end of a right sample outlet pipe 17, the upper end of the right sample outlet pipe 17 is communicated with the front surface of a runner layer, a sample valve is connected and an external sample outlet pipeline, the lower end of the right downlink hot runner 18 is communicated with the upper end of a right detection tank 20, the lower end of the right detection tank 20 is communicated with the upper end of a serpentine right cooling reflux runner 21, the lower end of the right cooling reflux runner 21 is communicated with the upper end of a right downlink cold runner 23, the other end of the right downlink cold runner 23 is communicated with a right uplink cold runner 24, the common end of the right uplink cold runner is communicated with the upper end of a right sample inlet pipe 25, the lower end of the right sample inlet pipe 25 is communicated with the front surface of the runner layer, the sample inlet valve is connected and the external sample inlet pipeline, and the upper end of the right uplink cold runner 24 is communicated with the lower end of the right storage tank 22, so as to form a right uplink hot runner 19-right uplink hot runner-right downlink hot runner 18-right detection tank-right uplink hot runner 20-right downlink hot runner 21-right downlink hot runner 18-right detection tank-right downlink hot runner 20-right downlink cold runner 21-right detection tank 22 23-right ascending cold runner 24-right storage tank 22, thus forming a left and a right set of symmetrical and independent runner systems.
The basal layer front face is attached to the back face of the flow channel layer, the position of the left optical detection area 3 is overlapped with the position of the left detection groove 11, the position of the right optical detection area 4 is overlapped with the position of the right detection groove 20, the position of the left temperature control heating sheet 5 is overlapped with the position of the left storage groove 13, the position of the right temperature control heating sheet 6 is overlapped with the position of the right storage groove 22, and the position of the cooling sheet 7 is overlapped with the positions of the left cooling circulation flow channel and the right cooling circulation flow channel.
The main body of the substrate layer is made of quartz glass.
The sensing interfaces on the left and right optical detection areas are uniform gold films with the thickness of 50 nm.
The temperature control heating sheet is a temperature controllable heating body formed by compounding a resistance type heating sheet and a thermocouple.
The cooling fin is a semiconductor cooler.
The flow channel layer is polymethyl methacrylate (PMMA).
The inner volume of the single set of channels of the left and right independent circulation channel systems is 189 microliters.
And the base layer and the flow channel layer are bonded after the binding surfaces are treated by a plasma cleaner.
The working process is as follows: firstly, under the state that all valves are opened, the whole pipeline of the microfluidic chip is filled with fluid through a sample injection pipe, then all valves are closed, a flow channel is closed, the left and right sets of independent circulating flow channel systems are heated by external power supply and control systems through independent temperature control heating sheets respectively, and carry out circulating cooling through a common semiconductor cooling sheet, so that the fluid circulates in the microfluidic pipeline system under the synergistic action of density difference and gravity formed in different temperature areas, and the requirements of fluid heat convection circulation in the flow channel systems in the sheet and repeated sample flowing through a detection groove are met.
Example two
A method for detecting a polynucleic acid sample by using a microfluidic chip is a method for detecting a polynucleic acid sample by using the microfluidic chip for molecular cycle adsorption and continuous optical detection, which comprises the following specific steps:
(1) all valves are opened, and 200. mu.l of bio-probe reagent containing 0.5OD for the target nucleic acid is injected into the micro flow channels of the left and right two sets of flow channel systems, respectively.
(2) And closing all the valves, starting the left and right temperature control heating sheets and the cooling sheet, controlling the temperature at 25 ℃, 45 ℃ or 95 ℃ through the temperature control heating sheets, and fully modifying the biological probe molecules on the sensing interface after keeping for 90 minutes, 10 minutes or 1 minute.
(3) Stopping heating and cooling, opening all valves, injecting 800 microliters of pure water into the left group of micro-channels and the right group of micro-channels respectively for cleaning, and closing all valves for standby after the cleaning is finished.
(4) All samples to be tested were prepared as 200 microliters solutions, respectively, using pure water.
(5) All valves are opened, the No. 1 sample solution is injected into the left flow channel, and pure water in the left flow channel is discharged.
(6) And closing all valves, starting the left temperature control heating sheet and the cooling sheet, controlling the temperature of the fluid flowing through the left storage tank at 25 ℃, 45 ℃ or 70 ℃, keeping for 90 minutes, 5 minutes or 1 minute, adjusting the front surface of the test beam to irradiate the left optical detection area, and acquiring sample detection information through detection and analysis of the transmission beam.
(7) And opening all valves, injecting the No. 2 sample solution into the right flow channel, simultaneously injecting a cleaning solution into the left flow channel, and discharging the sample solution in the left flow channel.
(8) And closing all the valves, starting the right temperature control heating sheet and the cooling sheet, controlling the temperature of the fluid flowing through the right storage tank to be 25 ℃, 45 ℃ or 70 ℃, simultaneously starting the left temperature control heating sheet, controlling the temperature of the fluid flowing through the left storage tank to be 75 ℃, 85 ℃ or 95 ℃, keeping for 90 minutes, 5 minutes or 1 minute, adjusting the front surface of the test light beam to irradiate the right optical detection area during the period, and acquiring the detection information of the sample 2 through the detection and analysis of the transmission beam.
(9) And opening all valves, injecting the No. 3 sample solution into the left flow channel, discharging the cleaning solution in the left flow channel, simultaneously injecting the cleaning solution into the right flow channel, and discharging the sample solution in the right flow channel.
(10) And closing all the valves, starting the left temperature control heating sheet and the cooling sheet, controlling the temperature of the fluid flowing through the left storage tank to be 25 ℃, 45 ℃ or 70 ℃, simultaneously starting the right temperature control heating sheet, controlling the temperature of the fluid flowing through the right storage tank to be 75 ℃, 85 ℃ or 95 ℃, keeping for 90 minutes, 5 minutes or 1 minute, adjusting the front surface of the test light beam to irradiate the left optical detection area during the period, and acquiring the detection information of the sample 3 through the detection analysis of the transmission beam.
(11) And (5) circularly executing the step (7) to the step (10) until all the sample solution is detected.
The above embodiments are only preferred embodiments of the present invention, and the structure and method of the present invention are not limited to the forms illustrated in the above embodiments, and any modifications, equivalents and the like within the spirit and principle of the present invention should be included in the protection scope of the present invention.
Claims (10)
1. A micro-fluidic chip for molecular cycle adsorption and continuous optical detection comprises a substrate layer and a flow channel layer, and is characterized in that:
the front surface of the substrate layer is provided with an optical detection area, a sensing interface is arranged on the optical detection area, and the back surface is attached with a temperature control heating sheet and a cooling sheet;
the back of the runner layer is provided with an independent circulating runner system which comprises a storage tank, a hot runner, a detection tank, a cooling return runner, a cold runner, a sample inlet pipe and a sample outlet pipe, one end of the storage tank is communicated with one end of the hot runner, the other end of the hot runner is communicated with one end of the detection tank, the other end of the detection tank is communicated with one end of the cooling return runner, the other end of the cooling return runner is communicated with one end of the cold runner, the other end of the cold runner is communicated with the other end of the storage tank, thereby forming an in-chip circulation flow channel of a storage tank, a hot runner, a detection tank, a cooling reflux flow channel, a cold runner and the storage tank, one end of the sample inlet pipe is communicated with the cold runner, the other end of the sample inlet pipe is communicated with the front surface of the runner layer, the sample inlet valve and the external sample inlet pipeline are connected, one end of the sample outlet pipe is communicated with the hot runner, and the other end of the sample outlet pipe is communicated with the front surface of the runner layer, is connected with the sample outlet valve and an external sample outlet pipeline;
the front surface of the basal layer is attached to the back surface of the flow passage layer, the position of the optical detection area is overlapped with the position of the detection groove, the position of the temperature control heating sheet is overlapped with the position of the storage groove, and the position of the cooling sheet is overlapped with the position of the cooling circulation flow passage.
2. The microfluidic chip for molecular cycling adsorption and continuous optical detection according to claim 1, wherein: the independent circulating flow channel system is provided with a plurality of sets, the sample inlet pipe is communicated with the middle section of the cold flow channel, the sample outlet pipe is communicated with the middle section of the hot flow channel, and the sample outlet pipe is higher than the sample inlet pipe.
3. The microfluidic chip for molecular cycling adsorption and continuous optical detection according to claim 2, wherein: the micro-fluidic chip is placed in an inclined or vertical fixed mode during working.
4. The microfluidic chip for molecular cycling adsorption and continuous optical detection according to claim 2, wherein: the bulk of the substrate layer is a crystalline or amorphous, ceramic or glass material, such as quartz glass or the like, that is transparent to a selected wavelength of light.
5. The microfluidic chip for molecular cycling adsorption and continuous optical detection according to claim 2, wherein: the optical detection area is provided with a plurality of optical detection areas, each optical detection area is provided with a sensing interface, and the sensing interface is a nanoscale film or a microarray or a film-microarray multilayer composite structure.
6. The microfluidic chip for molecular cycling adsorption and continuous optical detection according to claim 2, wherein: the temperature control heating sheets are a plurality of temperature control heating bodies formed by compounding resistance type heating sheets and thermocouples.
7. The microfluidic chip for molecular cycling adsorption and continuous optical detection according to claim 2, wherein: one or more of the cooling fins are semiconductor coolers.
8. The microfluidic chip for molecular cycling adsorption and continuous optical detection according to claim 2, wherein: the flow channel layer is made of a polymer light-transmitting material, and the material of the flow channel layer is a transparent material which can transmit light beams in a specific wavelength range and is convenient for processing a corresponding flow channel in the modes of machining, forming, 3D printing, hot pressing, etching and the like, such as polymethyl methacrylate (PMMA) and the like.
9. The microfluidic chip for molecular cycling adsorption and continuous optical detection according to claim 2, wherein: the independent circulation flow channel system is provided with a left set and a right set, the optical detection area is provided with a left set and a right set, and the temperature control heating plate is provided with a left set and a right set.
10. A method for detecting a polynucleic acid sample by using a microfluidic chip is characterized by comprising the following steps: the method for detecting the multiple nucleic acid samples by using the microfluidic chip for molecular cycle adsorption and continuous optical detection as claimed in claim 9 comprises the following specific steps:
(1) opening all valves, and injecting biological probe reagents aiming at target nucleic acid into the micro flow channels of the left and right flow channel systems respectively;
(2) closing all valves, starting the left and right temperature control heating sheets and the cooling sheet, controlling the temperature of the fluid flowing through the left and right storage tanks to be 25-95 ℃ through the temperature control heating sheets, and fully modifying the biological probe molecules on a sensing interface after keeping for 1-90 minutes;
(3) stopping heating and cooling, opening all valves, respectively injecting pure water into each micro-channel for cleaning, and closing all valves for standby after the cleaning is finished;
(4) respectively preparing all samples to be detected into solutions by using pure water;
(5) opening all valves, injecting the No. 1 sample solution into the left flow channel, and discharging pure water in the left flow channel;
(6) closing all valves, starting a left temperature control heating sheet and a cooling sheet, controlling the temperature of fluid flowing through a left storage tank to be 25-70 ℃, keeping for 1-90 minutes, adjusting the front surface of a test beam to irradiate a left optical detection area during the period, and obtaining sample detection information through detection and analysis of a transmission beam;
(7) opening all valves, injecting the No. 2 sample solution into the right flow channel, simultaneously injecting a cleaning solution into the left flow channel, and discharging the sample solution in the left flow channel;
(8) closing all valves, starting a right temperature control heating sheet and a cooling sheet, controlling the temperature of fluid flowing through a right storage tank to be 25-70 ℃, simultaneously starting a left temperature control heating sheet, controlling the temperature of fluid flowing through a left storage tank to be 75-95 ℃, keeping for 1-90 minutes, adjusting the front surface of a test beam to irradiate a right optical detection area, and obtaining detection information of a sample 2 through detection and analysis of a transmission beam;
(9) opening all valves, injecting No. 3 sample solution into the left flow channel, discharging the cleaning solution in the left flow channel, simultaneously injecting the cleaning solution into the right flow channel, and discharging the sample solution in the right flow channel;
(10) closing all valves, starting a left temperature control heating sheet and a cooling sheet, controlling the temperature of fluid flowing through a left storage tank to be 25-70 ℃, simultaneously starting a right temperature control heating sheet, controlling the temperature of fluid flowing through a right storage tank to be 75-95 ℃, keeping for 1-90 minutes, adjusting the front surface of a test beam to irradiate a left optical detection area, and obtaining detection information of a sample 3 through detection and analysis of a transmission beam;
(11) and (5) circularly executing the step (7) to the step (10) until all the sample solution is detected.
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| US20030008308A1 (en) * | 2001-04-06 | 2003-01-09 | California Institute Of Technology | Nucleic acid amplification utilizing microfluidic devices |
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| CN217042644U (en) * | 2021-12-10 | 2022-07-26 | 广西科技大学 | Micro-fluidic chip for molecular cycle adsorption and continuous optical detection |
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| US20030008308A1 (en) * | 2001-04-06 | 2003-01-09 | California Institute Of Technology | Nucleic acid amplification utilizing microfluidic devices |
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| CN202072690U (en) * | 2011-05-23 | 2011-12-14 | 北京工业大学 | Polymerase chained type reactor and real-time optical array detecting device |
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