CN110296963A - A kind of fluorescence detection device and fluorescence detection method - Google Patents

Abstract

The invention belongs to the technical field of bioelectronics, and mainly provides a fluorescence detection device and a fluorescence detection method. A plurality of injected sample droplets are fused through a transparent microfluidic chip to obtain corresponding fused droplets, and the fused The droplet is driven to the optical signal detection area, and then the excitation light source is used to output excitation light to the fusion droplet, so that the fusion droplet generates a corresponding fluorescent signal under the action of the excitation light, and the fluorescence signal is passed through the fluorescence receiver Receive the fluorescent signal, convert the fluorescent signal into a corresponding data signal, output the corresponding detection result based on the data signal and preset processing conditions through the data processing system, and solve the problem that the traditional digital microfluidic platform cannot simultaneously The problem of exciting the fluorescent molecules in the liquid droplet on the electrode and collecting the fluorescent signal in the vertical direction.

Description

Fluorescence detection device and fluorescence detection method

technical field

The invention belongs to the technical field of bioelectronics, in particular to a fluorescence detection device and a fluorescence detection method.

Background technique

In biological experiments, fluorescent signals are very important results indicating tools. From one end, a certain wavelength of light is used to excite the fluorescent molecules in the droplet, and the other end uses a receiver to obtain a fluorescent signal of a specific wavelength, so as to judge whether the experimental results meet expectations. On the traditional digital microfluidic platform, the reaction solution is usually pre-added to the reservoir for 4-5 reactions by manual control. In the experiment, the single liquid is pulled out from the reservoir with electrodes, and then optically The signal detection system detects the liquid after the reaction.

However, traditional digital microfluidic platforms usually use opaque metal electrodes to drive droplets, which cannot simultaneously excite and collect fluorescence signals from fluorescent molecules in droplets on the electrodes in the vertical direction, greatly reducing fluorescence detection efficiency.

Contents of the invention

The purpose of the present invention is to provide a fluorescence detection device and a fluorescence detection method, aiming to solve the problem that traditional digital microfluidic platforms usually use opaque metal electrodes to drive droplets, and cannot detect the droplets on the electrodes in the vertical direction at the same time. Exciting and collecting fluorescent signals of fluorescent molecules in the system greatly reduces the problem of fluorescence detection efficiency.

An embodiment of the present application provides a fluorescence detection device, which includes:

excitation light source;

The transparent microfluidic chip is used to fuse the injected multiple sample droplets to obtain corresponding fusion droplets, and drive the fusion droplets to the optical signal detection area, wherein the optical signal detection area is set at On the optical path of the excitation light output by the excitation light source, the fused droplet generates a corresponding fluorescent signal under the action of the excitation light;

a fluorescence receiver, configured to receive the fluorescence signal and convert the fluorescence signal into a corresponding data signal; and

A data processing module is connected to the fluorescence receiver, and the data processing module is used to output corresponding detection results based on the data signal and preset processing conditions.

Optionally, the transparent microfluidic chip includes a stacked transparent lower plate and a transparent upper plate, wherein an electrode layer is formed on the surface of the transparent lower plate, and a storage liquid is formed on the surface of the transparent upper plate. tank, the liquid storage tank is set between the transparent lower plate and the transparent upper plate;

The electrode layer is used to drive the sample droplets in the liquid storage tank according to the received driving signal.

Optionally, the fluorescence detection device also includes:

The peristaltic pump system is used to adjust the volume of the sample droplets in the liquid storage tank according to the instruction input by the user.

Optionally, the peristaltic pump system includes a peristaltic pump, a pump tube and a suction needle connected in sequence;

The pump tube is used to store sample droplets of a preset volume;

The peristaltic pump generates corresponding air pressure according to an instruction input by the user, so as to adjust the volume of the sample droplet in the liquid storage tank through the suction.

Optionally, the fluorescence detection device also includes: a host computer, a microcontroller, a relay module, and a power drive module;

The upper computer is used to edit the power-on sequence based on user operations to obtain the control parameters of the transparent microfluidic chip;

The microcontroller is used to convert the control parameters into corresponding level signals;

The relay module is used to close the controlled terminal when the level signal acts on the control terminal, so that the power drive module, the controlled terminal of the relay module, and the transparent microfluidic chip form an electrical circuit;

The power drive module is used to provide a driving signal to the transparent microfluidic chip when the controlled terminal of the relay module is closed, so as to drive the liquid droplets on the transparent microfluidic chip to move.

Optionally, the host computer is also used to control the working voltage and power switch state of the peristaltic pump system.

The embodiment of the present application also provides a fluorescence detection method, the fluorescence detection method comprising:

A transparent microfluidic chip is used to fuse the injected multiple sample droplets to obtain corresponding fused droplets, and drive the fused droplets to the optical signal detection area; wherein the optical signal detection area is set in the On the optical path of the excitation light output by the excitation light source, the fused droplet generates a corresponding fluorescent signal under the action of the excitation light;

receiving the fluorescent signal through a fluorescent receiver, and converting the fluorescent signal into a corresponding data signal;

A corresponding detection result is output by the data processing module based on the data signal and preset processing conditions.

Optionally, the fluorescence detection method also includes:

The volume of the sample droplet in the liquid storage tank on the transparent microfluidic chip is adjusted by the peristaltic pump system according to the instruction input by the user.

Optionally, the fluorescence detection method also includes:

Obtaining the control parameters of the transparent microfluidic chip by editing the power-on sequence based on user operations through the host computer;

converting the control parameter into a corresponding level signal through a microcontroller;

When the level signal acts on the control terminal through the relay module, the controlled terminal is closed, so that the power drive module, the controlled terminal of the relay module, and the transparent microfluidic chip form an electrical circuit;

When the controlled end of the relay module is closed, the power drive module provides a driving signal to the transparent microfluidic chip, so as to drive the droplet on the transparent microfluidic chip to move.

Optionally, the fluorescence detection method also includes:

The operating voltage and power switch state of the peristaltic pump system are adjusted through the host computer.

The embodiment of the present application provides a fluorescence detection device and a fluorescence detection method. A plurality of injected sample droplets are fused through a transparent microfluidic chip according to an input driving signal to obtain corresponding fused droplets, and the fused The droplet is driven to the optical signal detection area, and then the excitation light source is used to output the excitation light to the fusion droplet to obtain the corresponding fluorescence signal, and the fluorescence signal received by the fluorescence receiver is converted into a corresponding data signal through the photoelectric conversion system , through the data processing system to output the corresponding detection results based on the data signal and preset processing conditions, it solves the problem that the traditional digital microfluidic platform cannot simultaneously excite and collect the fluorescent molecules in the droplets on the electrodes in the vertical direction Problems with fluorescent signals.

Description of drawings

FIG. 1 is a schematic structural diagram of a fluorescence detection device provided by an embodiment of the present application.

Fig. 2 is a schematic flowchart of a fluorescence detection method provided by an embodiment of the present application.

Detailed ways

In order to make the object, technical solution and advantages of the present invention clearer, the present invention will be further described in detail below in conjunction with the accompanying drawings and embodiments. It should be understood that the specific embodiments described here are only used to explain the present invention, not to limit the present invention.

Fig. 1 is a schematic structural diagram of a fluorescence detection device provided by an embodiment of the present application. Referring to Fig. 1, an embodiment of the present application provides a fluorescence detection device, and the fluorescence detection device includes:

Excitation light source 10;

The transparent microfluidic chip 20 is used to fuse a plurality of injected sample droplets to obtain corresponding fusion droplets, and drive the fusion droplets to the optical signal detection area, wherein the optical signal detection area is set On the optical path of the excitation light output by the excitation light source 10, the fused droplet generates a corresponding fluorescent signal under the action of the excitation light;

A fluorescence receiver 30, configured to receive the fluorescence signal and convert the fluorescence signal into a corresponding data signal; and

A data processing module 40 is connected to the fluorescence receiver 30, and the data processing module 40 is configured to output corresponding detection results based on the data signal and preset processing conditions.

In this embodiment, by disposing the transparent microfluidic chip 20 between the excitation light source 10 and the fluorescence receiver 30, and making the optical signal detection area set on the transparent microfluidic chip 20 be located at the area emitted by the excitation light source 10 The optical path of the excitation light, so that the transparent microfluidic chip 20 can directly turn on the excitation light source 10 to irradiate the fusion droplets after driving the fusion droplets to the optical signal detection area, and receive the fusion droplets through the fluorescence receiver 30. Fluorescent signals generated by excitation under excitation light irradiation, and converting the fluorescent signals into corresponding data signals, and finally output corresponding detection results through the data processing module 40 based on the data signals and preset processing conditions, the preset processing conditions It can be set according to the user's detection needs. For example, in the application test of using liposomes to transfer plasmids with green fluorescent protein sequences into cells, multiple sample droplets include droplets containing cells, and liposomes containing plasmids. The droplet of the above-mentioned three kinds of samples and the droplet of the cell culture medium are respectively injected through different injection holes provided on the transparent microfluidic chip 20, and the transparent microfluidic chip 20 will contain the cell according to the driving signal received. The droplets containing the liposomes containing the plasmid are fused to obtain the corresponding fusion droplets, and the fusion droplets are moved to the optical signal detection area, and then the excitation light source 10 is turned on to irradiate the blue-violet excitation light, and then pass The fluorescent receiver 30 receives the fluorescent signal generated by the excitation of the fused droplet. The preset processing condition is to compare the data signal obtained by converting the fluorescent signal with the green fluorescent signal to determine whether the green fluorescent protein is expressed in the cell, thereby To further determine whether the plasmid is successfully transfected into the cell, that is, the preset processing conditions can be the contrast fluorescent signal preset by the user, so that the fluorescent signal received by the fluorescent receiver is converted into a corresponding data signal to compare with the preset fluorescent signal Compare to determine whether the experiment was successful.

In one embodiment, the transparent microfluidic chip 20 can also separate the fusion droplets according to the driving signal input by the user, so as to separate the preset sample droplets from the fusion droplets, for example, the transparent microfluidic chip 20. According to the received driving signal, the droplets containing cells and the droplets containing liposomes containing plasmids are fused to obtain the corresponding first fusion droplets. After the cells adhere to the wall, the liquid of fresh cell culture medium can be continuously added The droplet is fused with the first fusion droplet to obtain the second fusion droplet, and at the same time, a medium droplet of the same volume is separated from the second fusion droplet, so as to ensure that the cells have fresh medium and achieve continuous culture. Purpose.

In one embodiment, the fluorescence detection device in this embodiment may also include a temperature control system, which is used to adjust the ambient temperature of the transparent microfluidic chip 20, so that the transparent microfluidic chip 20 can simultaneously To complete the continuous culture process and detection process of bacteria, the diversity detection of bacterial droplets can be completed by setting the preset processing conditions in the data processing module 40, for example, by driving the bacterial droplets to the optical signal detection area, the culture can be directly The absorbance of the bacterial droplets is detected to determine the density of the bacteria, the growth of the bacteria or the gene expression level can be judged by the fluorescence profile, and the drug resistance of the bacteria to be tested can also be detected by the absorbance of the bacteria. Bacteria droplets and droplets containing different drugs are injected from multiple injection points, and by providing corresponding driving signals to the transparent microfluidic chip 20, each droplet containing drugs is mixed with a sample containing bacteria to be tested. After the droplets are fully mixed, these droplets are transplanted into different optical signal detection areas, and the excitation light source 10 at one end emits ultraviolet light at intervals, and the fluorescent receiver 30 at the other end receives ultraviolet light, so that the data processing module 40 Calculate the absorbance of the bacterial droplet to be tested, thereby deduce the density of the bacteria in the droplet, and further judge whether the bacteria are resistant to a certain drug. Compared with the existing manual experiments, the use of the fluorescence detection device in this example can automatically control a large amount of work with strong repeatability, greatly liberating the labor of the experimenters, and can also complete drug resistance faster than manual experiments. sex detection test. In addition, compared with the existing instruments on the market that can automate the detection of drug resistance, the fluorescence detection device in this embodiment uses a very low amount of reagents for a single experiment, and can increase the amount of simultaneous detection by increasing the number of injection points. drug.

In one embodiment, the transparent microfluidic chip 20 includes a stacked transparent lower plate and a transparent upper plate, wherein an electrode layer is formed on the surface of the transparent lower plate, and an electrode layer is formed on the surface of the transparent upper plate. There is a liquid storage tank, and the liquid storage tank is arranged between the transparent lower plate and the transparent upper plate;

The electrode layer is used to drive the sample droplets in the liquid storage tank according to the received driving signal.

In this embodiment, the transparent microfluidic chip 20 includes a stacked transparent lower plate and a transparent upper plate, wherein an electrode layer is formed on the upper surface of the lower plate, and the electrode layer includes a plurality of electrodes for receiving driving Signal, each electrode is covered by a dielectric material and a hydrophobic material, the dielectric material acts as electrical insulation, and provides electrowetting force and dielectrophoretic force to drive the droplet movement when the electrode is energized. The upper plate is made of a transparent conductive material (for example, indium tin oxide glass or FTO glass), and the surfaces of the upper plate and the lower plate are coated with a superhydrophobic material, such as Teflon. In the absence of electricity, the surface of the electrode cannot be wetted. By applying a driving voltage to the electrode, the surface of the electrode that is energized will undergo electrowetting, and the droplet will move under the action of electrode electrowetting.

In this embodiment, the droplet size on the transparent microfluidic chip 20 can be adjusted by setting the number of electrodes, that is to say, a driving voltage can be applied to multiple adjacent electrodes simultaneously so that they drive a large droplet It is also possible to apply a driving voltage to a single electrode to drive a small droplet to move. The sizes of large and small droplets are controllable, large droplets can be separated into small droplets, and small droplets can be mixed into large droplets.

In one embodiment, the electrode layer on the upper surface of the lower plate can be obtained by wet etching. First, a layer of positive photoresist is evenly spread on the ITO coating of ITO glass with a homogenizer. The photoresist is heat-fixed, and then exposed to ultraviolet rays to obtain a preset electrode pattern. After the excess photoresist is washed away with a developer, the ITO glass is placed in the etching solution for 2-5 minutes for etching. After the etching, the ITO glass is cleaned with deionized water to remove the residual positive glue, so that an electrode layer with a preset electrode pattern is formed on the surface of the lower plate.

In one embodiment, the ITO glass in the above embodiment can be replaced by FTO glass, and the ITO coating is replaced by the fluorine-doped tin oxide coating on the surface of the FTO glass.

In one embodiment, a plurality of liquid storage tanks are arranged on the upper machine board, and the plurality of liquid storage tanks correspond to a plurality of sampling holes, and the user can measure the volume of the droplets in the corresponding liquid storage tanks through the sampling holes as required. adjust.

In one embodiment, the fluorescence detection device further includes:

The peristaltic pump system is used to adjust the volume of the sample droplets in the liquid storage tank according to the instruction input by the user.

In this embodiment, the user can adjust the volume of the sample droplet in the liquid storage tank provided on the transparent microfluidic chip 20 through the peristaltic pump system, for example, control the pressure difference of the peristaltic pump system according to the instruction input by the user, In this way, the volume of the sample droplet can be precisely controlled. Further, the positive and negative poles in the peristaltic pump system can be set according to the instructions input by the user to adjust the rotation direction of the peristaltic pump, so as to achieve the purpose of removing the excess sample in the storage tank. The purpose of the droplet pumping back to the reagent tube not only saves the cost of the reagent, but also avoids the decompression and booster gas pump and valve required by the traditional gas pump, saving the space occupied by the liquid nitrogen bottle.

In an embodiment, the instruction input by the user may also include a pump tube switching command, by switching pump tubes with different inner diameters in the peristaltic pump system, the purpose of adjusting the flow rate of the sample droplets output by the pump tube is achieved. In this embodiment, the pump tube can be a silicone tube. By using the high temperature resistance, low temperature resistance, oil resistance and good physiological stability of the silicone tube, errors caused by deformation of the pump tube during the working process of the peristaltic pump system can be avoided. In one embodiment, the peristaltic pump system includes a peristaltic pump, a pump tube and a suction needle connected in sequence;

The pump tube is used to store sample droplets of a preset volume;

The peristaltic pump generates corresponding air pressure according to an instruction input by the user, so as to adjust the volume of the sample droplet in the liquid storage tank through the suction.

In this embodiment, the pump tube is used to store the sample droplets, and the peristaltic pump rotates according to the instructions input by the user to generate corresponding air pressure to drive the suction to adjust the volume of the sample droplets in the storage tank. Specifically, the peristaltic pump generates corresponding air pressure to suck the air column in the pump tube, thereby controlling the movement direction of the sample droplets in the pump tube.

In one embodiment, the aspiration needle in this embodiment is in sealing connection with the liquid storage tank, which can avoid the sampling error caused by the vibration of the fluorescence detection device.

In one embodiment, a peristaltic pump can be connected to multiple pump tubes at the same time, and the multiple pump tubes correspond to multiple aspiration needles respectively, and different types of sample droplets can be stored in the multiple pump tubes. It is necessary to connect the pump tube storing the required sample droplets to the peristaltic pump at the same time. By turning on the peristaltic pump, multiple sample droplets can be input at the same time, which not only saves the number of peristaltic pumps, but also injects multiple samples at the same time. The required sample droplet can be accurately controlled to inject the sample droplet, and the experimental error can be eliminated by reducing the time difference between different sample droplet injections.

In one embodiment, the pump tube in this embodiment may include a plurality of silicone tubes with different inner diameters, so that the type of sample droplet and the output flow rate of the sample droplet can be controlled by switching the silicone tubes, so as to achieve the control of the sample addition process. For the purpose of precise control, further, the tail of the silicone tube can be set as a semi-cone, which is beneficial to the connection between the silicone tube and the aspiration needle. In one embodiment, the peristaltic pump system further includes a droplet reservoir, one end of the peristaltic pump sucks the sample droplet from the droplet reservoir, and outputs the sample droplet from the other end into the pump tube, so that the sample droplet can be passed through Adding samples to the droplet storage achieves the purpose of continuously providing sample droplets for the pump tube, and realizes the effect of continuous sample cultivation and fluorescence detection.

In this embodiment, the other end of the peristaltic pump adopts the structure of a pump tube and a suction needle, and the suction needle can extend vertically near the surface of the chip, and pressurize the peristaltic pump so that the sample droplets are ejected, and the sample droplets stick to the A liquid storage tank other than the electrodes in the droplet reaction area, the liquid storage tank can be a preset liquid storage tank electrode, so that the microliter-level droplets can respond to the dielectrophoretic effect generated by the transparent microfluidic chip 20, Specifically, since the transparent microfluidic chip 20 adopts a superhydrophobic structure, it cannot be added on the surface of the chip in a manner similar to a pipette tip. By turning on the electrodes of the liquid storage tank, the surface of the liquid storage tank will become hydrophilic, so that a small amount of sample droplets from the aspiration needle can be successfully injected into the transparent microfluidic chip 20 .

In one embodiment, the aspiration needle in this embodiment can be an injection needle, and the injection needle has a diameter difference of a hemispherical structure. Further, the peristaltic pump system in this embodiment can also include a needle for fixing the aspiration needle. The fixture structure can fix multiple aspiration needles at the same time, so as to prevent the aspiration needles from vibrating and causing errors during the process of injecting sample droplets.

Furthermore, the fixture structure in this embodiment can also be used for multiple fixed pump tubes, so as to prevent the pump tubes from sloshing when the peristaltic pump is working, resulting in errors in the injection of sample droplets.

In this embodiment, the sample droplet flows from the silicone tube to the injection needle, and the sample droplet gradually becomes larger through the inclined cone at the tip of the injection needle, and then spreads on the liquid storage tank on the transparent upper plate that the injection needle contacts , and then drive the sample droplet into the reaction area of the transparent microfluidic chip through the transparent microfluidic chip to complete the sample injection. Further, when the user needs to extract the sample droplet, the transparent microfluidic chip drives the sample droplet into the storage area. The liquid tank, and then adjust the peristaltic pump to generate negative pressure, so that the sample droplets are output into the pump tube.

In one embodiment, the fluorescence detection device further includes: a host computer, a microcontroller, a relay module, and a power drive module;

The host computer is used to edit the power-on sequence based on user operations to obtain the control parameters of the transparent microfluidic chip 20;

The microcontroller is used to convert the control parameters into corresponding level signals;

The relay module is used to close the controlled terminal when the level signal acts on the control terminal, so that the power drive module, the controlled terminal of the relay module, and the transparent microfluidic chip 20 form an electrical circuit;

The power drive module is used to provide a driving signal to the transparent microfluidic chip 20 when the controlled terminal of the relay module is closed, so as to drive the liquid droplets on the transparent microfluidic chip 20 to move.

In this embodiment, the upper computer, the microcontroller, the relay module, the power drive module and the transparent microfluidic chip 20 form a digital microfluidic system. The multiple electrodes on the chip 20 are electrically connected, and each electrode includes an electrode serial number. For example, the number of electrodes on the transparent microfluidic chip 20 can be 92, 94 or 96. The relay module includes a plurality of relay switches, the control end of each relay switch is electrically connected to the microcontroller, and the controlled end of each relay switch is electrically connected to the power drive module and is connected to the transparent microfluidic chip 20 through pogo pins. One electrode is electrically connected. The upper computer is used to edit the power-on sequence based on user operations to obtain the control parameters of the transparent microfluidic chip 20, and send the control parameters to the microcontroller, wherein the control parameters can include the target electrode serial number, single-step power-on time and single-step step interval. Therefore, the host computer is also used to edit the power-on sequence based on user operations to obtain the target electrode serial number, as well as the single-step power-on time and single-step interval time of the target electrode, and send them to the microcontroller.

In this embodiment, the host computer runs code compilation software for droplet driving, and the code compilation software can compile the target electrode serial number, single-step power-on time and single-step interval time obtained based on user operation and editing power-on sequence , get the code including power-on sequence, target electrode serial number, single-step power-on time and single-step interval time and store it in the form of TXT file. When the host computer is running, the host computer invokes the TXT file that stores the code, according to the power-on sequence in the code, the target electrode serial number, the single-step power-on time and the single-step interval time, realize the closing sequence of the relay switch in the relay module and interval, so as to send corresponding driving signals to the transparent microfluidic chip 20 .

In one embodiment, the host computer is also used to control the optical detection system composed of the excitation light source 10, the fluorescence receiver 30 and the data processing module 40, so that the optical detection system and the transparent microfluidic chip 20 are powered on at the same time, And when the transparent microfluidic chip 20 drives the fusion droplet to the optical signal detection area, the excitation light source 10 is turned on at the same time, so as to avoid errors caused by detection delay.

In one embodiment, the host computer is also used to control the working voltage and power switch state of the peristaltic pump system.

In this embodiment, the host computer is also used to control the peristaltic pump system. Specifically, the host computer can adjust the working state of the peristaltic pump system by adjusting the operating voltage and power switch state of the peristaltic pump system. For example, whether the peristaltic pump system is started is controlled by controlling the power switch state of the peristaltic pump system, and the rotation speed of the peristaltic pump is controlled by controlling the operating voltage of the peristaltic pump system, so as to complete the control of the flow rate of the sample droplets and achieve the goal of maintaining the stability of the liquid storage tank. The volume of the sample droplet in the sample is adjusted for the purpose.

In one embodiment, the upper computer may be a personal computer, such as a desktop or a notebook computer, and meanwhile, the upper computer is equipped with Labview software or an EXE program file set in the FPGA chip.

In one embodiment, the host computer can also be equipped with a human-computer interaction interface, which provides automatic operation and status monitoring functions for the transparent microfluidic chip 20, and the user can select the target electrode through the human-computer interaction interface. The TXT file stored with the code can be called through the human-computer interaction interface, and the code in the TXT file can be sent to the microcontroller, so that the transparent microfluidic chip 20 can be powered on in accordance with the power-on sequence, target electrode serial number, and single-step power-on specified in the code. The closing sequence and interval of the relay switches in the relay module are realized by the electric time and the single-step interval time. In addition, when the transparent microfluidic chip 20 is running, the user can also monitor the on-off status of each electrode, the power-on time and the execution of each step in real time through the human-computer interaction interface.

In one embodiment, the digital microfluidic system further includes a magnetic module, the magnetic module communicates with the Arduino, the transparent microfluidic chip 20 is arranged at the magnetic position of the magnetic module, and the magnetic module is used to turn the transparent The magnetic beads on the microfluidic chip 20 are separated and mixed with the sample droplets. Magnetic beads are widely used in biochemical analysis (such as immune reaction, nucleic acid detection, protein enrichment) and cell separation due to their advantages of large surface area, stable chemical properties, surface modification of other molecules, and easy manipulation in an external magnetic field. Combining liquid droplets with magnetic beads in a digital microfluidic system can further increase the throughput of the reaction and shorten the reaction time. For example, by linking antibodies or DNA to magnetic beads through chemical modification, specific interactions will be formed between target molecules or cells and magnetic beads. Under the action of an external magnetic field, that is, the magnetic module, the captured target molecules or cells are easily eluted, and then used for the next step of reaction or analysis. That is to say, the magnetic beads are adsorbed by the magnetic module and the droplet is driven by the driving voltage, so that the separation of the droplet and the magnetic bead can be realized, and when the magnetic module is not involved, the separation of the droplet and the droplet can be realized by only using the driving voltage to drive the droplet. Mixing of magnetic beads.

Figure 2 is a schematic flow chart of the fluorescence detection method provided by one embodiment of the present application, as shown in Figure 2, the fluorescence detection method in this embodiment includes:

Step S10: Use the transparent microfluidic chip 20 to fuse the injected multiple sample droplets to obtain corresponding fused droplets, and drive the fused droplets to the optical signal detection area; wherein, the optical signal detection area Set on the optical path of the excitation light output by the excitation light source 10, the fused droplet generates a corresponding fluorescent signal under the action of the excitation light;

Step S20: receiving the fluorescence signal through the fluorescence receiver 30, and converting the fluorescence signal into a corresponding data signal;

Step S30: output the corresponding detection result through the data processing module 40 based on the data signal and preset processing conditions.

In this embodiment, by disposing the transparent microfluidic chip 20 between the excitation light source 10 and the fluorescence receiver 30, and making the optical signal detection area set on the transparent microfluidic chip 20 be located at the area emitted by the excitation light source 10 The optical path of the excitation light, so that the transparent microfluidic chip 20 can directly turn on the excitation light source 10 to irradiate the fusion droplets after driving the fusion droplets to the optical signal detection area, and receive the fusion droplets through the fluorescence receiver 30. Fluorescent signals generated by excitation under excitation light irradiation, and converting the fluorescent signals into corresponding data signals, and finally output corresponding detection results through the data processing module 40 based on the data signals and preset processing conditions, the preset processing conditions It can be set according to user needs. For example, in the application test of using liposomes to transfer plasmids with green fluorescent protein sequences into cells, multiple sample droplets include droplets containing cells, liquid droplets containing liposomes containing plasmids, and liposomes containing plasmids. droplet and droplet of cell culture medium, the above-mentioned three kinds of sample droplets are respectively injected through different injection holes provided on the transparent microfluidic chip 20, and the transparent microfluidic chip 20 injects the liquid containing the cells according to the driving signal received. Droplets, droplets containing liposomes containing plasmids are fused to obtain corresponding fused droplets, and the fused droplets are moved to the optical signal detection area, and then the excitation light source 10 is turned on to irradiate blue-violet excitation light, and then received by fluorescence The device 30 receives the fluorescent signal generated by the excitation of the fusion droplet. The preset processing condition is to compare the data signal obtained by converting the fluorescent signal with the green fluorescent signal to determine whether the green fluorescent protein is expressed in the cell, so as to further determine Whether the plasmid was successfully transfected into the cells.

In one embodiment, the transparent microfluidic chip 20 can also separate the fusion droplets according to the driving signal input by the user, so as to separate the preset sample droplets from the fusion droplets, for example, the transparent microfluidic chip 20. According to the received driving signal, the droplets containing cells and the droplets containing liposomes containing plasmids are fused to obtain the corresponding first fusion droplets. After the cells adhere to the wall, the liquid of fresh cell culture medium can be continuously added The droplet is fused with the first fusion droplet to obtain the second fusion droplet, and at the same time, a medium droplet of the same volume is separated from the second fusion droplet, so as to ensure that the cells have fresh medium and achieve continuous culture. Purpose.

In one embodiment, the fluorescence detection method also includes:

Step S11: adjusting the volume of the sample droplet in the liquid storage tank on the transparent microfluidic chip 20 according to the instruction input by the user through the peristaltic pump system.

In one embodiment, the fluorescence detection method also includes:

Step S12: Obtain the control parameters of the transparent microfluidic chip by editing the power-on sequence based on user operations through the host computer;

Step S13: converting the control parameter into a corresponding level signal through a microcontroller;

Step S14: when the level signal acts on the control terminal through the relay module, the controlled terminal is closed, so that the power drive module, the controlled terminal of the relay module, and the transparent microfluidic chip 20 form an electrical circuit;

Step S15 : providing a drive signal to the transparent microfluidic chip 20 through the power drive module when the controlled end of the relay module is closed, so as to drive the droplet on the transparent microfluidic chip 20 to move.

In this embodiment, the upper computer, the microcontroller, the relay module, the power drive module and the transparent microfluidic chip 20 form a digital microfluidic system. The multiple electrodes on the chip 20 are electrically connected, and each electrode includes an electrode serial number. For example, the number of electrodes on the transparent microfluidic chip 20 can be 92, 94 or 96. The relay module includes a plurality of relay switches, the control end of each relay switch is electrically connected to the microcontroller, and the controlled end of each relay switch is electrically connected to the power drive module and is connected to the transparent microfluidic chip 20 through pogo pins. One electrode is electrically connected. The upper computer is used to edit the power-on sequence based on user operations to obtain the control parameters of the transparent microfluidic chip 20, and send the control parameters to the microcontroller, wherein the control parameters can include the target electrode serial number, single-step power-on time and single-step step interval. Therefore, the host computer is also used to edit the power-on sequence based on user operations to obtain the target electrode serial number, as well as the single-step power-on time and single-step interval time of the target electrode, and send them to the microcontroller.

In one embodiment, the fluorescence detection method also includes:

Step S111: Adjust the operating voltage and power switch state of the peristaltic pump system through the host computer.

In this embodiment, the upper computer can adjust the working state of the peristaltic pump system by adjusting the working voltage and the power switch state of the peristaltic pump system, for example, by controlling the power switch state of the peristaltic pump system to control the peristaltic pump system Whether to start, by controlling the operating voltage of the peristaltic pump system to control the rotation speed of the peristaltic pump, thereby completing the control of the flow rate of the sample droplet, and achieving the purpose of adjusting the volume of the sample droplet in the liquid storage tank.

The embodiment of the present application provides a fluorescence detection device and a fluorescence detection method, in which multiple injected sample droplets are fused through a transparent microfluidic chip 20 to obtain corresponding fused droplets, and the fused droplets are driven to The optical signal detection area, and then output excitation light to the fusion droplet through the excitation light source 10, so that the fusion droplet generates a corresponding fluorescence signal under the action of the excitation light, and receives the fluorescence signal through the fluorescence receiver 30 The fluorescent signal is converted into a corresponding data signal, and the corresponding detection result is output by the data processing system based on the data signal and preset processing conditions, which solves the problem that the traditional digital microfluidic platform cannot simultaneously The problem of exciting the fluorescent molecules in the liquid droplets on the electrode and collecting the fluorescent signals in the direction of the electrode.

The above descriptions are only preferred embodiments of the present invention, and are not intended to limit the present invention. Any modifications, equivalent replacements and improvements made within the spirit and principles of the present invention should be included in the protection of the present invention. within range.

Claims (10)

1. A fluorescence detection device, characterized in that, the fluorescence detection device comprises: excitation light source; The transparent microfluidic chip is used to fuse the injected multiple sample droplets to obtain corresponding fusion droplets, and drive the fusion droplets to the optical signal detection area, wherein the optical signal detection area is set at On the optical path of the excitation light output by the excitation light source, the fusion droplet generates a corresponding fluorescent signal under the action of the excitation light; a fluorescence receiver, configured to receive the fluorescence signal and convert the fluorescence signal into a corresponding data signal; and A data processing module is connected to the fluorescence receiver, and the data processing module is used to output corresponding detection results based on the data signal and preset processing conditions. 2. The fluorescent detection device according to claim 1, wherein the transparent microfluidic chip comprises a stacked transparent lower plate and a transparent upper plate, wherein electrodes are formed on the surface of the transparent lower plate layer, a liquid storage tank is formed on the surface of the transparent upper pole plate, and the liquid storage tank is arranged between the transparent lower pole plate and the transparent upper pole plate; The electrode layer is used to drive the sample droplets in the liquid storage tank according to the received driving signal. 3. The fluorescent detection device according to claim 2, wherein the fluorescent detection device further comprises: The peristaltic pump system is used to adjust the volume of the sample droplets in the liquid storage tank according to the instruction input by the user. 4. The fluorescent detection device according to claim 3, wherein the peristaltic pump system comprises a peristaltic pump, a pump tube and a suction needle connected in sequence; The pump tube is used to store sample droplets of a preset volume; The peristaltic pump generates corresponding air pressure according to an instruction input by the user, so as to adjust the volume of the sample droplet in the liquid storage tank through the suction. 5. The fluorescence detection device according to claim 4, wherein the fluorescence detection device further comprises: a host computer, a microcontroller, a relay module and a power drive module; The host computer is used to edit the power-on sequence based on user operations to obtain the control parameters of the transparent microfluidic chip; The microcontroller is used to convert the control parameters into corresponding level signals; The relay module is used to close the controlled terminal when the level signal acts on the control terminal, so that the power drive module, the controlled terminal of the relay module, and the transparent microfluidic chip form an electrical circuit; The power drive module is used to provide a driving signal to the transparent microfluidic chip when the controlled terminal of the relay module is closed, so as to drive the liquid droplets on the transparent microfluidic chip to move. 6 . The fluorescence detection device according to claim 5 , wherein the host computer is also used to control the working voltage and power switch state of the peristaltic pump system. 7. A fluorescence detection method, characterized in that, the fluorescence detection method comprises: A transparent microfluidic chip is used to fuse the injected multiple sample droplets to obtain corresponding fusion droplets, and drive the fusion droplets to the optical signal detection area; wherein the optical signal detection area is set at the excitation light On the optical path of the excitation light output by the light source, the fused droplet generates a corresponding fluorescent signal under the action of the excitation light; receiving the fluorescent signal through a fluorescent receiver, and converting the fluorescent signal into a corresponding data signal; A corresponding detection result is output by the data processing module based on the data signal and preset processing conditions. 8. fluorescence detection method as claimed in claim 7, is characterized in that, described fluorescence detection method also comprises: The volume of the sample droplet in the liquid storage tank on the transparent microfluidic chip is adjusted by the peristaltic pump system according to the instruction input by the user. 9. fluorescence detection method as claimed in claim 7, is characterized in that, described fluorescence detection method also comprises: Obtaining the control parameters of the transparent microfluidic chip by editing the power-on sequence based on user operations through the host computer; converting the control parameter into a corresponding level signal through a microcontroller; When the level signal acts on the control terminal through the relay module, the controlled terminal is closed, so that the power drive module, the controlled terminal of the relay module, and the transparent microfluidic chip form an electrical circuit; When the controlled end of the relay module is closed, the power drive module provides a driving signal to the transparent microfluidic chip to drive the droplet on the transparent microfluidic chip to move. 10. fluorescence detection method as claimed in claim 8, is characterized in that, described fluorescence detection method also comprises: The operating voltage and power switch state of the peristaltic pump system are adjusted through the host computer.