A microfluidic chip, device and method for selectively detecting microorganisms

Abstract

The present invention relates to a microfluidic chip, device and method for selectively detecting microorganisms. The microfluidic chip includes a sample loading hole layer, a detection hole and channel layer, and a base layer arranged in sequence from top to bottom; the detection hole and channel layer are provided with a plurality of channels arranged in a radial shape, and the ends of the plurality of channels are provided with detection holes; the detection holes are pre-buried with freeze-dried powder of a reagent for selectively detecting microorganisms. The microfluidic chip provided by the present invention is provided with a plurality of detection holes for different types of target microorganisms, and different microorganisms can be detected simultaneously. The device and method containing the microfluidic chip can realize rapid selective detection of a variety of specific microorganisms, which can greatly simplify the operation steps, and the device is portable.

Description

Technical Field

[0001] The present invention belongs to the technical field of microorganism detection, and in particular relates to a microfluidic chip, a device and a method for selectively detecting microorganisms. Background Art

[0002] Infectious diseases caused by microbial infection pose a major risk to global food safety and public health, and have become a major issue of global concern. Timely monitoring and control of microbial contamination is an important task in the food hygiene industry.

[0003] Bioluminescence methods based on adenosine triphosphate (ATP) have been widely used in the food industry and environmental inspections to monitor and evaluate the cleanliness of various environmental surfaces. ATP is a molecule found in living cells that can produce bioluminescent signals in the presence of luciferase, luciferin, magnesium ions and oxygen, and the luminescent signal produced is proportional to the amount of ATP released by the lysed bacteria. Generally speaking, the content of ATP contained in each bacterium is relatively constant, so the luminescent signal produced is proportional to the concentration of bacteria in the sample.

[0004] Compared with the traditional culture-based detection of microorganisms, the detection of microorganisms based on ATP bioluminescence has the advantages of being simple and rapid. However, this method has limited application in the selective detection and quantification of specific bacteria because it cannot distinguish between different types of bacteria or distinguish whether the measured ATP comes from beneficial or harmful bacteria. Therefore, the current methods for measuring ATP levels cannot provide detailed information for identifying target bacteria and cannot correctly assess microbial risks.

[0005] Microfluidic chips have received extensive attention in the field of rapid testing in recent years due to their small size, small reagent usage, and easy operation. In addition, the high throughput of microfluidic chips helps to achieve simultaneous detection of multiple microorganisms in a sample, which can greatly reduce the cost of testing. Summary of the invention

[0006] The present invention aims at the deficiencies of the prior art and provides a microfluidic chip, device and method for selectively detecting microorganisms based on the detection principle of ATP bioluminescence. The device and method containing the microfluidic chip for selectively detecting microorganisms can realize the selective detection of specific microorganisms, and can also realize the distinction between ATP present in the matrix of the sample to be tested and ATP in the microorganisms.

[0007] To this end, the first aspect of the present invention provides a microfluidic chip for selectively detecting microorganisms, which comprises a sample loading hole layer, a detection hole and channel layer, and a base layer arranged in sequence from top to bottom;

[0008] The detection hole and channel layer is provided with a plurality of channels arranged in a radial shape, and the ends of the plurality of channels are all provided with detection holes; freeze-dried powder of a reagent for selectively detecting microorganisms is pre-embedded in the detection holes;

[0009] The sample adding hole layer is provided with sample adding holes, and the sample adding holes are used to inject the sample to be tested into the detection holes and each channel on the channel layer through the sample adding holes, and further enter each detection hole.

[0010] In the present invention, since the detection holes are arranged at the ends of the channels, the number of the detection holes is the same as the number of the channels.

[0011] In some embodiments of the present invention, the number of the detection holes is multiple, which can achieve simultaneous detection of multiple microorganisms in the sample to be detected.

[0012] In some embodiments of the present invention, the number of the detection holes is 3 or more; more preferably, the number of the detection holes is 4 to 11.

[0013] In some specific embodiments of the present invention, the detection wells include a positive control well, a negative control well, and a plurality of (eg, 1 to 9) detection wells for different types of target microorganisms.

[0014] The present invention does not clearly limit the number of detection holes, and the number can be 11 or even more. In some specific embodiments of the present invention, the number of detection holes is 11, of which holes 1, 2, 3, 4, 6, 7, 8, and 9 are experimental holes, respectively pre-embedded with freeze-dried photothermal conversion nanomaterials (such as gold nanorods) and ATP bioluminescent substrates coated with different types of antibodies, corresponding to the detection of different target microorganisms; hole 10 is a negative control hole, pre-embedded with photothermal conversion nanomaterials (such as gold nanorods) and ATP bioluminescent substrates without antibody coating; hole 11 is a positive control hole, pre-embedded with bacterial lysis reagents and ATP bioluminescent substrates.

[0015] In some embodiments of the present invention, the reagent is a mixture of an antibody-coated photothermal conversion nanomaterial and an ATP bioluminescent substrate, and the antibody can specifically bind to the target microorganism to be detected.

[0016] In some preferred embodiments of the present invention, the mass concentration of the antibody-coated photothermal conversion nanomaterial in the mixed solution is 0.025-0.1 mg / mL, and the mass concentration of the ATP bioluminescent substrate is 3-5 mg / mL.

[0017] In some specific embodiments of the present invention, the mass concentration of the antibody-coated photothermal conversion nanomaterial in the mixed solution may be 0.025 mg / mL, and the mass concentration of the ATP bioluminescent substrate may be 5 mg / mL.

[0018] In the present invention, the preparation method of the above-mentioned mixed solution can be: 0.01 mg of the antibody-coated photothermal conversion nanomaterial is mixed with ATP bioluminescent substrate (a total of 2 mg of mixed substrate powder of luciferase, luciferin and magnesium ions), dissolved in 400 μL of buffer, and then a mixed solution including the antibody-coated photothermal conversion nanomaterial and the ATP bioluminescent substrate is obtained, wherein the mass concentration of the antibody-coated photothermal conversion nanomaterial in the mixed solution is 0.025 mg / mL, and the mass concentration of the ATP bioluminescent substrate is 5 mg / mL. The type of antibody on the photothermal conversion nanomaterial is selected according to the test requirements.

[0019] In some embodiments of the present invention, the photothermal conversion nanomaterial is a gold nanorod; preferably, the aspect ratio of the gold nanorod is 4 to 4.3, for example, 4.1.

[0020] In some embodiments of the invention, the ATP bioluminescent substrate comprises thermostable luciferase, luciferin and magnesium ions.

[0021] The present invention does not clearly limit the dosage relationship of the thermostable luciferase, luciferin and magnesium ions in the ATP bioluminescent substrate, and those skilled in the art can make routine selections as needed.

[0022] In some embodiments of the present invention, the detection hole has a diameter of 3 to 5 mm and a depth of 1 to 2 mm.

[0023] In some specific embodiments of the present invention, the detection hole has a diameter of 5 mm and a depth of 2 mm.

[0024] In other embodiments of the present invention, the channel has a width of 0.5 to 1 mm and a depth of 50 to 100 μm.

[0025] In some specific embodiments of the present invention, the channel has a width of 1 mm and a depth of 100 μm.

[0026] In some embodiments of the present invention, the diameter of the sample loading holes on the sample loading hole layer is 2-3 mm.

[0027] In some specific embodiments of the present invention, the diameter of the sample loading holes on the sample loading hole layer is 3 mm.

[0028] In some preferred embodiments of the present invention, a vent valve hole is provided on the sample loading hole layer at a position corresponding to the detection hole and the detection hole on the channel layer. The vent valve hole is used to maintain the balance of air pressure inside and outside the chip.

[0029] In some other preferred embodiments of the present invention, the diameter of the vent valve hole is 0.5-1 mm. In some specific embodiments of the present invention, the diameter of the vent valve hole is 1 mm.

[0030] In some embodiments of the present invention, the sample loading hole layer, the detection hole and channel layer, and the base layer are all made of transparent materials. In some specific embodiments of the present invention, the sample loading hole layer, the detection hole and channel layer are made of polydimethylsiloxane, and the base layer is made of glass.

[0031] In other embodiments of the present invention, the sample loading hole layer, the detection hole and channel layer, and the base layer are combined together by plasma bonding.

[0032] That is, the sample loading hole layer is fixed together with the detection hole and the channel layer by plasma bonding, and the sample loading hole layer serves as the top cover of the detection hole, and the base layer is fixed together with the detection hole and the channel layer by plasma bonding, and the base layer serves as the bottom cover of the detection hole.

[0033] In the present invention, the microfluidic chip is provided with a plurality of detection holes for different types of target microorganisms, and the photothermal conversion nanomaterials containing antibodies in the reagents pre-buried in the detection holes can specifically bind to specific microorganisms. Under the irradiation of a near-infrared light source, the nanomaterials on the surface of the microorganisms convert light into heat and cause the microorganisms to lyse, thereby releasing ATP. The released ATP can produce bioluminescence with luciferase, luciferin and magnesium ions in the reagents in the presence of oxygen, and the signal is captured by an optical detection element. Therefore, the microfluidic chip described in the present invention can realize the simultaneous detection of different microorganisms.

[0034] In order to prevent the reagents embedded in the microfluidic chip from denaturing or inactivating, the constructed microfluidic chip needs to be sealed and packaged, and then unpacked when it is used.

[0035] The second aspect of the present invention provides a device for selectively detecting microorganisms, which includes a light-proof box, which is a hollow cuboid consisting of a box body and a box cover, and the interior of which is divided into two areas; a near-infrared light source is installed in one of the areas, a bracket for a detection chip is arranged above the near-infrared light source, the bracket is loaded with the microfluidic chip as described in the first aspect, an optical detection element is arranged above the microfluidic chip, so that each detection hole of the microfluidic chip coaxially corresponds to a near-infrared light source and an optical detection element; a signal conversion element is installed in the other area, and the signal conversion element is connected to the optical detection element;

[0036] A display screen is installed on the cover of the light-proof box, and the display screen is connected to the signal conversion element. The display screen can directly display the detected signal value.

[0037] In some embodiments of the present invention, a chip socket is provided on the box body on the side of the light-proof box, and the microfluidic chip is inserted into or extracted from the chip socket.

[0038] In some embodiments of the present invention, the near-infrared light source is an LED light source or a laser diode. Each near-infrared light source is coaxially corresponding to each detection hole of the microfluidic chip.

[0039] In some other embodiments of the present invention, the optical detection element is a photodiode. Each optical detection element is coaxially corresponding to each detection hole of the microfluidic chip.

[0040] In some embodiments of the present invention, the signal conversion element includes a transimpedance amplifier, a signal acquisition card and a single-chip microcomputer connected in series. The transimpedance amplifier is connected to the optical detection element and is used to convert the optical signal into a voltage signal. The voltage signal is further converted into an instrument readable value by the signal acquisition card and the single-chip microcomputer connected to the transimpedance amplifier and displayed on the display screen on the box cover.

[0041] In the present invention, the display screen is also called a data display, which is used to display the detected signal value for easy direct reading.

[0042] In the present invention, the device further comprises a power supply, and the power supply is used to supply power to the device.

[0043] The third aspect of the present invention provides a method for selectively detecting microorganisms using the device according to the second aspect of the present invention, comprising the following steps:

[0044] S1, adding a sample to be tested containing microorganisms into a sample addition hole of the microfluidic chip;

[0045] S2, turning on the optical detection element and recording the first detection signal value of each detection hole;

[0046] S3, after incubating the microfluidic chip, turning on the near-infrared light source to illuminate each detection well for a period of time, turning on the optical detection element again, and recording the second detection signal value of each detection well;

[0047] S4, calculating the difference between the second detection signal value and the first detection signal value of each detection well, and judging whether the sample to be tested contains the target microorganism and / or calculating the concentration of the target microorganism according to the difference.

[0048] In step S1, the sample to be tested enters the detection holes and each channel on the channel layer through the sample addition hole, and further enters each detection hole.

[0049] In the present invention, the first detection signal value is the ATP present in the matrix of the sample to be tested, which has nothing to do with the microorganisms; the second detection signal value is the sum of the ATP released by the microorganisms in the sample to be tested and the ATP present in the matrix of the sample to be tested; the difference between the second detection signal value and the first detection signal value is the ATP released by the microorganisms in the sample to be tested.

[0050] In the present invention, when the sample to be tested contains target microorganisms, the target microorganisms can be combined with the antibodies coated on the photothermal conversion nanomaterials in the reagent through incubation, so that the antibody-coated photothermal conversion nanomaterials are attached to the surface of the microorganisms. Then, through irradiation with a near-infrared light source, the photothermal conversion nanomaterials attached to the surface of the microorganisms convert light energy into heat energy, thereby increasing the surface temperature of the microorganisms, and then causing the microorganisms to thermally decompose and release ATP.

[0051] In some embodiments of the present invention, when the difference between the second detection signal value and the first detection signal value of a detection well is greater than 0, it indicates that the sample to be tested contains the target microorganism detected by the detection well.

[0052] In other embodiments of the present invention, when the difference between the second detection signal value and the first detection signal value of a detection well is greater than 0, the difference of the detection well is brought into the standard curve of the concentration of the target microorganism detected corresponding to the detection well and the detection signal difference, and the concentration of the target microorganism is calculated.

[0053] In the present invention, the standard curve of the concentration of the target microorganism and the difference in the detection signal can be obtained by detecting a series of target microorganism samples with known concentrations.

[0054] In the present invention, the detection signal difference refers to the difference between the second detection signal value and the first detection signal value of a target microorganism sample with a known concentration.

[0055] In some embodiments of the present invention, in step S3, the incubation time is 15 to 20 minutes (for example, 20 minutes), and the incubation temperature is 15 to 37°C.

[0056] In some other embodiments of the present invention, in step S3, the irradiation time is 5 to 10 minutes, for example, 6 minutes.

[0057] In some specific embodiments of the present invention, the specific operation of the method is: add the sample to be tested containing microorganisms to the sample well of the microfluidic chip, and the sample to be tested enters the channel of the middle layer from the upper sample well, and then enters each detection well. Immediately turn on the optical detection element and record the first detection signal value. This signal value is the ATP present in the matrix of the sample to be tested, which has nothing to do with the microorganism, that is, the background signal. Incubate at 15 to 37°C for 15 to 20 minutes, and the antibody-coated photothermal conversion nanomaterial (such as gold nanorods) binds to the surface of the target microorganism. Turn on the near-infrared light source to irradiate the detection well for 5 to 10 minutes and then turn it off. The photothermal conversion nanomaterial (such as gold nanorods) attached to the surface of the microorganism converts light energy into heat energy, so that the surface temperature of the microorganism increases until the microorganism is thermally cracked and releases ATP. Turn on the optical detection element again and record the second detection signal value. This signal value is the sum of the ATP released by the microorganism in the sample to be tested and the ATP in the matrix of the sample to be tested, and the difference between the two is the ATP released by the microorganism in the sample to be tested. The difference can then be used to determine whether the sample to be tested contains the target microorganism and / or calculate the concentration of the target microorganism.

[0058] The beneficial effects of the present invention are as follows: the microfluidic chip provided by the present invention is provided with a plurality of detection holes for different types of target microorganisms, and can detect different microorganisms simultaneously. The device and method containing the microfluidic chip can realize rapid selective detection of multiple specific microorganisms, which can greatly simplify the operation steps, and the device is portable. BRIEF DESCRIPTION OF THE DRAWINGS

[0059] The present invention will be further described below in conjunction with the accompanying drawings.

[0060] Figure 1 The present invention is a schematic diagram of the structure of the microfluidic detection chip used in Example 1; the meanings of the figure numbers in the figure are as follows: 101-microfluidic detection chip; 1-sample loading hole layer; 2-detection hole and channel layer; 3-base layer; 4-sample loading hole; 5-vent valve hole; 6-channel; 7-first detection hole; 8-second detection hole; 9-third detection hole; 10-fourth detection hole; 11-fifth detection hole; 12-sixth detection hole; 13-seventh detection hole; 14-eighth detection hole; 15-ninth detection hole; 16-tenth detection hole; 17-eleventh detection hole.

[0061] Figure 2 This is an appearance diagram of the microfluidic detection chip used in Example 1.

[0062] Figure 3 This is an external structural diagram of the device for selectively detecting microorganisms used in Example 1; the meanings of the figure numbers in the figure are as follows: 24 - display screen; 26 - chip socket.

[0063] Figure 4 This is a diagram of the internal structure of the device for selectively detecting microorganisms used in Example 1; the meanings of the figure numbers in the figure are as follows: 18-LED light source; 19-bracket; 20-photodiode; 21-transimpedance amplifier; 22-signal acquisition card; 23-single-chip microcomputer; 24-display screen; 25-power supply; 101-microfluidic detection chip.

[0064] Figure 5 This is a standard curve diagram of the concentration of microorganisms corresponding to Escherichia coli, Staphylococcus aureus, Salmonella and Bacillus cereus in Example 1 and the difference in detection signals. DETAILED DESCRIPTION

[0065] In order to make the present invention easier to understand, the present invention will be further described in detail below in conjunction with examples, which are merely illustrative and are not intended to limit the scope of application of the present invention. The raw materials or components used in the present invention can be obtained by commercial routes or conventional methods unless otherwise specified.

[0066] The structural schematic diagram and appearance diagram of the microfluidic chip 101 for selectively detecting microorganisms used in the following embodiment 1 are respectively as follows: Figure 1 and 2 As shown, it includes a sample loading hole layer 1, a detection hole and channel layer 2, and a base layer 3 which are sequentially arranged from top to bottom; the sample loading hole layer 1, the detection hole and channel layer 2, and the base layer 3 are combined together by plasma bonding;

[0067] The detection hole and channel layer 2 is provided with 11 channels 6 arranged in a radial shape, and the ends of the 11 channels are provided with detection holes, namely the first detection hole 7, the second detection hole 8, the third detection hole 9, the fourth detection hole 10, the fifth detection hole 11, the sixth detection hole 12, the seventh detection hole 13, the eighth detection hole 14, the ninth detection hole 15, the tenth detection hole 16 and the eleventh detection hole 17; the detection holes are pre-buried with freeze-dried powder of a reagent for selective detection of microorganisms;

[0068] The sample adding hole layer 1 is provided with a sample adding hole 4, and the sample adding hole 4 is used to inject the sample to be tested into the detection hole and each channel 6 on the channel layer 2 through the sample adding hole 4, and further enter each detection hole; the sample adding hole layer 1 is also provided with a vent valve hole 5 at a position corresponding to the detection hole and the detection hole on the channel layer 2;

[0069] The detection hole has a diameter of 5 mm and a depth of 2 mm, the channel has a width of 1 mm and a depth of 100 μm; the sample addition hole has a diameter of 2 mm, and the ventilation valve aperture has a diameter of 1 mm.

[0070] The external structure diagram and internal structure diagram of the selective microorganism detection device used in the following Example 1 are respectively as follows: Figure 3 and 4 As shown; it includes a light-proof box, which is a hollow cuboid consisting of a box body and a box cover, and its interior is divided into two areas; an LED light source 18 is installed in one area, and a bracket 19 for a detection chip is arranged above the LED light source 18, and a microfluidic chip 101 is loaded on the bracket 19, and a photodiode 20 is arranged above the microfluidic chip 101, so that each detection hole of the microfluidic chip 101 coaxially corresponds to an LED light source 18 and a photodiode 20 respectively; a signal conversion element is installed in the other area, and the signal conversion element is connected to the photodiode 20;

[0071] A display screen 24 is installed on the cover of the light-proof box, and the display screen 24 is connected to the signal conversion element;

[0072] A chip socket 25 is provided on the box body on the side of the light-proof box, and the microfluidic chip 101 is inserted or extracted from the chip socket 25;

[0073] The signal conversion element includes a transimpedance amplifier 21, a signal acquisition card 22 and a single-chip microcomputer 23 connected in series in sequence; the transimpedance amplifier is connected to the photodiode 20, and is used to convert the optical signal into a voltage signal; the voltage signal is further converted into an instrument-readable value through the signal acquisition card 22 and the single-chip microcomputer 23 connected to the transimpedance amplifier 21 and displayed on the display screen 24 on the box cover;

[0074] The device further comprises a power supply 25, wherein the power supply 25 is used to supply power to the device.

[0075] Example 1: Rapid and selective detection of Escherichia coli, Staphylococcus aureus, Salmonella, and Bacillus cereus in milk samples.

[0076] The first detection hole of the microfluidic detection chip is pre-buried with freeze-dried powder of a mixture of gold nanorods coated with Escherichia coli antibodies and ATP bioluminescent substrates, the second detection hole is pre-buried with freeze-dried powder of a mixture of gold nanorods coated with Staphylococcus aureus antibodies and ATP bioluminescent substrates, the third detection hole is pre-buried with freeze-dried powder of a mixture of gold nanorods coated with Salmonella antibodies and ATP bioluminescent substrates, the fourth detection hole is pre-buried with freeze-dried powder of a mixture of gold nanorods coated with Bacillus cereus antibodies and ATP bioluminescent substrates, the fifth detection hole is pre-buried with freeze-dried powder of a mixture of gold nanorods without antibody coating and ATP bioluminescent substrates, and the sixth detection hole is pre-buried with freeze-dried powder of a mixture of bacterial lysis reagent and ATP bioluminescent substrates. The fifth detection hole is a negative control hole, because its gold nanorods are not coated with antibodies, so no gold nanorods are attached to the bacterial surface, so no bacteria are lysed and ATP is released. The sixth detection well is a positive control well, and the bacterial lysis reagent can lyse all bacteria to release ATP. The mass concentration of the gold nanorods coated with the corresponding antibodies in the pre-embedded mixed solution in the first detection well to the fourth detection well is 0.025 mg / mL, and the aspect ratio of the gold nanorods is 4.1, and the mass concentration of the ATP bioluminescent substrate (mixed substrate powder of luciferase, luciferin and magnesium ions) is 5 mg / mL.

[0077] The detection steps are specifically as follows: after the milk sample is injected into each detection hole from the sample addition hole during the detection, a photodiode is immediately used to detect the signal and record the first detection signal values ​​of the six detection holes, wherein the first detection signal values ​​in the first detection hole-the fourth detection hole are 0.25, 0.26, 0.19, and 0.3, respectively, and the signal value at this time is the ATP present in the milk matrix. The microfluidic chip is incubated at room temperature for 20 minutes to allow the antibody-coated gold nanorods to bind to the surface of the target microorganism. Then, the six LED light sources located at the bottom of the microfluidic chip are turned on to illuminate the corresponding detection holes for 6 minutes and then turned off, so that the gold nanorods attached to the surface of the bacteria in the six different detection holes undergo photothermal conversion, thereby lysing the bacteria to release ATP. The photodiode is used again to detect the signal and record the second detection signal values ​​of the six detection holes, wherein the second detection signal values ​​in the first detection hole-the fourth detection hole are 2.09, 1.782, 1.8, and 2.26, respectively, and the signal value at this time is the sum of the ATP released by the corresponding microorganism in the milk sample and the ATP in the matrix.

[0078] After calculation, the difference between the second detection signal value and the first detection signal value in the first detection well to the fourth detection well (ATP released by the corresponding specific microorganism in the milk sample) is 1.84, 1.522, 1.61, and 1.96, respectively, and the above difference is brought into the corresponding standard curve (such as Figure 5As shown), the concentrations of Escherichia coli, Staphylococcus aureus, Salmonella and Bacillus cereus in the milk samples were obtained.

[0079] The concentrations of Escherichia coli, Staphylococcus aureus, Salmonella and Bacillus cereus in milk samples were calculated to be 10 5 CFU / mL, 10 5 CFU / mL, 10 5 CFU / mL, 10 5 CFU / mL.

[0080] It should be noted that the embodiments described above are only used to explain the present invention and do not constitute any limitation to the present invention. The present invention has been described with reference to typical embodiments, but it should be understood that the words used therein are descriptive and explanatory words, rather than restrictive words. The present invention may be modified as specified within the scope of the claims of the present invention, and the present invention may be revised without departing from the scope and spirit of the present invention. Although the present invention described therein relates to specific methods, materials and embodiments, it does not mean that the present invention is limited to the specific examples disclosed therein, on the contrary, the present invention can be extended to all other methods and applications with the same functions.

Claims

1. A microfluidic chip for selectively detecting microorganisms, comprising a sample loading hole layer, a detection hole and channel layer, and a base layer arranged in sequence from top to bottom; The detection hole and channel layer is provided with a plurality of channels arranged in a radial shape, and the ends of the plurality of channels are all provided with detection holes; freeze-dried powder of a reagent for selectively detecting microorganisms is pre-embedded in the detection holes; The sample adding hole layer is provided with a sample adding hole, and the sample adding hole is used to inject the sample to be tested into the detection hole and each channel on the channel layer through the sample adding hole, and further enter each detection hole; The reagent is a mixed solution including an antibody-coated photothermal conversion nanomaterial and an ATP bioluminescent substrate, wherein the antibody can specifically bind to the target microorganism to be detected; the photothermal conversion nanomaterial is a gold nanorod; and the ATP bioluminescent substrate includes a thermostable luciferase, luciferin and magnesium ions; The mass concentration of the antibody-coated photothermal conversion nanomaterial in the mixed solution is 0.025-0.1 mg / mL, and the mass concentration of the ATP bioluminescent substrate is 3-5 mg / mL; Under the irradiation of a near-infrared light source, the nanomaterial on the surface of the target microorganism converts light into heat and causes the target microorganism to lyse, thereby releasing ATP; the released ATP can react with the luciferase, luciferin and magnesium ions in the reagent to produce bioluminescence in the presence of oxygen, and the signal is captured by an optical detection element.

2. The microfluidic chip according to claim 1, characterized in that: The number of the detection holes is 3 or more.

3. The microfluidic chip according to claim 2, characterized in that: The number of the detection holes is 4 to 11.

4. The microfluidic chip according to claim 1, characterized in that: The aspect ratio of the gold nanorods is 4-4.

3.

5. The microfluidic chip according to any one of claims 1 to 4, characterized in that: The diameter of the detection hole is 3-5 mm and the depth is 1-2 mm; The channel has a width of 0.5-1 mm and a depth of 50-100 μm.

6. The microfluidic chip according to any one of claims 1 to 4, characterized in that: The diameter of the sample adding holes on the sample adding hole layer is 2-3 mm.

7. The microfluidic chip according to claim 6, characterized in that: The positions of the sample adding hole layer corresponding to the detection holes and the detection holes on the channel layer are provided with ventilation valve holes.

8. The microfluidic chip according to claim 7, characterized in that: The diameter of the ventilation valve hole is 0.5-1 mm.

9. The microfluidic chip according to any one of claims 1 to 4, characterized in that: The sample addition hole layer, detection hole and channel layer, and base layer are all made of transparent materials; The sample addition hole layer, the detection hole and channel layer, and the base layer are combined together through plasma bonding.

10. A device for selectively detecting microorganisms, comprising a light-proof box, the light-proof box being a hollow cuboid consisting of a box body and a box cover, the interior of which is divided into two areas; a near-infrared light source is installed in one area, a bracket for a detection chip is arranged above the near-infrared light source, the bracket is loaded with a microfluidic chip as claimed in any one of claims 1 to 9, an optical detection element is arranged above the microfluidic chip, so that each detection hole of the microfluidic chip coaxially corresponds to a near-infrared light source and an optical detection element; a signal conversion element is installed in the other area, and the signal conversion element is connected to the optical detection element; A display screen is installed on the box cover of the light-proof box, and the display screen is connected to the signal conversion element.

11. The device according to claim 10, characterized in that A chip socket is arranged on the box body at the side of the light-proof box, and the microfluidic chip is inserted into or taken out from the chip socket.

12. A method for selectively detecting microorganisms using the device according to claim 10 or 11, comprising the following steps: S1, adding a sample to be tested containing microorganisms into a sample addition hole of the microfluidic chip; S2, turning on the optical detection element and recording the first detection signal value of each detection hole; S3, after incubating the microfluidic chip, turning on the near-infrared light source to illuminate each detection well for a period of time, turning on the optical detection element again, and recording the second detection signal value of each detection well; S4, calculating the difference between the second detection signal value and the first detection signal value of each detection well, and judging whether the sample to be tested contains the target microorganism or calculating the concentration of the target microorganism according to the difference.

13. The method according to claim 12, characterized in that When the difference between the second detection signal value and the first detection signal value of the detection well is greater than 0, it indicates that the sample to be tested contains the target microorganism detected by the detection well; or When the difference between the second detection signal value and the first detection signal value of a detection well is greater than 0, the difference of the detection well is brought into the standard curve of the concentration of the target microorganism detected corresponding to the detection well and the detection signal difference, and then the concentration of the target microorganism is calculated.

14. The method according to claim 12 or 13, characterized in that In step S3, the incubation time is 15 to 20 minutes, the incubation temperature is 15 to 37° C., and the irradiation time is 5 to 10 minutes.

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