An acoustic fluidic polystyrene microsphere enrichment chip and its use method and application

Abstract

The present invention discloses an acoustofluidic polystyrene microsphere enrichment chip and its use method and application, and relates to the technical field of acoustofluidic enrichment chips. It includes: a piezoelectric transducer, which is connected to a signal generator and used to convert the electrical energy generated by the signal generator into acoustic mechanical energy; a microfluidic channel, which is connected to a piezoelectric transducer and used to receive the acoustic mechanical energy generated by the piezoelectric transducer, and an acoustic node is generated in the center of the microfluidic channel under the action of acoustic waves; the microfluidic channel is provided with a microfluidic channel inlet and a microfluidic channel outlet, and a solution containing modified polystyrene microspheres is injected through the microfluidic channel inlet, and the modified polystyrene microspheres are effectively enriched in the center of the microfluidic channel under the action of acoustic waves. The polystyrene microsphere enrichment chip provided by the present invention has the advantages of small device size and low cost, and the use method is simple, and can be used for the separation and detection of food or drug components, with good repeatability and high separation and detection efficiency.

Description

Technical Field

[0001] The present application relates to the technical field of acoustofluidic enrichment chips, and in particular to an acoustofluidic polystyrene microsphere enrichment chip and a use method and application thereof. Background Art

[0002] Polystyrene (PS) microspheres have a wide range of applications. Studies have shown that they can be used for particle tracking and particle image velocimetry in rheological experiments; they can be used as carriers for drug delivery and targeted release in the biomedical field, as well as for the separation and purification of biomolecules; they can be used to prepare new materials in the field of materials science; and they can be used in emerging applications such as in vitro diagnostics through functional modification of their surfaces. However, unlike magnetic particles in a magnetic field or charged particles in an electric field, the manipulation of PS microspheres often relies on the drive of an external field, while acoustic fields that are not affected by electromagnetic radiation, non-contact, and selective often have the advantage of manipulating PS microspheres. In addition, the development of a fast and simple method for aggregating polystyrene microspheres will help promote the development of related research.

[0003] In the prior art, polystyrene microsphere enrichment chips based on acoustic fluidics have the disadvantages of complicated preparation, high cost and large sample amount required. Summary of the invention

[0004] The present invention aims to solve at least one of the technical problems existing in the prior art or related art.

[0005] The first aspect of the present invention provides an acoustofluidic polystyrene microsphere enrichment chip, comprising:

[0006] A piezoelectric transducer, the piezoelectric transducer is connected to the signal generator and is used to convert the electrical energy generated by the signal generator into acoustic mechanical energy;

[0007] A microfluidic channel is connected to the piezoelectric transducer and is used to receive the acoustic mechanical energy generated by the piezoelectric transducer. An acoustic node is generated at the center of the microfluidic channel under the action of the acoustic wave. The microfluidic channel is provided with a microfluidic channel inlet and a microfluidic channel outlet. A solution containing modified polystyrene microspheres is injected through the microfluidic channel inlet. Under the action of the acoustic wave, the modified polystyrene microspheres are effectively enriched at the center of the microfluidic channel.

[0008] Furthermore, the acoustofluidic polystyrene microsphere enrichment chip further comprises:

[0009] A substrate, on which the piezoelectric transducer is arranged.

[0010] Furthermore, a gasket is provided between the piezoelectric transducer and the substrate so that a gap is left between the piezoelectric transducer and the substrate.

[0011] Furthermore, the microfluidic channel is a square glass capillary, the inner diameter of the square glass capillary is 0.18 μm~0.22 μm, and the outer diameter of the square glass capillary is 0.38 μm~0.42 μm.

[0012] The second aspect of the present invention provides a method for using the above-mentioned acoustofluidic polystyrene microsphere enrichment chip, comprising:

[0013] injecting the solution containing the modified polystyrene microspheres from the inlet of the microfluidic channel;

[0014] Turning on the signal generator to transmit the acoustic wave in the microfluidic channel and generate an acoustic node at the center of the microfluidic channel, and the modified polystyrene microspheres are effectively enriched in the center of the microfluidic channel within 2 minutes;

[0015] Detection analysis was performed based on image processing after enrichment.

[0016] Furthermore, the signal generator is controlled to output a sine wave frequency of 3.0 MHz to 3.8 MHz, and the signal generator is controlled to output a sine wave amplitude range of 1 Vpp to 20 Vpp.

[0017] Furthermore, the volume of the solution containing the modified polystyrene microspheres in the microfluidic channel is controlled to be no less than 25 μL, and the concentration of the modified polystyrene microspheres in the solution containing the modified polystyrene microspheres is 50 μg / mL~1000 μg / mL.

[0018] Furthermore, the particle size of the polystyrene microspheres in the solution containing the modified polystyrene microspheres is 450 nm to 10.5 μm.

[0019] Furthermore, in the data collection process for measuring the fluorescence intensity, the signal generator is turned on after the solution containing the modified polystyrene microspheres in the microfluidic channel becomes stationary.

[0020] The third aspect of the present invention provides the use of the above-mentioned acoustofluidic polystyrene microsphere enrichment chip for rapid separation and detection of food or drug components.

[0021] Compared with the prior art, the present invention has at least the following beneficial effects:

[0022] The acoustofluidic polystyrene microsphere enrichment chip provided by the present invention effectively enriches the modified polystyrene microspheres in the microfluidic channel at the center of the microfluidic channel through the transmission of acoustic waves in the microfluidic channel and the generation of acoustic nodes.

[0023] The device has the advantages of small size, low cost and simple use. It can be used for separation and detection of food or drug components with good repeatability and high analysis and detection efficiency. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] Various other advantages and benefits will become apparent to those of ordinary skill in the art by reading the following detailed description of the preferred embodiment.The accompanying drawings are only for the purpose of illustrating the preferred embodiments and are not to be considered as limiting the present application.

[0025] Figure 1 This is a schematic diagram of the structure of the acoustic fluidic polystyrene microsphere enrichment chip provided in Example 1 of the present application.

[0026] Figure 2 A finite element simulation diagram of the sound pressure in the cross section of a small chip made of the microfluidic channel and piezoelectric transducer provided in Example 1 of the present application.

[0027] Figure 3 This is a finite element simulation diagram of the motion trajectory of the microspheres in the cross section of the microfluidic channel provided in Example 1 of the present application.

[0028] Figure 4 This is a graph showing the test results of the resonance frequency required for the polystyrene microsphere enrichment chip provided in Example 1 of the present application to cause the polystyrene microsphere enrichment phenomenon.

[0029] Figure 5 This is a comparison chart of the fluorescence intensity of fluorescent microspheres after aggregation in a solution containing polystyrene microspheres in a flowing or static state provided in Example 1 of the present application.

[0030] Figure 6 Ten sets of parallel images and fluorescence intensity comparisons collected when testing fluorescent polystyrene microspheres provided in Example 1 of the present application.

[0031] Figure 7 This is a diagram showing the enrichment effect of polystyrene microspheres of different particle sizes provided in Example 1 of the present application.

[0032] Figure 8 This is a graph of the fluorescence intensity in the microfluidic channel provided in Example 1 of the present application.

[0033] The reference numerals are as follows: 1. piezoelectric transducer; 2. microfluidic channel; 201. microfluidic channel inlet; 202. microfluidic channel outlet; 3. gasket; 4. substrate. DETAILED DESCRIPTION

[0034] In order to better understand the above-mentioned technical scheme, the technical scheme of the embodiments of the present application is described in detail below through the accompanying drawings and specific embodiments. It should be understood that the embodiments of the present application and the specific features in the embodiments are detailed descriptions of the technical scheme of the embodiments of the present application, rather than limitations on the technical scheme of the present application. In the absence of conflict, the embodiments of the present application and the technical features in the embodiments may be combined with each other.

[0035] According to a first aspect of an embodiment of the present invention, there is provided an acoustic fluidic polystyrene microsphere enrichment chip, comprising:

[0036] A piezoelectric transducer 1, the piezoelectric transducer 1 is connected to a signal generator and is used to convert the electrical energy generated by the signal generator into acoustic mechanical energy;

[0037] A microfluidic channel 2, wherein the microfluidic channel 2 is connected to the piezoelectric transducer 1 and is used to receive the acoustic mechanical energy generated by the piezoelectric transducer 1, and an acoustic node is generated at the center of the microfluidic channel 2 under the action of the acoustic wave; the microfluidic channel 2 is provided with a microfluidic channel inlet 201 and a microfluidic channel outlet 202, a solution containing modified polystyrene microspheres is injected through the microfluidic channel inlet 201, and the modified polystyrene microspheres are effectively enriched at the center of the microfluidic channel 2 under the action of the acoustic wave.

[0038] The acoustofluidic polystyrene microsphere enrichment chip provided in the embodiment of the present invention realizes the transmission of acoustic waves and the generation of acoustic nodes in the microfluidic channel 2 by connecting the piezoelectric transducer 1 with the microfluidic channel 2, thereby being able to act on the modified polystyrene microspheres in the microfluidic channel to effectively enrich them in the center of the microfluidic channel. Compared with the existing reported acoustic wave microfluidic enrichment chips, the present invention achieves the advantages of low cost, rapid preparation, contactless and rapid enrichment.

[0039] Specifically, the signal generator can be a portable signal generator, the piezoelectric transducer 1 can be a piezoelectric transducer PZT made of lead zirconate titanate material, and the microfluidic channel 2 can be connected to the piezoelectric transducer 1 by adhesion (for example, the microfluidic channel 2 is adhered to the piezoelectric transducer 1 by epoxy resin glue). Driven by the signal generator, the piezoelectric transducer 1 generates a bulk acoustic wave, and the solution containing modified polystyrene microspheres flowing through the microfluidic channel 2 is rebounded by the inner wall of the opposite microchannel, thereby generating two sound waves with the same frequency but opposite transmission directions in the microchannel, i.e., standing waves. Therefore, at a preset frequency, an acoustic pressure node can be generated in the center of the microfluidic channel 2, so that the modified polystyrene microspheres can converge quickly to the node, thereby achieving effective enrichment.

[0040] In some embodiments, the acoustofluidic polystyrene microsphere enrichment chip further comprises: a substrate 4, and the piezoelectric transducer 1 is disposed on the substrate 4. Preferably, a gasket 3 is disposed between the piezoelectric transducer 1 and the substrate 4, so that a gap is left between the piezoelectric transducer 1 and the substrate 4.

[0041] Specifically, a gap can be provided between the piezoelectric transducer 1 and the substrate 4 to reduce the transmission of acoustic wave energy from the piezoelectric transducer 1 to the microfluidic channel 2, so that the acoustofluidic polystyrene microsphere enrichment chip has a higher quality factor at the resonance frequency. For example, a gasket 3 is provided between the piezoelectric transducer 1 and the substrate 4 so that a gap is provided between the piezoelectric transducer 1 and the substrate 4. The gasket 3 can be a plastic gasket.

[0042] In some embodiments, the microfluidic channel 2 is a square glass capillary, the inner diameter of the square glass capillary is 0.18 μm to 0.22 μm, and the outer diameter of the square glass capillary is 0.38 μm to 0.42 μm. Preferably, the inner diameter of the square glass capillary is 0.2 μm, and the outer diameter of the square glass capillary is 0.4 μm.

[0043] Specifically, the microfluidic channel 2 is a glass capillary with a square cross section, the inner diameter of the capillary is 0.18 μm ~ 0.22 μm, the outer diameter of the square glass capillary is 0.38 μm ~ 0.42 μm, and the length is 40±2 mm. The advantage of selecting this glass capillary is that it is low in cost, and the μm-level inner diameter can greatly reduce the volume required for detection, solving the defect of large sample volume required for polystyrene microsphere enrichment chips based on acoustic fluidics in the prior art.

[0044] A second aspect of an embodiment of the present invention provides a method for using the above-mentioned acoustofluidic polystyrene microsphere enrichment chip, comprising:

[0045] Injecting the solution containing the modified polystyrene microspheres from the microfluidic channel inlet 201;

[0046] Turning on the signal generator to transmit the acoustic wave in the microfluidic channel 2 and generate an acoustic node at the center of the microfluidic channel 2, and the modified polystyrene microspheres are effectively enriched at the center of the microfluidic channel 2 within 2 minutes;

[0047] Detection analysis was performed based on image processing after enrichment.

[0048] The method for using the acoustofluidic polystyrene microsphere enrichment chip provided in the embodiment of the present invention is simple to use, can achieve rapid aggregation of modified polystyrene microspheres within 2 minutes, and has high detection efficiency and good repeatability.

[0049] In some embodiments, the control signal generator outputs a sine wave with a frequency of 3.0 MHz to 3.8 MHz, preferably 3.36 MHz, and the control signal generator outputs a sine wave with an amplitude of 1 Vpp to 20 Vpp, preferably 20 Vpp.

[0050] In some embodiments, the volume of the solution containing the modified polystyrene microspheres in the microfluidic channel 2 is controlled to be no less than 25 μL, and the concentration of the modified polystyrene microspheres in the solution containing the modified polystyrene microspheres is 50 μg / mL-1000 μg / mL.

[0051] Specifically, considering the internal volume of the microfluidic channel 2 and the internal volume of the hose connecting the inlet and outlet of the microfluidic channel 2, the volume of the solution containing the modified polystyrene microspheres in the microfluidic channel 2 is controlled to be no less than 25 μL. In the selection of the concentration range, the lowest concentration of 50 μg / mL is the concentration point corresponding to the maximum cross-section of the enrichment zone being only a thin line, and the highest concentration of 1000 μg / mL is selected based on the concentration point corresponding to the maximum cross-section of the enrichment zone being approximately half of the center of the microchannel. And the experiment found that: when the volume of the solution containing the modified polystyrene microspheres is no less than 25 μL, and the concentration of the modified polystyrene microspheres in the solution containing the modified polystyrene microspheres is 50 μg / mL~1000 μg / mL, the rapid convergence of the modified polystyrene microspheres can be achieved, and the detection efficiency is high and the repeatability is good.

[0052] In some embodiments, the particle size of the polystyrene microspheres in the solution containing modified polystyrene microspheres is 450 nm to 10.5 μm.

[0053] Specifically, due to the excellent enrichment effect of the polystyrene microsphere enrichment chip, the acoustic field can manipulate the particle size range from micrometer level to nanometer level. The experiment found that the particle size of polystyrene microspheres between 450 nm and 10.5 μm can achieve excellent enrichment effect.

[0054] In some embodiments, during the data collection process for measuring fluorescence intensity, the signal generator is turned on after the solution containing the modified polystyrene microspheres in the microfluidic channel 2 becomes stationary.

[0055] Specifically, the fluorescence microscope has a specific excitation light source and filter system, and the modified polystyrene microspheres are converted into visible images of fluorescence signals through the signal detection system, and then data analysis is performed. In our experiment, we found that the solution containing modified polystyrene microspheres in the microfluidic channel 2 can converge at the center of the microfluidic channel 2 in both flowing and static states, but the fluorescence intensity measured by the convergence of the modified polystyrene microsphere solution after static is stronger than that in the flowing state. Therefore, in the data collection process of measuring fluorescence intensity, it is preferred to test in a static state.

[0056] A third aspect of the embodiments of the present invention provides an application of the above-mentioned acoustofluidic polystyrene microsphere enrichment chip for rapid separation and detection of food or drug components.

[0057] The acoustic fluidic polystyrene microsphere enrichment chip provided in the embodiment of the present invention can be applied to nano-scale targets. Compared with the prior art, the manipulated size is reduced from the micron level to the nano-level. In addition, in applications such as in vitro diagnosis, disease, pathogen detection, and foodborne pathogen detection based on image processing after enrichment of modified polystyrene microspheres, the pre-treatment process of sample preparation (i.e., the polystyrene microsphere modification process: such as the preparation of signal probes coupled to polystyrene microspheres and antibody capture of coupled polystyrene microspheres, etc.) is combined to improve the sensitivity (the effect of signal amplification is achieved after the signal polystyrene microspheres are aggregated), which is of significant significance to improving the efficiency of diagnosis and detection. At the same time, it can also be applied to the aggregation of polystyrene microspheres coupled to nucleic acid probes to achieve selective detection.

[0058] Example 1 An acoustic fluidic polystyrene microsphere enrichment chip and its use method

[0059] (a) As Figure 1 As shown, an acoustic fluidic PS microsphere enrichment chip includes: a piezoelectric transducer 1, a microfluidic channel 2, a gasket 3, and a substrate 4. Among them, the piezoelectric transducer 1 is a piezoelectric transducer PZT, with a size of 20mm×15mm×2.1mm; the microfluidic channel 2 is a square glass capillary, with an inner diameter of 0.2μm and an outer diameter of 0.4μm, and is provided with a microfluidic channel inlet 201 and a microfluidic channel outlet 202; the gasket 3 is a plastic gasket; and the substrate 4 is a glass substrate.

[0060] A piezoelectric transducer 1 and a microfluidic channel 2 are sequentially arranged above the substrate 4 . The microfluidic channel 2 is connected to the piezoelectric transducer 1 by a bonding method, and a gasket 3 made of plastic is arranged between the piezoelectric transducer 1 and the substrate 4 .

[0061] The piezoelectric transducer 1 is connected to the signal generator and is used to convert the electrical energy generated by the signal generator into acoustic mechanical energy; the microfluidic channel 2 receives the acoustic mechanical energy generated by the piezoelectric transducer 1, and an acoustic node is generated in the center of the microfluidic channel 2 under the action of the acoustic wave. A solution containing modified polystyrene microspheres is injected through the microfluidic channel inlet 201, and the modified polystyrene microspheres are effectively enriched in the center of the microfluidic channel 2 under the action of the acoustic wave, and excess solution flows out from the microfluidic channel outlet 202, and the microfluidic channel outlet 202 is connected to a waste liquid bottle.

[0062] (II) A method for using an acoustofluidic polystyrene microsphere enrichment chip, comprising:

[0063] (1) The acoustic wave generated by the piezoelectric transducer PZT and the acoustic node formed by the reflection back to the center of the microfluidic channel 2 are simulated and analyzed. The simulation result shows that the sound pressure at the center of the cross section of the microfluidic channel 2 (square glass capillary) is 0 ( Figure 2 ), the simulation diagram of the PS microsphere particle trajectory shows that the particles at each position of the cross section converge toward the center ( Figure 3 ).Depend on Figure 2 and Figure 3 The finite element simulation results further verified the feasibility of this chip enrichment.

[0064] (2) Network analyzer measurement of the resonant frequency of the piezoelectric transducer PZT: In order to achieve the best operating frequency of the piezoelectric transducer PZT, the reflection coefficient in the range of 3.0~3.8 MHz is analyzed using S parameters. Figure 4 The spectrum shows that the S11 reflection coefficient is the smallest when the input frequency is 3.36 MHz, which indicates that this frequency is the optimal operating frequency.

[0065] (3) Set the output frequency of the signal generator: Refer to the optimal operating frequency measured by the network analyzer and set the output sine wave frequency to 3.36 MHz.

[0066] (4) Setting the output sine wave amplitude of the signal generator: The setting range is 1 Vpp~20 Vpp. By testing the aggregation of PS microspheres at 1 Vpp, 5 Vpp, 10 Vpp, 15 Vpp and 20 Vpp, it was found that when the voltage range is set to 1 Vpp~20 Vpp, the aggregation effect of fluorescent-modified polystyrene microspheres can be achieved.

[0067] (5) Data acquisition image determination: Figure 5As shown, during the test, the solution containing fluorescently modified polystyrene microspheres in the microfluidic channel 2 can achieve central convergence in both the flowing and static states. However, the fluorescence intensity measured when the solution converges after being static is stronger than that in the flowing state. Therefore, in the process of collecting data for measuring fluorescence intensity, it is preferred to conduct the test in the static state.

[0068] (6) Under static conditions, the fluorescence intensity of the fluorescent PS microspheres and the good repeatability of the experiment were analyzed with the help of imageJ software, such as Figure 6 As shown, the enrichment area in the center of the channel is uniform in shape, and enrichment was achieved at the same position in 10 repeated experiments.

[0069] (7) In order to verify that the present invention has a strong enrichment ability, the particle size range of the PS microspheres that can be enriched is from micrometer to nanometer. PS microspheres with five different particle sizes of 10 μm, 5 μm, 2 μm, 1 μm and 500 nm were selected and enriched using the parameter settings of Example 1, as shown in FIG. Figure 7 As shown, the results show that different PS microsphere particle sizes can achieve excellent enrichment effects.

[0070] (8) In order to verify that the present invention has a good quantitative effect, we used PS microspheres with a particle size of 500 nm and prepared seven concentrations of fluorescent PS microsphere solutions of 50 μg / mL, 75 μg / mL, 100 μg / mL, 250 μg / mL, 500 μg / mL, 750 μg / mL and 1000 μg / mL. After the fluorescent PS microsphere solutions were injected into the microfluidic channel, a CCD camera was used to record the fluorescence image in the microfluidic channel during aggregation, and a curve was drawn according to the corresponding fluorescence intensity, as shown in FIG. Figure 8 As shown, the chip has a good quantitative effect, and the correlation coefficient of the quantitative curve drawn is ( r 2 ) is greater than 0.99.

[0071] It is easy for those skilled in the art to understand that, under the premise of no conflict, the above-mentioned advantageous methods can be freely combined and superimposed. The above are only preferred embodiments of the present application and are not intended to limit the present application. Any modifications, equivalent substitutions and improvements made within the spirit and principles of the present application should be included in the protection scope of the present application. The above are only preferred implementation methods of the present application. It should be pointed out that for ordinary technicians in this technical field, several improvements and variations can be made without departing from the technical principles of the present application, and these improvements and variations should also be regarded as the protection scope of the present application.

Claims

1. A method for using an acoustofluidic polystyrene microsphere enrichment chip, characterized in that: include: A solution containing modified polystyrene microspheres is injected from the inlet (201) of the microfluidic channel; Turning on the signal generator to transmit the acoustic wave in the microfluidic channel (2) and generate an acoustic node at the center of the microfluidic channel (2), so that the modified polystyrene microspheres are effectively enriched at the center of the microfluidic channel (2) within 2 minutes; Detection and analysis based on image processing after enrichment; The volume of the solution containing the modified polystyrene microspheres in the microfluidic channel (2) is controlled to be no less than 25 μL, and the concentration of the modified polystyrene microspheres in the solution containing the modified polystyrene microspheres is 50 μg / mL to 1000 μg / mL; The particle size of the polystyrene microspheres in the solution containing the modified polystyrene microspheres is 450 nm to 10.5 μm; Controlling the signal generator to output a sine wave frequency of 3.0 MHz to 3.8 MHz, and controlling the signal generator to output a sine wave amplitude range of 1 Vpp to 20 Vpp; Wherein, the acoustofluidic polystyrene microsphere enrichment chip comprises: A piezoelectric transducer (1), the piezoelectric transducer (1) being connected to a signal generator and used for converting the electrical energy generated by the signal generator into acoustic mechanical energy; A microfluidic channel (2), wherein the microfluidic channel (2) is connected to the piezoelectric transducer (1) and is used to receive acoustic mechanical energy generated by the piezoelectric transducer (1), and to generate an acoustic node at the center of the microfluidic channel (2) under the action of the acoustic wave; the microfluidic channel (2) is provided with a microfluidic channel inlet (201) and a microfluidic channel outlet (202); a solution containing modified polystyrene microspheres is injected through the microfluidic channel inlet (201), and the modified polystyrene microspheres are effectively enriched at the center of the microfluidic channel (2) under the action of the acoustic wave; A substrate (4), the piezoelectric transducer (1) being arranged on the substrate (4); The microfluidic channel (2) is a square glass capillary, the inner diameter of the square glass capillary is 0.18 μm to 0.22 μm, and the outer diameter of the square glass capillary is 0.38 μm to 0.42 μm.

2. The method of use according to claim 1, characterized in that: During the data collection process for measuring the fluorescence intensity, after the solution containing the modified polystyrene microspheres in the microfluidic channel (2) becomes stationary, the signal generator is turned on.

3. The method of use according to claim 1 or 2, characterized in that: A gasket (3) is provided between the piezoelectric transducer (1) and the substrate (4), so that a gap is left between the piezoelectric transducer (1) and the substrate (4).

4. An application of a method for using an acoustofluidic polystyrene microsphere enrichment chip, characterized in that: The method for using the acoustofluidic polystyrene microsphere enrichment chip according to any one of claims 1 to 3 is used for rapid separation and detection of food or drug components.

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