TW202026429A - Multi-color fluorescent excitation and detection device - Google Patents

Abstract

The present invention discloses a multi-color fluorescent excitation and detection device. The multi-color fluorescent excitation and detection device comprises at least one illumination module, a cartridge, a liquid lens module and at least one detection module. The illumination module provides illumination light at a specified range of wavelengths. The cartridge comprises a detection chip comprising plural detection wells. Each of the detection wells is adapted for accommodating a corresponding fluorescent sample therein. Each of the detection wells includes a first wall and a second wall. The illumination light transmits through the first wall to excite the fluorescent sample so as to generate a fluorescent signal. The liquid lens module comprising at least one liquid lens attached on the second wall. The liquid lens module is configured to enhance the fluorescent signal while the fluorescent signal passes through the second wall and the liquid lens. The detection module receives the fluorescent signal and converts the enhanced fluorescent signal to an electrical signal.

Description

Multicolor fluorescent excitation and detection device

This case relates to a fluorescent detection device, especially a multi-color fluorescent excitation and detection device with liquid lens modules.

In order to obtain a large number of specific fragments of DNA in response to different needs, the method has been booming in recent years. Scientists have been striving to find efficient ways to meet their goals. Polymerase chain reaction (PCR) is one of them. One of the most economical and rapid technology that can obtain one billion copies of specific DNA fragments in a short time. PCR technology can be applied in a variety of fields, such as selective DNA isolation for genetic identification, forensic analysis of ancient DNA in archaeology, medical application of genetic testing and tissue types, rapid and accurate diagnosis of infectious diseases in hospitals and research institutions , Food safety environmental hazard inspections and genetic fingerprints used to investigate criminals, etc. PCR technology only needs to use a small amount of DNA sample extracted from blood or tissue, and add fluorescent dye to the nucleic acid solution, and the amplified DNA fragments can be detected through fluorescent molecules.

Dye and fluorescence detection technology is a widely used technology to simultaneously detect and analyze whether target nucleic acid molecules are present in a batch of biological samples. When excited by light of a specific wavelength, when a fluorescent signal is emitted from a target nucleic acid molecule with a DNA-binding dye or a fluorescent-binding probe, this signal represents the presence of the target nucleic acid molecule. This technique is applied to a new type of PCR technique called isothermal amplification method. Among them, the optical device is an indispensable tool in the qPCR detection technology to detect the fluorescence emitted by a specific nucleic acid fragment. The optical device must provide a light source to excite the fluorescent probe at a specific wavelength and simultaneously detect the secondary probe The fluorescent signal emitted.

Another alternative method of isothermal amplification is that it does not require thermal cycling control, but instead relies on proteins that utilize the DNA/RNA synthesis mechanism in the body and are dominated by enzyme activity. Therefore, the miniaturized isothermal system has the advantages of simple design and low energy consumption. At present, a variety of analysis complexity (multiple enzymes or primers), detection sensitivity and specificity acceptable isothermal amplification methods have been developed, including nucleic acid sequence-based amplification (NASBA), Strand displacement amplification (SDA), helicase-dependent amplification (HAD), loop-mediated isothermal amplification (LAMP), recombinant enzyme polymerase amplification Increase technology (recombinase polymerase amplification, RPA), and nicking enzyme amplification reaction (nicking enzyme amplification reaction, NEAR).

Fluorescence detection systems have been maturely developed in many fields, such as the application of fluorescence spectroscopy and fluorescence microscopy. Using a monochromatic light source with a set of filters and optical elements, it can be easily applied to monochromatic fluorescent probes. However, most of the existing fluorescent detection systems have the disadvantages of large size, complex structure and high cost. What's more, the fluorescent signal produced by most fluorescent detection systems has a low Signal-to-Noise ratio (SNR). The current fluorescent detection device design of the portable isothermal amplification method still lags behind the development of its biochemical technology. Since isothermal amplification has a high tolerance for sample purification, most isothermal platforms on the market focus on creating a stable temperature environment and a detection method with medium to high throughput. More importantly, most devices using isothermal amplification prefer to be in a motion detection environment, so the system needs to have a high degree of integration. Therefore, although most current optical element designs have been widely used, they are not suitable Apparatus for isothermal amplification.

One of the objectives of this project is to provide a multi-color fluorescent excitation and detection device that concentrates the fluorescent signals to the detection module through the liquid lens module to obtain a high signal-to-noise ratio, reduce the overall volume and weight, and compare The low cost provides better performance of the portable isothermal PCR system and allows the detection of tank deviation.

Another purpose of this case is to provide a multi-color fluorescent excitation and detection device, which has a compact optical structure to avoid difficulties in alignment and assembly, and has a high-performance and solid structure to avoid loss caused by light transmission and maintain High signal-to-noise ratio.

According to the concept of this case, this case provides a multi-color fluorescent excitation and detection device, which includes at least one light emitting module, a cassette, a liquid lens module, and at least one detection module. Each of the light-emitting modules provides illumination light having a specific wavelength range. The cassette contains a detection chip, and the detection chip contains a plurality of detection slots, which are arranged around the periphery of the detection chip. Each detection slot contains a corresponding fluorescent sample, and each detection slot contains The first wall surface and the second wall surface, wherein the illuminating light penetrates the first wall surface to illuminate the fluorescent sample in the detection tank, and excites the fluorescent sample to generate a fluorescent signal to penetrate the second wall surface. The liquid lens module includes at least one liquid lens attached to the second wall surface to enhance the fluorescent signal when the fluorescent signal penetrates the second wall surface and the liquid lens. At least one detection module receives the enhanced fluorescent signal and converts the fluorescent signal into an electrical signal.

Some typical embodiments embodying the features and advantages of this case will be described in detail in the following description. It should be understood that this case can have various changes in different aspects, which do not deviate from the scope of the case, and the descriptions and diagrams therein are essentially for illustrative purposes, rather than limiting the case.

This case provides a multi-color fluorescence excitation and detection device suitable for nucleic acid analysis equipment. To further explain, the nucleic acid analysis equipment applicable to the multi-color fluorescence excitation and detection device in this case integrates a fluid delivery unit, a temperature control unit, a rotation drive unit, and a multi-color fluorescence excitation and detection device on a single device, so Nucleic acid analysis can be performed on a single device in real time to achieve the functions of sample purification, nucleic acid extraction, nucleic acid diffusion, and nucleic acid detection.

Figure 1 is a schematic structural diagram of a nucleic acid analysis device using a multi-color fluorescent excitation and detection device according to an embodiment of the present invention. Figure 2 is a schematic structural diagram of the nucleic acid analysis device shown in Figure 1 when the tank is opened. The cassette is schematically removed from the nucleic acid analysis equipment. As shown in FIGS. 1 and 2, the nucleic acid analysis equipment 100 includes a tank 1, a fluid conveying unit 2, a temperature control unit 3, a rotation driving unit 4 and a multi-color fluorescence excitation and detection device 9. The multi-color fluorescent excitation and detection device 9 includes a cassette 6, at least one light-emitting module 7, a liquid lens module 5, and at least one detection module 8 (of which light-emitting module 7 and liquid lens module 5, please refer to 3 pictures). The tank 1 can be opened to install the cassette 6 therein. The fluid conveying unit 2 is connected to the tank body 1 and is suitable for conveying reagents in the cassette 6 for sample purification and/or nucleic acid extraction. The temperature control unit 3 is arranged in the tank 1 and is adapted to provide a preset temperature for nucleic acid amplification. The rotary drive unit 4 is connected to the tank body 1 and can rotate the cassette 6 in the tank body 1 in a preset program. In an embodiment, the rotary drive unit 4 can clamp the cassette 6. At least one light-emitting module 7 and at least one detection module 8 are disposed on the tank 1, each light-emitting module 7 includes at least one optical element for laser, and each detection module 8 includes at least one for detection Optical element for nucleic acid detection or sample reaction detection.

In one embodiment, the tank body 1 includes a top tank body 11 and a bottom tank body 12. The top tank body 11 and the bottom tank body 12 are connected by a hinge 13, but not limited to this. The bottom tank 12 has a cavity 121 which is specially designed for installing the cassette 6 therein. The top tank 11 can be opened so that the cassette 6 can be placed in the cavity 121 of the bottom tank 12. When the top tank 11 is closed, a closed space is formed in the tank 1. In an embodiment, the shape of the tank body 1 may be but not limited to cylindrical, spherical, cubic, cone or olive shape, and the tank body 1 may be, but not limited to, metal, ceramic, polymer, polymer compound, wood , Glass, or other materials that can provide good thermal insulation.

The bottom tank 12 is connected to the fluid delivery unit 2 through a pipe or a flow channel. When the cassette 6 is installed in the bottom tank 12, the cassette 6 will be locked and tightly contacted with the fluid delivery unit 2 to avoid leakage. For example, the cassette 6 can be locked to the bottom slot 12 through at least one fixing element, wherein the fixing element may include a clip, but it is not limited thereto.

As shown in FIG. 2, the cassette 6 includes a detection chip 62 and a reagent storage body 61, and the detection chip 62 is disposed on the top of the reagent storage body 61. The detection chip 62 is a flat fluid chip. The detection chip 62 includes a plurality of detection grooves 625 (as shown in FIG. 4 ), at least one first channel 63 and at least one second channel 64. At least one first channel 63 is connected to the detection slot 625 via at least one second channel 64. In one embodiment, a plurality of detection grooves 625 are arranged around the periphery of the detection chip 62 and contain samples or reagents for nucleic acid amplification and/or detection. For example, the detection tank 625 can be coated with samples or reagents for nucleic acid amplification and/or detection, such as reagents including different fluorescent dyes. The number of detection slots 625 is not limited, and can be as many as 40 or more slots, so the equipment in this case can perform multiplexing nucleic acid analysis. In some embodiments, the shape of the detection chip 62 is substantially circular, so that the detection chip 62 has a plurality of curved sides for aligning with the at least one detection module 8 to facilitate focusing.

The reagent storage body 61 includes a plurality of reagent tanks (not shown), which store reagents for sample purification and/or nucleic acid extraction. The reagent storage body 61 also includes a plurality of flow channels which are connected to the reagent tank for fluid delivery. In an embodiment, the reagent storage body 61 may be, but is not limited to, a cylindrical body. The detection chip 62 further includes at least one opening 65 provided on the top surface of the detection chip 62, and the opening 65 is aligned with and communicated with at least one reagent tank of the reagent storage body 61 for adding a sample to the cassette 6.

FIG. 3 is a schematic diagram of the structure of the multi-color fluorescence excitation and detection device of the preferred embodiment of the present invention, and FIG. 4 is a schematic diagram of a partially enlarged structure of the multi-color fluorescence excitation and detection device shown in FIG. 3. As shown in FIGS. 1, 2, 3, and 4, the multi-color fluorescent excitation and detection device 9 includes a cassette 6, at least one light emitting module 7, a liquid lens module 5, and at least one detection module 8. The multi-color fluorescent excitation and detection device 9 can be, but is not limited to, include four light-emitting modules 7 and four detection modules 8. Each light-emitting module 7 is disposed in the accommodating space 14 of the bottom tank 12 (Please refer to Figure 2), and provide illumination light with a specific wavelength range.

Referring to FIG. 4, each detecting groove 625 includes a first wall surface 623, a second wall surface 621, a third wall surface 622, a fourth wall surface 624, a fifth wall surface, and a sixth wall surface (not shown). The first wall surface 623 is opposite to the third wall surface 622, the second wall surface 621 is opposite to the fourth wall surface 624, and the fifth wall surface is opposite to the sixth wall surface. The second wall surface 621, the fourth wall surface 624, the fifth wall surface and the sixth wall surface are connected and disposed between the first wall surface 623 and the third wall surface 622. In this embodiment, the first wall surface 623 is a bottom wall surface, the second wall surface 621 is a front wall surface, the third wall surface 622 is an upper wall surface, the fourth wall surface 624 is a rear wall surface, and the fifth wall surface is Is the first side wall, and the sixth wall is the second side wall.

Fig. 5 is a schematic diagram of the structure of the detecting tank and the liquid lens module shown in Fig. 4. As shown in FIGS. 4 and 5, in this embodiment, the liquid lens module 5 includes a liquid lens 51 and a first lens 52. The liquid lens module 5 is used for enhancing the fluorescent signal when the fluorescent signal penetrates the second wall 621 and the liquid lens 51. The first lens 52 is a condensing lens and is attached to the first wall surface 623 (meaning the bottom wall surface) of the detecting groove 625, whereby the illuminating light generated by the light emitting module 7 penetrates and attaches to the first wall surface When the first lens 52 on the 623, the illuminating light can be concentrated upwards to illuminate the fluorescent sample in the detection tank 625, and excite the fluorescent sample to generate a fluorescent signal.

The liquid lens 51 is attached to the second wall surface 621 (that is, the front wall surface) of the detecting groove 625, and includes a second lens 511, a third lens 513, and a filling arrangement between the second lens 511 and the third lens 513 The liquid layer 512. In this embodiment, the liquid layer 512 is filled with oil, such as mineral oil, but not limited to this, it can also be filled with water, glue, or other liquid materials. And, in some embodiments, the refractive index of the filling oil filled in the liquid layer 512 is 1.46, and it has a difference of at least 0.05 or more from the refractive index of the box material.

In other embodiments, the second lens 511 is used to focus the light in the X direction, and the third lens 513 is used to focus the light in the Y direction. And, because the second lens 511, the liquid layer 512, and the third lens 513 together constitute the liquid lens 51, and the surfaces of the second lens 511 and the third lens 513 can be deformable films, such as thin polymers or soft elastomers. Therefore, when a voltage is applied to the liquid lens 51, the pressure or volume of the liquid layer 512 will correspondingly change, and the curvature of the second lens 511 and/or the third lens 513 can be changed, and the focal position can be adjusted. The optical curvature of the second lens 511 and the third lens 513 with double-cone surface design technology can maximize the transmission of light energy and enhance the performance of the system, and the variable focal length optical device can enhance the fluorescence detection of different wavelengths. Therefore, when the fluorescent signal generated by the fluorescent sample penetrates the second wall 621 of the detection groove 625, the liquid lens 51 concentrates, enhances, and converges the fluorescent signal to the detection module 8. The detection module 8 receives the enhanced fluorescent signal transmitted by the liquid lens 51, and then converts the fluorescent signal into an electrical signal.

Please refer to FIGS. 3 and 4 again, the light emitting module 7 is disposed adjacent to the first wall surface 623 of the detecting groove 625, and the optical axis of the light emitting module 7 is aligned with the first wall surface of the detecting groove 625 623. Therefore, the first lens 52 attached to the first wall surface 623 of the detecting groove 625 can receive the illumination light having a specific wavelength range emitted by the light-emitting module 7. The detection module 8 is adjacently arranged on the second wall 621 of the detection slot 625, and the optical axis of the detection module 8 is aligned with the second wall 621 of the detection slot 625, so the detection module 8 can receive The fluorescent signal penetrates the second wall 621 of the detecting groove 625 and the liquid lens 51.

In one embodiment, the light-emitting module 7 includes a light source 71 and a first filter 72. The light source 71 can be a light emitting diode (LED) or a laser diode (laser diode), and the light source 71 is used to generate illuminating light with a broad wavelength band. The first filter 72 is arranged between the light source 71 and the first wall surface 623, and makes the illumination light of a specific wavelength range generated by the light source 71, for example: the illumination light in the first wavelength range, passes through and prevents the light source 71 The generated illumination light of unnecessary wavelength passes.

The light-emitting module 7 further includes a first pinhole 73 disposed between the light source 71 and the first filter 72, and the first pinhole 73 of the light-emitting module 7 guides the illumination light generated by the light source 71 to the first filter The first wall 623 of the detector 72 and the detecting groove 625. In some embodiments, the diameter of the first pinhole 73 ranges from 2.0 mm to 3.0 mm, but it is not limited thereto.

In some embodiments, the first channel 63 is connected to the detection slot 625 via the corresponding second channel 64, wherein the first channel 63 is used to send the sample to the detection slot 625. The cross-sectional area of the second channel 64 is preferably smaller than the cross-sectional area of the first channel 63, so the second channel 64 has a capillary valve for passive flow control.

In one embodiment, the third wall surface 622 and the first wall surface 623 of the detecting groove 625 are both made of optical film, and the thickness of the optical film on the third wall surface 622 and the thickness of the optical film on the first wall surface 623 can both be between Between 0.1mm and 0.2mm, but not limited to this. The refractive index of the optical film on the third wall surface 622 and the refractive index of the optical film on the first wall surface 623 can both be between 1.3 and 1.6, but not limited thereto.

In some embodiments, the volume of the detection groove 625 of the detection chip 62 can be between 10 μL and 50 μL, but it is not limited thereto. The detection chip 62 may be composed of polycarbonate (PC), polymethyl methacrylate (PMMA) or cyclic olefin copolymer (COC). The refractive index of the detecting groove 625 of the detecting chip 62 can be between 1.3 and 1.6, but is not limited to this.

The detection module 8 includes a second filter 81 and a detector 82. The second filter 81 is used to receive the fluorescent signal transmitted by the liquid lens 51, and allow the fluorescent signal of another specific wavelength range, for example: the fluorescent signal of the second specific wavelength range, to pass, and prevent unnecessary The wavelength passes. The detector 82 is used for receiving the fluorescent signal having the second wavelength range passing through the second filter 81 and converting the fluorescent signal into an electrical signal. In an embodiment, the detector 82 can be, but is not limited to, a photodiode (PD), avalanche photodiode (APD), a charge coupled device (CCD) or a complementary Type metal-oxide semiconductor (complementary metal-oxide semiconductor, CMOS).

The detection module 8 may further include a second pinhole 83. The second pinhole 83 is disposed between the second wall 621 of the detection groove 625 and the second filter 81. The second pinhole of the detection module 8 83 guide the fluorescent signal generated by the fluorescent sample to the detection module 8. In other embodiments, the diameter of the second pinhole 83 ranges from 2.0 mm to 3.0 mm, but it is not limited thereto.

In some embodiments, the multi-color fluorescent excitation and detection device 9 includes a plurality of light-emitting modules 7 and a plurality of detection modules 8, such as, but not limited to, four light-emitting modules 7 and four detection modules. Group 8, where a plurality of light-emitting modules 7 provide different colors of illuminating light to the corresponding detection slot 625 for fluorescent detection, and a plurality of detection modules 8 receive corresponding fluorescent signals, so a plurality of detections The module 8 can detect multiple samples at the same time and realize multiple detection.

Please refer to Figure 6 and Figure 7. Figure 6 shows the excitation spectrum of fluorescent dye FAM, and Figure 76 shows the excitation spectrum of fluorescent dye Cy5. In this example, FAM and Cy5 dyes are used for demonstration. These dyes are standard fluorescent dyes, but the system in this case is not limited to these fluorescent dyes. As shown in Figures 6 and 7, compared with the prior art, the intensity of the fluorescent signal and signal-to-noise ratio detected in this case is higher.

Table 1 shows the comparison between the signal-to-noise ratio of the two fluorescent dyes applied to the multi-color fluorescent excitation and detection device 9 of this case and the conventional technique. The concentrations of the two fluorescent dyes are 320 nM. Table 1 clearly shows that the signal-to-noise ratio of the two fluorescent dyes applied to the multi-color fluorescence excitation and detection device 9 in this case is quite high, even as high as 77 and 30, which shows that the multi-color fluorescence excitation The sensitivity of the detection device 9 is quite good, and the signal from the target fluorescent dye is highly distinctive. Table 1 FAM Cy5 Know-how SNR_320nM 19.9 17.7 Signal-to-noise ratio in this case_320nM 77.10 30.77

In one embodiment, the light-emitting modules 7 are arranged in the accommodating space 14 of the bottom tank 12. During operation, each light-emitting module 7 is located in one of the detection slots 625 of the cassette 6 to provide Effective illumination light for detection. The detection module 8 is arranged on the outer edge of the top tank 11 to realize optical detection, so that the sample can be detected in real time during the process of nucleic acid diffusion. Once the cassette 6 is clamped and locked, the detection module 8 is aligned with one of the detection slots 625 on the cassette 6, so the result of nucleic acid analysis can be read, and the cassette 6 rotates so that Each detection slot 625 passes through different light-emitting modules 7 and detection modules 8 in sequence. In one embodiment, each light-emitting module 7 and detection module 8 can provide a unique color of illuminating light and detection to provide different colors to complete fluorescence detection, so that the nucleic acid analysis device 100 can simultaneously detect multiple Target and achieve multiple detection.

In actual operation, when the optical axis of the light-emitting module 7 or the optical axis of the detection module 8 is aligned with the detection groove 625, some deviation may occur. The experimental results show that when the deviation angle of the fluorescent sample is within ±2 degrees, the signal-to-noise ratio of the system on the target fluorescent dye still maintains a good performance, which means that when the optical axis of the light-emitting module 7 or the detection When the optical axis pair of the module 8 is located in the detection slot 625, the multi-color fluorescent excitation and detection device 9 receives a slight deviation. In some embodiments, the acceptable deviation angle may be within ±3.5 degrees.

In summary, this project provides a multi-color fluorescence excitation and detection device, which can be applied to many fields that use fluorescent dyes as a medium, such as qPCR, isothermal PCR, fluorescent Light microscopy, fluorescence spectroscopy and other fields. The multi-color fluorescence excitation and detection device in this case integrates the light-emitting module, cassette and detection module in a single device, so that the multi-color fluorescence excitation and detection device in this case has a compact structure and a small volume And the advantage of lighter weight. In addition, the multi-color fluorescence excitation and detection device in this case is equipped with liquid lens modules to enhance fluorescence detection signals of different wavelengths, thereby maximizing light energy transmission and improving system performance. Therefore, the multi-color fluorescence excitation and detection device does not require expensive optical components, so the cost of the multi-color fluorescence excitation and detection device is lower. What's more, due to the configuration of multiple light-emitting modules, multiple detection slots, and multiple detection modules, this project can achieve multiple nucleic acid analysis equipment and multiple fluorescent detection. In addition, the signal-to-noise ratio of the multi-color fluorescent excitation and detection device in this case is relatively high, and a slight deviation can be allowed when the cassette rotates.

100: Nucleic acid analysis equipment 1: tank 11: Top tank 12: bottom tank 121: Chamber 13: Hinge 14: Housing space 2: Fluid delivery unit 3: Temperature control unit 4: Rotary drive unit 5: Liquid lens module 51: Liquid lens 511: second lens 512: Liquid layer 513: Third lens 52: The first lens 6: Cassette 61: Reagent storage body 62: Detection chip 625: Detection Slot 623: First Wall 621: second wall 622: Third Wall 624: The Fourth Wall 63: First channel 64: second channel 65: opening 7: Light-emitting module 71: light source 72: first filter 73: The first pinhole 8: Detection module 81: second filter 82: Detector 83: second pinhole 9: Multi-color fluorescence excitation and detection device

Figure 1 is a structural diagram of a nucleic acid analysis device using a multi-color fluorescent excitation and detection device according to an embodiment of the present invention.

Figure 2 is a schematic diagram of the structure of the nucleic acid analysis device shown in Figure 1 when the tank is opened.

Figure 3 is a schematic diagram of the structure of the multi-color fluorescent excitation and detection device of the preferred embodiment of the present invention.

Fig. 4 is a schematic diagram of a partially enlarged structure of the multi-color fluorescent excitation and detection device shown in Fig. 3.

Fig. 5 is a schematic diagram of the structure of the detecting tank and the liquid lens module shown in Fig. 4.

Figure 6 shows the excitation spectrum of the fluorescent dye FAM.

Figure 7 shows the emission spectrum of the fluorescent dye Cy5.

5: Liquid lens module

51: Liquid lens

511: second lens

512: Liquid layer

513: Third lens

52: The first lens

625: Detection Slot

623: First Wall

621: second wall

622: Third Wall

624: The Fourth Wall

Claims (17)

A multi-color fluorescence excitation and detection device, which includes: At least one light emitting module, providing illumination light having a specific wavelength range; A cassette includes a detection chip, the detection chip includes a plurality of detection grooves arranged around the periphery of the detection chip, and each of the detection grooves accommodates a corresponding fluorescent sample, And each of the detection grooves includes a first wall surface and a second wall surface, wherein the illumination light penetrates the first wall surface to illuminate the fluorescent sample in the detection groove and excite the fluorescent sample to Generate a fluorescent signal to penetrate the second wall; A liquid lens module, including at least one liquid lens attached to the second wall surface, so that the fluorescent signal can be enhanced when the fluorescent signal penetrates the second wall surface and the liquid lens; and At least one detection module receives the enhanced fluorescent signal and converts the fluorescent signal into an electrical signal. The multi-color fluorescent excitation and detection device according to claim 1, wherein the light-emitting module is arranged adjacent to the first wall surface, and the optical axis of the light-emitting module is positioned on the first wall surface, and the The detection module is adjacently arranged on the second wall surface, and the optical axis of the detection module is located on the second wall surface. The multi-color fluorescent excitation and detection device according to claim 1, wherein the liquid lens module further includes a first lens, the first lens is attached to the first wall surface and is a condenser lens. The multi-color fluorescent excitation and detection device according to claim 1, wherein the liquid lens module further comprises a second lens, a liquid layer and a third lens, wherein the liquid layer is sandwiched between the second lens And the third lens. The multicolor fluorescent excitation and detection device according to claim 4, wherein the liquid layer is filled with oil, and the surfaces of the second lens and the third lens are composed of deformable films. The multi-color fluorescent excitation and detection device according to claim 5, wherein the refractive index of the liquid layer filled oil is 1.46, and the refractive index of the material of the box body has a difference of 0.05 or more. The multicolor fluorescent excitation and detection device according to claim 1, wherein the first wall surface is a bottom wall surface, and the second wall surface is a front wall surface. The multicolor fluorescent excitation and detection device according to claim 7, wherein each of the detection grooves further includes a third wall, a fourth wall, a fifth wall, and a sixth wall, wherein the third wall The wall surface is opposite to the first wall surface, the second wall surface is opposite to the fourth wall surface, the fifth wall surface is opposite to the sixth wall surface, wherein the third wall surface is an upper wall surface, and the fourth wall surface is A rear wall surface, the fifth wall surface is a first side wall, and the sixth wall surface is a second side wall. The multicolor fluorescent excitation and detection device according to claim 8, wherein the third wall surface and the first wall surface are respectively composed of an optical film, wherein the thickness of the optical film on the third wall surface and the first wall surface The thickness of the optical film on the wall is between 0.1mm and 0.2mm, and the refractive index of the optical film on the third wall and the refractive index of the optical film on the first wall are respectively between 1.3 and 1.6 between. The multi-color fluorescent excitation and detection device according to claim 1, wherein the light-emitting module includes: A light source that generates the illuminating light with a broad wavelength band; and A first filter is arranged between the light source and the first wall surface, and allows the illumination light in the first wavelength range to pass. The multi-color fluorescent excitation and detection device according to claim 10, wherein the light-emitting module further comprises a first pinhole disposed between the light source and the first filter, wherein the first pinhole The aperture size is between 2.0mm and 3.0mm. The multi-color fluorescence excitation and detection device according to claim 1, wherein the volume of the detection tank is between 10 μL and 50 μL. The multicolor fluorescence excitation and detection device according to claim 1, wherein the detection chip is composed of polycarbonate, polymethyl methacrylate or cyclic olefin copolymer, and the refraction of the detection groove The rate is between 1.3 and 1.6. The multi-color fluorescent excitation and detection device according to claim 1, wherein the detection module includes: A second filter that receives the fluorescent signal and allows the fluorescent signal in the second wavelength range to pass through; and A detector receives the fluorescent signal with a second wavelength range, and converts the fluorescent signal into the electrical signal. The multi-color fluorescence excitation and detection device according to claim 14, wherein the detection module further includes a second pinhole disposed between the second wall surface and the second filter, wherein the second The aperture size of the pinhole is between 2.0mm and 3.0mm. The multi-color fluorescence excitation and detection device according to claim 14, wherein the detector is a photodiode, an avalanche photodiode, a photosensitive coupling element or a complementary metal oxide semiconductor. The multi-color fluorescence excitation and detection device according to claim 1, wherein the multi-color fluorescence excitation and detection device is applied to at least one of qPCR, isothermal PCR, fluorescence microscopy, and fluorescence spectroscopy.

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