A microfluidic Cryptococcus sorting device and detection method

Abstract

The present invention discloses a microfluidic Cryptococcus sorting device and a detection method. The device includes: a focusing section for receiving a fluid solution and focusing the particles in the fluid solution; a sinking section for receiving the fluid solution after the focused particles and performing stratification; a liquid separation section for separating cerebrospinal fluid from the stratified fluid solution; a capture member for receiving the fluid solution after separating the cerebrospinal fluid and capturing the particles therein; an outlet section for discharging the fluid solution in the capture member; the focusing section is serrated and is composed of a plurality of inverted V-shaped components. A focusing channel is arranged in each inverted V-shaped component, and adjacent two inverted V-shaped components are connected in series through a common end point and the internal focusing channels are communicated; through the unique structure provided by the present invention, precise focusing, separation and enrichment of Cryptococcus can be achieved in a short time, so as to achieve the purpose of detection.

Description

Technical Field

[0001] The present invention relates to the technical field of medical equipment, and in particular to a microfluidic cryptococcal sorting device and a detection method. Background Art

[0002] Cryptococcal meningitis (CM) is a serious infectious disease caused by Cryptococcus neoformans and Cryptococcus gattii, which is mainly seen in patients with impaired immune function, especially in those infected with HIV. The early symptoms of CM are usually hidden, and its diagnosis is not only crucial for the management and treatment of the disease, but also can assist in HIV screening to a certain extent. However, due to the high mortality rate of CM and the lack of specificity of symptoms, clinical diagnosis faces great challenges and is prone to misdiagnosis.

[0003] Common symptoms of CM patients include chronic or subacute headaches, positive meningeal irritation signs, and significantly elevated cerebrospinal fluid pressure. However, these symptoms overlap with the manifestations of other central nervous system diseases and lack specificity. In addition, the biological characteristics of Cryptococcus also make diagnosis difficult. For example, the thick polysaccharide capsule of Cryptococcus accounts for about 70% of its total cell mass, and the cell wall structure is tough. These characteristics make the traditional nucleic acid extraction and detection process complicated and inefficient.

[0004] At present, traditional detection methods for CM mainly include direct examination and isolation and culture. Direct examination such as ink smear microscopy requires the collection of samples by centrifugation and precipitation before testing. Although this method is simple and fast, the sample is easily lost, the sensitivity is low, and the false negative rate is high. Although the isolation and culture method is considered the "gold standard" for CM detection, its diagnostic cycle usually takes 2 to 7 days, which cannot meet the needs of clinical rapid diagnosis.

[0005] Microfluidics is an emerging biomedical detection method. By designing precise fluid channels at the microscopic scale, microfluidics can effectively achieve functions such as sample separation, enrichment, and particle focusing. Microfluidics has shown great potential in the fields of cell counting, biomolecule detection, sample sorting, and biosensor optimization. Compared with traditional detection methods, microfluidics has the advantages of low cost, low energy consumption, and efficient operation, and can achieve precise control of droplets, cells, and microorganisms at the microscopic scale.

[0006] In microfluidics, particle focusing is an important function. Its core goal is to arrange dispersed particles into regular single or multi-column sequences to meet the requirements of subsequent operations, such as the enrichment, separation, or manipulation of target particles. According to the operating principle, microfluidic particle focusing methods can be divided into three categories: active focusing, sheath-assisted focusing, and passive focusing. Active focusing relies on external forces such as acoustic waves, magnetic fields, or optical forces to manipulate particle movement. Although it has high precision, it usually requires a complex external energy field control system and can only ensure sufficient focusing under low flow rate conditions, which limits the flux improvement. Sheath-assisted focusing compresses the target sample into a narrow single sheath flow through an additional sheath flow, thus achieving high-throughput focusing. However, it requires additional microchannel structures and fluid input devices, reducing the integration of microfluidic devices. Passive focusing, on the other hand, realizes natural particle focusing without external driving forces through the inertia of the fluid, the viscoelasticity of non-Newtonian fluids, or the micro-vortex effect induced by the channel structure. This method can effectively avoid cell damage during cell operation and can also achieve continuous and high-throughput particle operation. In the application of small particle focusing, due to the significant change in the cross-sectional width at the channel corner, the Dean drag force (FD) plays a dominant role, accelerating the focusing of particles towards the center of the channel along the counter-rotating vortex streamlines. Among them, the Dean drag force tends to entrain particles to follow the counter-rotating vortex streamlines.

[0007] From the perspective of cell sorting, sorting techniques can be mainly divided into two categories: label-free sorting and labeled sorting. Label-free sorting differentiates based on the physical characteristics of cells (such as size, density, and shape, etc.), but it is difficult to distinguish cells with similar physical properties but different biological characteristics. Labeled sorting, on the other hand, realizes the identification and separation of cells by introducing specific markers (such as fluorescent dyes or magnetic beads). However, this method may have a certain impact on the activity of cells, and due to the need for multiple cell washes during the operation, the capture efficiency will be reduced. For the focusing of small particles, the significant increase in the cross-sectional width at the channel corner makes the Dean drag force increase and dominate, and the particles focus towards the center of the channel faster.

[0008] In the existing CM detection methods, the direct inspection method is complex in operation, the sample is easily lost, the sensitivity is low, and the false negative rate is high. The isolation and culture method has a long diagnostic cycle. Therefore, a new technology is needed to improve this phenomenon. Summary of the Invention

[0009] Aiming at the deficiencies of the prior art, the present invention provides a microfluidic Cryptococcus sorting device and detection method, aiming to solve the problems in the background technology.

[0010] To achieve the above object, the present invention provides the following technical solution: A microfluidic Cryptococcus sorting device, comprising:

[0011] Focusing section, for receiving a fluid solution and focusing the particles in the fluid solution;

[0012] Sinking section, which is connected to the output end of the serrated-like component, for receiving the fluid solution after focusing the particles and performing layering;

[0013] Liquid separation section, which is connected to and communicates with the sinking section, for separating cerebrospinal fluid from the layered fluid solution in the sinking section;

[0014] Transition section, which is connected to the liquid separation section, for receiving the fluid solution after separating cerebrospinal fluid in the liquid separation section and transmitting it;

[0015] Capture member, which is connected to the transition section, for receiving the fluid solution transmitted by the transition section and capturing the particles in the fluid solution;

[0016] Outlet section, which is connected to the capture member, for discharging the fluid solution in the capture member;

[0017] The focusing section is serrated-like and composed of a plurality of inverted V-shaped components. A focusing channel is provided in each inverted V-shaped component. Adjacent two inverted V-shaped components are connected in series through a common end point and the internal focusing channels are communicated with each other.

[0018] Furthermore, each inverted V-shaped component is composed of an isosceles trapezoid component with a missing lower base and four isosceles trapezoid components with a missing upper base; two obliquely arranged isosceles trapezoid components with a missing upper base are respectively connected to the two ends of the bottom of the isosceles trapezoid component with a missing lower base in sequence, and two adjacent and connected isosceles trapezoid components with a missing upper base are inverted with each other;

[0019] One first connection end is respectively provided at both ends of the missing part of the isosceles trapezoid component with a missing lower base, and one second connection end is respectively provided at both ends of the missing part of the isosceles trapezoid component with a missing upper base; the specific connection relationship between the isosceles trapezoid component with a missing lower base and the isosceles trapezoid component with a missing upper base is: the two first connection ends of the isosceles trapezoid component with a missing lower base are respectively connected to one second connection end of an isosceles trapezoid component with a missing upper base, and the other second connection end of the isosceles trapezoid component with a missing upper base is connected to one second connection end of an adjacent isosceles trapezoid component with a missing upper base;

[0020] Adjacent inverted V-shaped components are connected by an inverted isosceles trapezoid component with a missing lower base, that is, one unconnected second connection end of the isosceles trapezoid component with a missing upper base at the bottom of two adjacent inverted V-shaped components is respectively connected to the two first connection ends of the inverted isosceles trapezoid component with a missing lower base.

[0021] Further, one second connection end of the isosceles trapezoidal component with the upper base missing at the bottom in the first inverted V-shaped component in the focusing section, which is not connected, is connected to one third connection end of the isosceles trapezoidal component with half of the lower base missing. The other third connection end of the isosceles trapezoidal component with half of the lower base missing is used as the entrance of the focusing section, that is, the entrance of the sorting device.

[0022] Focusing channels are provided in the isosceles trapezoidal component with the lower base missing, the isosceles trapezoidal component with the upper base missing, and the isosceles trapezoidal component with half of the lower base missing, and they are connected and communicated with each other.

[0023] Further, the corner angle range of the focusing channel in the isosceles trapezoidal component with the upper base missing is set to 55° - 75°.

[0024] Let the width of the focusing channels in the isosceles trapezoidal component with the lower base missing, the isosceles trapezoidal component with the upper base missing, and the isosceles trapezoidal component with half of the lower base missing be Z, the inner wall length of the focusing channel in the isosceles trapezoidal component with the lower base missing be S, and the inner wall length of the focusing channel in the isosceles trapezoidal component with the upper base missing be L, and it satisfies L = S + Z. Among them, the widths of the focusing channel, the sinking channel, and the transition channel are all Z, and the width of the liquid separation channel satisfies 1.5Z.

[0025] Further, a sinking channel is provided in the sinking section. An inner cavity is provided in the sinking channel. The sinking channel is connected to one second connection end of the isosceles trapezoidal component with the upper base missing at the bottom in the last inverted V-shaped component in the focusing section, which is not connected, and is connected and communicated with the focusing channel in this isosceles trapezoidal component with the upper base missing. From the input end to the output end of the inner cavity of the sinking channel, the first layer of steps, the second layer of steps, the third layer of steps, the fourth layer of steps, the fifth layer of steps, the sixth layer of steps, and the seventh layer of steps are arranged in sequence. Among them, the bottom of the inner cavity of the sinking channel is used as the 0th layer of steps. It descends layer by layer from the 0th layer of steps to the 5th layer of steps and ascends layer by layer from the 5th layer of steps to the 7th layer of steps. Among them, the first layer of steps, the second layer of steps, the third layer of steps, the fourth layer of steps, the fifth layer of steps, the sixth layer of steps, and the seventh layer of steps are set with the same width.

[0026] The sixth layer of steps is flush with the 0th layer of steps. Let the vertical distance from the 5th layer of steps to the 0th layer of steps be A, and the vertical distance from the 5th layer of steps to the 7th layer of steps be B, and it satisfies B > A.

[0027] The sinking channel is connected to one second connection end of the isosceles trapezoidal component with the upper base missing at the bottom in the last inverted V-shaped component in the focusing section through a plurality of successively connected and mutually inverted isosceles trapezoidal components with the upper base missing, and is connected and communicated with the focusing channel in this isosceles trapezoidal component with the upper base missing.

[0028] Further, there are two liquid separation sections. The input ends of the two liquid separation sections are respectively connected to both sides of the sinking section, and the output ends of the two liquid separation sections are connected to the outside; the two liquid separation sections are on the same axis; liquid separation channels are arranged in both liquid separation sections, and the liquid separation channels are connected to the sinking channel in the sinking section; the connection points between the liquid separation section and the sinking section are at the top of the fifth layer of steps in the sinking channel.

[0029] Further, a transition channel is arranged in the transition section, and the transition channel in the transition section is communicated with the sinking channel in the sinking section.

[0030] Further, a capture cavity is arranged in the capture part. The capture cavity has a semi-circular rectangular contour, that is, the capture cavity is formed by splicing a rectangular cavity and a semi-circular cavity; a vertical angled baffle array composed of several angled baffles is arranged in the capture cavity, and the openings of all the angled baffles in the vertical angled baffle array face the entrance end of the capture part; the rectangular cavity in the capture cavity is communicated with the transition channel.

[0031] The connection point between the transition section and the capture part is set on the central axis of the capture part; a depression is arranged at the connection between the capture part and the transition section, and the depth of the depression accounts for 1 / 5 - 1 / 4 of the longest distance between the two ends of the capture part.

[0032] Further, an outlet channel is arranged in the outlet section. One end of the outlet channel is communicated with the semi-circular cavity in the capture cavity, and the other end of the outlet channel penetrates through to the outside.

[0033] A microfluidic cryptococcus detection method is applied to a microfluidic cryptococcus sorting device, and includes the following steps:

[0034] S1: Mix the cerebrospinal fluid sample to be detected with acetic acid and ink to obtain a fluid solution.

[0035] S2: Inject the fluid solution into the entrance of the focusing section, and drive the fluid solution through the focusing channels in each isosceles trapezoid component with the lower base missing, isosceles trapezoid component with the upper base missing, and isosceles trapezoid component with half of the lower base missing in the focusing section, so that the particles in the fluid solution are focused.

[0036] Drive the fluid solution after focusing the particles through the sinking channel, and layer the fluid solution after focusing the particles, so that the cerebrospinal fluid in the fluid solution is focused on the upper layer of the sinking channel and discharged through the transition channel in the transition section, and the particles are focused on the lower layer of the sinking channel.

[0037] Drive the fluid solution after separating the cerebrospinal fluid to flow into the capture cavity through the transition section, and the fluid solution is enriched by the vertical angled baffle array.

[0038] S3: Observe through a microscope whether the particles in the capture cavity contain cryptococcus.

[0039] After all the fluid solution enters the focusing section, a syringe is used to continue injecting ink into the focusing section to drive all the fluid solution into the capture cavity.

[0040] Compared with the existing technologies, the present invention has the following beneficial effects: by combining the isosceles trapezoid structure with the lower base missing and the isosceles trapezoid structure with the upper base missing to form a quasi-inverted V-shaped structure, the zigzag particle focusing structure composed of multiple quasi-inverted V-shaped structures can better focus the fluid solution to the middle of the focusing channel. By arranging multiple steps with different heights in the inner cavity of the sinking channel, the cerebrospinal fluid and cryptococcus in the fluid solution can be stratified by the action of gravity stratification. By connecting a liquid separation channel in the sinking channel, the cerebrospinal fluid in the fluid solution can be separated; by arranging a vertical folding baffle array in the capture cavity, a larger eddy area can be generated when the fluid solution flows into the capture cavity. At the same time, the particles can also be effectively preserved in the capture cavity, which is convenient for subsequent observation through a microscope to check whether there is cryptococcus in the capture cavity. There is no need for centrifugation, the operation is simple, the sample is not easily lost, the sensitivity is high, and the false negative rate is low. It can achieve precise focusing, separation and enrichment of cryptococcus in a short time so as to achieve the purpose of detection. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 It is the overall structure diagram of the present invention.

[0042] Figure 2 It is the structure diagram of the quasi-inverted V-shaped component of the present invention.

[0043] Figure 3 It is the schematic diagram of the connection structure of the quasi-inverted V-shaped component of the present invention.

[0044] Figure 4 It is the comparison diagram of the focusing effects of four structures in series, namely, 5-cycle arc, rectangle, inverted V-shaped, and isosceles trapezoid with the lower base missing, in the embodiment of the present invention.

[0045] Figure 5 It is the velocity cloud diagram of the fluid solution in the quasi-inverted V-shaped component.

[0046] Figure 6 It is the cross-sectional view of the sinking channel of the present invention.

[0047] Figure 7 It is the schematic diagram of the connection structure between the sinking channel and the liquid separation channel of the present invention.

[0048] Figure 8 It is the schematic diagram of the internal structure of the capture part of the present invention.

[0049] In the figure, 1 is a quasi-inverted V-shaped component; 101 is an isosceles trapezoid component with a missing lower base; 1011 is the first connection end; 102 is an isosceles trapezoid component with a missing upper base; 1021 is the second connection end; 103 is an isosceles trapezoid component with a half-missing lower base; 1031 is the third connection end; 4 is the sinking section; 401 is the sinking channel; 411 is the first layer of steps; 412 is the second layer of steps; 413 is the third layer of steps; 414 is the fourth layer of steps; 415 is the fifth layer of steps; 416 is the sixth layer of steps; 417 is the seventh layer of steps; 5 is the liquid separation section; 6 is the transition section; 7 is the capture member; 71 is the capture cavity; 72 is the vertical angled baffle array; 721 is the angled baffle; 8 is the outlet section. Detailed implementation mode

[0050] As Figures 1 - 3 shown, the present invention provides a technical solution: a microfluidic cryptococcus sorting device, comprising:

[0051] A focusing section for receiving a fluid solution and focusing particles (cells, cryptococcus, etc.) in the fluid solution; the fluid solution is obtained by mixing a cerebrospinal fluid sample that may contain cryptococcus with acetic acid and ink, and the acetic acid is used to rupture red blood cells that may exist in the cerebrospinal fluid sample to reduce interference.

[0052] A sinking section 4, the sinking section 4 is connected to the output end of the quasi-serrated component, and is used for receiving the fluid solution after focusing the particles and performing layering.

[0053] A liquid separation section 5, the liquid separation section 5 is connected to and communicates with the sinking section 4, and is used for separating cerebrospinal fluid in the layered fluid solution in the sinking section 4.

[0054] A transition section 6, the transition section 6 is connected to the liquid separation section 5, and is used for receiving the fluid solution after the liquid separation section 5 separates the cerebrospinal fluid and transmitting it.

[0055] A capture member 7, the capture member 7 is connected to the transition section 6, and is used for receiving the fluid solution transmitted by the transition section 6 and capturing particles in the fluid solution.

[0056] An outlet section 8, the outlet section 8 is connected to the capture member 7, and is used for discharging the fluid solution in the capture member 7.

[0057] Among them, the focusing section is quasi-serrated, and is composed of a plurality of quasi-inverted V-shaped components 1. A focusing channel is arranged in each quasi-inverted V-shaped component 1. In this embodiment, 3 quasi-inverted V-shaped components 1 are provided, and adjacent two quasi-inverted V-shaped components 1 are connected in series through a common end point and the internal focusing channels are communicated with each other.

[0058] Among them, each inverted V-shaped component 1 is composed of an isosceles trapezoid component 101 with a missing lower base and four isosceles trapezoid components 102 with a missing upper base; at both ends of the bottom of the isosceles trapezoid component 101 with a missing lower base, two obliquely arranged isosceles trapezoid components 102 with a missing upper base are sequentially connected, and two adjacent and interconnected isosceles trapezoid components 102 with a missing upper base are inverted with respect to each other.

[0059] Among them, at both ends of the missing part of the isosceles trapezoid component 101 with a missing lower base, a first connection end 1011 is respectively provided, and at both ends of the missing part of the isosceles trapezoid component 102 with a missing upper base, a second connection end 1021 is respectively provided.

[0060] The specific connection relationship between the isosceles trapezoid component 101 with a missing lower base and the isosceles trapezoid component 102 with a missing upper base is as follows: the two first connection ends 1011 of the isosceles trapezoid component 101 with a missing lower base are respectively connected to a second connection end 1021 of an isosceles trapezoid component 102 with a missing upper base, and the other second connection end 1021 of the isosceles trapezoid component 102 with a missing upper base is connected to a second connection end 1021 of an adjacent isosceles trapezoid component 102 with a missing upper base.

[0061] Among them, adjacent inverted V-shaped components 1 are connected by an inverted isosceles trapezoid component 101 with a missing lower base, that is, the unconnected second connection ends 1021 of the isosceles trapezoid components 102 with a missing upper base at the bottom in two adjacent inverted V-shaped components 1 are respectively connected to the two first connection ends 1011 of the inverted isosceles trapezoid component 101 with a missing lower base.

[0062] Among them, an unconnected second connection end 1021 of the isosceles trapezoid component 102 with a missing upper base at the bottom in the first inverted V-shaped component 1 in the focusing section is connected to a third connection end 1031 on the half isosceles trapezoid component 103 with a missing lower base, and the other third connection end 1031 of the half isosceles trapezoid component 103 with a missing lower base is used as the entrance of the focusing section, that is, the entrance of the sorting device.

[0063] Among them, focusing channels are provided in the isosceles trapezoid component 101 with a missing lower base, the isosceles trapezoid component 102 with a missing upper base, and the half isosceles trapezoid component 103 with a missing lower base and are connected and communicated with each other.

[0064] By optimizing the angle of the focusing channel, it is found that the smaller the channel angle, the better the focusing effect, but the channel pressure drop increases. Therefore, 60° with a better focusing effect and a moderate pressure drop is selected as the optimal solution.

[0065] Among them, the corner angle range of the inner focusing channel of the isosceles trapezoidal component 102 with the upper base missing can be set to 55° to 75°, and the optimal corner angle of the inner focusing channel of the isosceles trapezoidal component 102 with the upper base missing is 60°.

[0066] Let the width of the inner focusing channels of the isosceles trapezoidal component 101 with the lower base missing, the isosceles trapezoidal component 102 with the upper base missing, and the isosceles trapezoidal component 103 with half of the lower base missing be Z, the inner wall length of the inner focusing channel of the isosceles trapezoidal component 101 with the lower base missing be S, and the inner wall length of the inner focusing channel of the isosceles trapezoidal component 102 with the upper base missing be L, satisfying L = S + Z; among them, the widths of the focusing channel, the sinking channel 401, and the transition channel are all Z, and the width of the liquid separation channel satisfies 1.5Z, so as to reduce the kinetic energy loss of the fluid solution after liquid separation in the liquid separation channel, so that it flows into the capture chamber 71 at a slower speed.

[0067] As Figure 4 shown, when the channel inlet velocities are all set to 0.1 m / s and the cross-sectional areas are the same, according to the comparison of the focusing effects of four series-connected structures of circular arc, rectangle, inverted V-shaped, and isosceles trapezoid with the base missing, the focusing effect of the isosceles trapezoid with the base missing is the best, and the inverted V-shaped is the second best; as Figure 5 shown, from the velocity distribution of the fluid solution at the corner of the inner channel of the inverted V-shaped and the inner channel of the isosceles trapezoidal structure with the base missing, due to the existence of wall shear stress, the fluid velocity close to the channel sidewall is relatively low, while the velocity in the central region of the channel is relatively high, resulting in particles (such as cells or cryptococcus, etc.) gradually concentrating in the central region of the channel to form a streamline arrangement during the flow process, and a significant eddy current region is formed at the corner of the channel of the isosceles trapezoidal structure with the base missing. In this region, the fluid generates a rotational motion under the action of Dean drag force, and the rotational and accelerating motions of the fluid solution enhance the collision and friction effects of the particles. When the fluid solution flows through these local corners, the magnitude and direction of the flow velocity change sharply, resulting in continuous changes in the velocity distribution law on the cross-section and generating additional friction; the setting of the isosceles trapezoidal structure with the base missing enables the flow velocity direction of the fluid solution to be adjusted multiple times, and at the same time, the eddy current region at the corner overlaps with the main flow velocity distribution region, further strengthening the friction effect of the fluid.

[0068] As Figures 6 - 7As shown, a sinking channel 401 is provided in the sinking section 4, and an inner cavity is provided in the sinking channel 401. The sinking channel 401 is connected to a second connection end 1021 that is not connected to the isosceles trapezoidal component 102 with the upper base missing at the bottom in the last inverted V-shaped component 1 in the focusing section, and is communicated with the focusing channel in the isosceles trapezoidal component 102 with the upper base missing; a first layer of steps 411, a second layer of steps 412, a third layer of steps 413, a fourth layer of steps 414, a fifth layer of steps 415, a sixth layer of steps 416, and a seventh layer of steps 417 are sequentially arranged from the input end to the output end of the inner cavity of the sinking channel 401. Among them, the bottom of the inner cavity of the sinking channel 401 is used as the 0th layer of steps, and it descends layer by layer from the 0th layer of steps to the fifth layer of steps 415, and ascends layer by layer from the fifth layer of steps 415 to the seventh layer of steps 417; among them, the first layer of steps 411, the second layer of steps 412, the third layer of steps 413, the fourth layer of steps 414, the fifth layer of steps 415, the sixth layer of steps 416, and the seventh layer of steps 417 are arranged with the same width.

[0069] Among them, the sixth layer of steps 416 is flush with the 0th layer of steps. Let the vertical distance from the fifth layer of steps 415 to the 0th layer of steps be A, and the vertical distance from the fifth layer of steps 415 to the seventh layer of steps 417 be B, and B > A is satisfied.

[0070] Among them, the sinking channel 401 is connected to a second connection end 1021 that is not connected to the isosceles trapezoidal component 102 with the upper base missing at the bottom in the last inverted V-shaped component 1 in the focusing section through a plurality of isosceles trapezoidal components 102 with the upper base missing that are sequentially connected and inverted, and is communicated with the focusing channel in the isosceles trapezoidal component 102 with the upper base missing.

[0071] Among them, there are two liquid separation sections 5. The input ends of the two liquid separation sections 5 are respectively connected to both sides of the sinking section 4, and the output ends of the two liquid separation sections 5 are connected to the outside; the two liquid separation sections 5 are on the same axis; liquid separation channels are provided in both liquid separation sections 5, and the liquid separation channels are connected to the sinking channel 401 in the sinking section 4; the connection points of the liquid separation section 5 and the sinking section 4 are at the top of the fifth layer of steps 415 in the sinking channel 401.

[0072] By setting five consecutive descending steps and two ascending steps, particles (such as cells, cryptococcus, etc.) can mainly converge on the lower side in the sinking channel 401 after passing through the sinking channel 401. When separated through the liquid separation channel, the particles (such as cells, cryptococcus, etc.) in the fluid solution maintain linear motion due to inertia, while the cerebrospinal fluid in the fluid solution is separated into the liquid separation channels on both sides, achieving an enrichment effect.

[0073] Among them, a transition channel is provided in the transition section 6, and the transition channel in the transition section 6 is communicated with the sinking channel 401 in the sinking section 4.

[0074] As shown Figure 8 in the figure, a capture cavity 71 is provided inside the capture member 7. The capture cavity 71 has a semi-rectangular contour, that is, the capture cavity 71 is formed by splicing a rectangular cavity and a semi-circular cavity. The capture cavity 71 composed of the rectangular cavity and the semi-circular cavity has the best eddy current effect, and it is easy for particles to enter but difficult to exit. Inside the capture cavity 71, there is a vertical angled baffle array 72 composed of several angled baffles 721. The openings of all the angled baffles 721 in the vertical angled baffle array 72 face the entrance end of the capture member 7. The rectangular cavity inside the capture cavity 71 is communicated with the transition channel. Through the design of the vertical angled baffle array 72, the fluid solution has a large eddy current area after entering the capture cavity 71, so that particle cells, cryptococcus, etc. can be effectively stored in the capture cavity 71.

[0075] Among them, the connection point between the transition section 6 and the capture member 7 is set on the central axis of the capture member 7; there is a depression at the connection between the capture member 7 and the transition section 6, and the depth of the depression accounts for 1 / 5 - 1 / 4 of the longest distance between the two ends of the capture member 7.

[0076] Among them, an outlet channel is provided inside the outlet section 8. One end of the outlet channel is communicated with the semi-circular cavity inside the capture cavity 71, and the other end of the outlet channel communicates with the outside.

[0077] A microfluidic cryptococcus detection method includes the following steps:

[0078] S1: Mix a cerebrospinal fluid sample that may contain cryptococcus with acetic acid and ink to obtain a fluid solution. Acetic acid is used to rupture the possible red blood cells in the cerebrospinal fluid sample to reduce interference.

[0079] S2: Inject the fluid solution into the entrance of the focusing section at a speed of 0.1 m / s through a micropump, and drive the fluid solution through the focusing channels in each isosceles trapezoid component 101 with the lower base missing, isosceles trapezoid component 102 with the upper base missing, and isosceles trapezoid component 103 with half of the lower base missing in the focusing section, so that the particles (cells, cryptococcus, etc.) in the fluid solution are focused.

[0080] Drive the fluid solution after focusing the particles through the sinking channel to layer the fluid solution after focusing the particles, so that the cerebrospinal fluid in the fluid solution is focused on the upper layer of the sinking channel and discharged through the transition channel in the transition section 6, and the particles (cells, cryptococcus, etc.) are focused on the lower layer of the sinking channel.

[0081] Drive the fluid solution after separating the cerebrospinal fluid to flow into the capture cavity through the transition section, and the fluid solution is enriched by the vertical angled baffle array.

[0082] S3: Observe through a microscope whether the particles in the capture cavity contain cryptococcus.

[0083] Wherein, after all the fluid solution enters the focusing section, a syringe is used to continue injecting ink into the focusing section to drive all the fluid solution into the capture cavity.

[0084] Although embodiments of the present invention have been shown and described, it will be understood by those of ordinary skill in the art that various changes, modifications, substitutions and variations can be made to these embodiments without departing from the principles and spirit of the present invention, and the scope of the present invention is defined by the appended claims and their equivalents.

Claims

1. A microfluidic Cryptococcus sorting device, characterized in that Comprising: A focusing section for receiving a fluid solution and focusing the particles in the fluid solution; A sinking section (4) connected to the output end of the serrated-like component for receiving the fluid solution after focusing the particles and performing layering; A liquid separation section (5) connected to and communicating with the sinking section (4) for separating cerebrospinal fluid from the layered fluid solution in the sinking section (4); A transition section (6) connected to the liquid separation section (5) for receiving the fluid solution after separating cerebrospinal fluid in the liquid separation section (5) and transmitting it; A capture member (7) connected to the transition section (6) for receiving the fluid solution transmitted by the transition section (6) and capturing the particles in the fluid solution; An outlet section (8) connected to the capture member (7) for discharging the fluid solution in the capture member (7); The focusing section is serrated-like and composed of a plurality of inverted V-shaped components (1). A focusing channel is provided in each inverted V-shaped component (1), and adjacent inverted V-shaped components (1) are connected in series through a common end point and the internal focusing channels are connected; Each inverted V-shaped component (1) is composed of an isosceles trapezoid component (101) with a missing lower base and four isosceles trapezoid components (102) with a missing upper base; Two obliquely arranged isosceles trapezoid components (102) with a missing upper base are sequentially connected to both ends of the bottom of the isosceles trapezoid component (101) with a missing lower base, and two adjacent and connected isosceles trapezoid components (102) with a missing upper base are inverted with respect to each other; A first connection end (1011) is provided at each of the two ends of the missing part of the isosceles trapezoid component (101) with a missing lower base, and a second connection end (1021) is provided at each of the two ends of the missing part of the isosceles trapezoid component (102) with a missing upper base. The specific connection relationship between the isosceles trapezoid component (101) with a missing lower base and the isosceles trapezoid component (102) with a missing upper base is as follows: The two first connection ends (1011) of the isosceles trapezoid component (101) with a missing lower base are respectively connected to a second connection end (1021) of an isosceles trapezoid component (102) with a missing upper base, and the other second connection end (1021) of the isosceles trapezoid component (102) with a missing upper base is connected to a second connection end (1021) of an adjacent isosceles trapezoid component (102) with a missing upper base; Adjacent inverted V-shaped components (1) are connected by an inverted isosceles trapezoid component (101) with a missing lower base, that is, the unconnected second connection ends (1021) of the isosceles trapezoid components (102) with a missing upper base at the bottom of two adjacent inverted V-shaped components (1) are respectively connected to the two first connection ends (1011) of the inverted isosceles trapezoid component (101); Connect a second connection end (1021) that is not connected on the upper base missing isosceles trapezoid component (102) at the bottom in the first inverted V-shaped component (1) within the focusing section to a third connection end (1031) on the half lower base missing isosceles trapezoid component (103), and use the other third connection end (1031) of the half lower base missing isosceles trapezoid component (103) as the entrance of the focusing section, that is, the entrance of the sorting device; Focusing channels are provided in the lower base missing isosceles trapezoid component (101), the upper base missing isosceles trapezoid component (102), and the half lower base missing isosceles trapezoid component (103) and are connected and communicated; The corner angle range of the focusing channel in the upper base missing isosceles trapezoid component (102) is set to 55° - 75°; Let the width of the focusing channels in the lower base missing isosceles trapezoid component (101), the upper base missing isosceles trapezoid component (102), and the half lower base missing isosceles trapezoid component (103) be Z, the inner wall length of the focusing channel in the lower base missing isosceles trapezoid component (101) be S, and the inner wall length of the focusing channel in the upper base missing isosceles trapezoid component (102) be L, satisfying L = S + Z; among them, the widths of the focusing channel, the sinking channel (401), and the transition channel are all Z, and the width of the liquid separation channel satisfies 1.5Z.

2. The microfluidic Cryptococcus sorting device according to claim 1, wherein: A sinking channel (401) is provided in the sinking section (4), an inner cavity is provided in the sinking channel (401), the sinking channel (401) is connected to a second connection end (1021) that is not connected on the upper base missing isosceles trapezoid component (102) at the bottom in the last inverted V-shaped component (1) within the focusing section and is connected and communicated with the focusing channel in the upper base missing isosceles trapezoid component (102); from the input end to the output end of the inner cavity of the sinking channel (401), a first layer of steps (411), a second layer of steps (412), a third layer of steps (413), a fourth layer of steps (414), a fifth layer of steps (415), a sixth layer of steps (416), and a seventh layer of steps (417) are sequentially provided. Among them, the bottom of the inner cavity of the sinking channel (401) is used as the 0th layer of steps, and it descends layer by layer from the 0th layer of steps to the fifth layer of steps (415), and ascends layer by layer from the fifth layer of steps (415) to the seventh layer of steps (417); among them, the first layer of steps (411), the second layer of steps (412), the third layer of steps (413), the fourth layer of steps (414), the fifth layer of steps (415), the sixth layer of steps (416), and the seventh layer of steps (417) are set to be of equal width; The sixth layer of steps (416) is flush with the 0th layer of steps. Let the vertical distance from the fifth layer of steps (415) to the 0th layer of steps be A, and the vertical distance from the fifth layer of steps (415) to the seventh layer of steps (417) be B, satisfying B > A; The sinking channel (401) is connected to a second connection end (1021) that is not connected to the upper-bottom missing isosceles trapezoid component (102) at the bottom of the last inverted-V-shaped component (1) in the focusing section through a plurality of successively connected and inverted upper-bottom missing isosceles trapezoid components (102), and is in communication with the focusing channel in the upper-bottom missing isosceles trapezoid component (102).

3. A microfluidic Cryptococcus sorting device according to claim 2, characterized in that: There are two liquid separation sections (5). The input ends of the two liquid separation sections (5) are respectively connected to both sides of the sinking section (4), and the output ends of the two liquid separation sections (5) are connected to the outside; the two liquid separation sections (5) are on the same axis; liquid separation channels are provided in both liquid separation sections (5), and the liquid separation channels are connected to the sinking channel (401) in the sinking section (4); the connection point between the liquid separation section (5) and the sinking section (4) is at the top of the 5th step (415) in the sinking channel (401).

4. A microfluidic Cryptococcus sorting device according to claim 3, characterized in that: A transition channel is provided in the transition section (6), and the transition channel in the transition section (6) is in communication with the sinking channel (401) in the sinking section (4).

5. The microfluidic Cryptococcus sorting device according to claim 4, wherein: A capture cavity (71) is provided in the capture member (7). The capture cavity (71) has a semi-rectangular contour, that is, the capture cavity (71) is formed by splicing a rectangular cavity and a semi-circular cavity; a vertical angled baffle array (72) composed of a number of angled baffles (721) is provided in the capture cavity (71), and the openings of all the angled baffles (721) in the vertical angled baffle array (72) face the entrance end of the capture member (7); the rectangular cavity in the capture cavity (71) is in communication with the transition channel. The connection point between the transition section (6) and the capture member (7) is set on the central axis of the capture member (7); a depression is provided at the connection between the capture member (7) and the transition section (6), and the depth of the depression accounts for 1 / 5 to 1 / 4 of the longest distance between the two ends of the capture member (7).

6. The microfluidic Cryptococcus sorting device according to claim 5, characterized in that: An exit channel is provided in the exit section (8). One end of the exit channel is in communication with the semi-circular cavity in the capture cavity (71), and the other end of the exit channel communicates with the outside.

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