Mediterranean anemia red blood cell form rapid analyzer based on micro-fluidic chip

Abstract

The invention relates to the field of medical diagnosis, in particular to a thalassemia red blood cell morphology rapid analyzer based on a micro-fluidic chip, which comprises a micro-fluidic chip module, a shooting module, an image recognition module, a temperature control module and a display module, the temperature control module is used for controlling the temperature of the micro-fluidic chip module, and the shooting module is used for shooting the movement and deformation process of red blood cells in the micro-fluidic chip module. The image recognition module is used for carrying out image recognition on the shot image and analyzing the performance of the red blood cells from dynamic and static directions according to an image recognition result; and the display module is used for displaying a recognition result and an analysis result of the image recognition module. According to the scheme, the deformation capacity and the recovery capacity of the red blood cells are judged from the static direction and the dynamic direction, and the recognition accuracy of diseases such as thalassemia is improved.

Description

Technical Field

[0001] The present invention relates to the field of medical diagnosis, and in particular to a rapid analyzer for thalassemia red blood cell morphology based on a microfluidic chip. Background Art

[0002] In recent years, microfluidics, an emerging micro-nano manipulation platform, has shown broad application prospects in biological sample processing and analysis. This technology enables precise control, sorting, and analysis of minute amounts of liquid by constructing micrometer-scale channels within the chip, offering advantages such as low sample volume, fast reaction times, and the ability to integrate automation. In the field of red blood cell analysis, microfluidics chips can utilize fluid dynamics and microscopic geometry to arrange red blood cells, measure their deformation, and capture images, thereby assisting in automated morphological recognition.

[0003] For example, the prior art disclosed in CN108855256B discloses a microfluidic chip and method for detecting red blood cell deformability. The present invention utilizes a wavy microfluidic channel, comprising an inlet, a first buffer straight channel, a first connecting portion, a wavy channel, a second connecting portion, a second buffer straight channel, and an outlet, which are sequentially connected. While providing continuous extrusion stress, red blood cells also bend and deform with the changes in the waves, providing another form of stress. A perfusate propulsion device, using a pressure gradient method controlled by height difference, is used to inject perfusate into the microfluidic chip for detecting red blood cell deformability, thereby promoting the flow of perfusate within the microfluidic chip.

[0004] Existing technologies generally analyze red blood cells using only static indicators such as the proportion of fragmented cells, without analyzing them dynamically. In reality, conditions such as iron deficiency anemia and hereditary spherocytosis can also lead to an elevated proportion of fragmented cells, but their dynamic properties (such as recovery rate) may differ from those of thalassemia. Relying solely on static indicators can lead to misdiagnosis of the cause of the disease. The present invention was developed to address these common problems in the field. Summary of the Invention

[0005] The purpose of the present invention is to address the current deficiencies and propose a rapid analyzer for thalassemia red blood cell morphology based on a microfluidic chip.

[0006] In order to overcome the deficiencies of the prior art, the present invention adopts the following technical solutions:

[0007] A rapid analyzer for thalassemia red blood cell morphology based on a microfluidic chip, comprising a microfluidic chip module, a shooting module, an image recognition module, a temperature control module, and a display module. The microfluidic chip module is used to construct a red blood cell flow and deformation channel, the temperature control module is used to control the temperature of the microfluidic chip module, the shooting module is used to capture the movement and deformation process of red blood cells within the microfluidic chip module, the image recognition module is used to perform image recognition on the captured image and analyze the performance of the red blood cells from both dynamic and static directions based on the image recognition results, and the display module is used to display the recognition and analysis results of the image recognition module.

[0008] Furthermore, the microfluidic chip module includes an inlet chamber, a pre-channel, a multi-level microchannel and an outlet collection pool. The inlet chamber is used to introduce a red blood cell sample. The pre-channel is used to guide the red blood cell sample into the multi-level microchannel and uniformize the sample flow rate. The multi-level microchannel includes microchannels of different widths, heights, bending radii and geometric configurations. The microchannel is used to construct multiple shear force gradient zones and induce deformation of red blood cells. The outlet collection pool is used to collect the deformed red blood cell sample and prevent its backflow.

[0009] Furthermore, the shooting module includes a high-speed camera and a synchronous lighting device, wherein the high-speed camera is used to continuously shoot images of red blood cells while they pass through the microchannel, and the synchronous lighting device is used to illuminate during the shooting process to improve image contrast.

[0010] Furthermore, the image recognition module includes a preprocessing unit, a deep learning unit and an analysis unit. The preprocessing unit is used to denoise and enhance the edges of the image. The deep learning unit is used to recognize the image and obtain the morphological indicators of the red blood cells through the deep learning algorithm and the processed image. The analysis unit is used to analyze the dynamic performance of the red blood cells based on the processed image.

[0011] Furthermore, the temperature control module includes a temperature sensor, a heating unit and an adjustment unit. The temperature sensor is used to detect the temperature of the microfluidic chip module, the heating unit is used to heat the microfluidic chip module, and the adjustment unit is used to feedback adjust the heating temperature of the heating unit through a PID adjustment algorithm according to the detection temperature of the temperature sensor.

[0012] Furthermore, the workflow of the system includes the following steps:

[0013] S1, the temperature control module controls the temperature of the microfluidic chip module;

[0014] S2, red blood cells pass through the microfluidic chip module and deform during the process;

[0015] S3, the shooting module captures images of red blood cells at various moments during the process of passing through the microfluidic chip module;

[0016] S4, the image recognition module performs image recognition on the captured image and analyzes the performance of the red blood cells based on the image recognition results;

[0017] S5, the display module displays the recognition results and analysis results of the image recognition module.

[0018] The present invention achieves the following beneficial effects: 1. A camera module captures continuous images of red blood cells in a microfluidic chip, and combined with an image recognition module, extracts both the dynamic and static properties of the red blood cells. This allows for simultaneous assessment of the red blood cell's deformability and resilience from both static and dynamic perspectives, improving the accuracy of identifying diseases such as thalassemia.

[0019] 2. The system integrates a microfluidic chip, high-speed imaging, temperature control, image recognition, and display modules to automate the entire process from sample entry to analysis result output. Compared to traditional manual microscopic analysis, it offers faster analysis and more objective results, making it suitable for high-throughput testing needs. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The present invention can be further understood from the following description in conjunction with the accompanying drawings. The components in the figures are not necessarily drawn to scale, but rather the emphasis is placed on illustrating the principles of the embodiments. In different views, the same reference numerals designate the same parts.

[0021] Figure 1 It is a structural schematic diagram of the present invention.

[0022] Figure 2 Schematic diagram of the structure of the microfluidic chip module of the present invention.

[0023] Figure 3 It is the workflow diagram of the present invention.

[0024] Figure 4 Graph showing the relationship between the estimated instantaneous membrane tension, equivalent radius, and fluid shear force of the present invention. DETAILED DESCRIPTION

[0025] The following is an explanation of the embodiments of the present invention through specific embodiments. Those skilled in the art can understand the advantages and effects of the present invention from the contents disclosed in this specification. The present invention can be implemented or applied through other different specific embodiments, and the details in this specification can also be modified and changed in various ways based on different viewpoints and applications without departing from the spirit of the present invention. In addition, the drawings of the present invention are only simple schematic illustrations and are not depicted in actual size. It is stated in advance. The following embodiments will further explain the relevant technical content of the present invention in detail, but the disclosed content is not intended to limit the scope of protection of the present invention.

[0026] Example 1: According to Figure 1 、 Figure 2 、 Figure 3 and Figure 4 This embodiment provides a rapid analyzer for thalassemia red blood cell morphology based on a microfluidic chip, comprising a microfluidic chip module, a shooting module, an image recognition module, a temperature control module, and a display module. The microfluidic chip module is used to construct a red blood cell flow and deformation channel, the temperature control module is used to control the temperature of the microfluidic chip module, the shooting module is used to capture the movement and deformation process of red blood cells within the microfluidic chip module, the image recognition module is used to perform image recognition on the captured image and analyze the performance of the red blood cells from both dynamic and static directions based on the image recognition results, and the display module is used to display the recognition and analysis results of the image recognition module.

[0027] Furthermore, the microfluidic chip module includes an inlet chamber, a pre-channel, a multi-level microchannel and an outlet collection pool. The inlet chamber is used to introduce a red blood cell sample. The pre-channel is used to guide the red blood cell sample into the multi-level microchannel and uniformize the sample flow rate. The multi-level microchannel includes microchannels of different widths, heights, bending radii and geometric configurations. The microchannel is used to construct multiple shear force gradient zones and induce deformation of red blood cells. The outlet collection pool is used to collect the deformed red blood cell sample and prevent its backflow.

[0028] Specifically, the microchannel induces deformation of red blood cells by forming a velocity gradient of the fluid, that is, an "interlayer pulling force" caused by the uneven distribution of wall viscosity and flow velocity of the liquid in the channel.

[0029] Furthermore, the shooting module includes a high-speed camera and a synchronous lighting device, wherein the high-speed camera is used to continuously shoot images of red blood cells while they pass through the microchannel, and the synchronous lighting device is used to illuminate during the shooting process to improve image contrast.

[0030] Furthermore, the image recognition module includes a preprocessing unit, a deep learning unit and an analysis unit. The preprocessing unit is used to denoise and enhance the edges of the image. The deep learning unit is used to recognize the image and obtain the morphological indicators of the red blood cells through the deep learning algorithm and the processed image. The analysis unit is used to analyze the dynamic performance of the red blood cells based on the processed image.

[0031] Specifically, the morphological indicators include the size atypia of red blood cells (the larger the size, the worse the performance), the proportion of target-shaped cells (the larger the size, the worse the performance), the proportion of fragmented cells (the larger the size, the worse the performance), the proportion of teardrop cells (the larger the size, the worse the performance), the abnormal rate of the central pale staining area (the larger the size, the worse the performance) and the standard deviation of the cell area (the larger the size, the worse the performance). The model used by the deep learning algorithm is obtained by those skilled in the art through the Internet or through self-training.

[0032] Furthermore, the temperature control module includes a temperature sensor, a heating unit and an adjustment unit. The temperature sensor is used to detect the temperature of the microfluidic chip module, the heating unit is used to heat the microfluidic chip module, and the adjustment unit is used to feedback adjust the heating temperature of the heating unit through a PID adjustment algorithm according to the detection temperature of the temperature sensor.

[0033] Furthermore, the workflow of the system includes the following steps:

[0034] S1, the temperature control module controls the temperature of the microfluidic chip module;

[0035] S2, red blood cells pass through the microfluidic chip module and deform during the process;

[0036] S3, the shooting module captures images of red blood cells at various moments during the process of passing through the microfluidic chip module;

[0037] S4, the image recognition module performs image recognition on the captured image and analyzes the performance of the red blood cells based on the image recognition results;

[0038] Specifically, the dynamic performance of red blood cells can be determined according to the following formula:

[0039]

[0040] E n (t) = FT n (t)*A n (t)

[0041]

[0042] F n (t)=NIAN*RATE n

[0043] Among them, MEMORY is the stress memory parameter of the red blood cell membrane. The larger the parameter, the better the dynamic performance. NUM is the number of red blood cell samples involved in the calculation. n is the recovery ability parameter of the nth red blood cell sample, which is used to reflect whether the red blood cells can recover their original shape after being subjected to shear force. The larger the parameter, the better the ability. n E is the time required for the nth red blood cell sample to recover from the maximum deformation to no longer deform. The shorter the time, the better the dynamic recovery ability of the red blood cell. n (t) is the recovery energy consumption parameter of the nth red blood cell sample at time t, which is used to reflect the integrity and functionality of red blood cells. The smaller the parameter, the better the performance. The total energy consumption parameter of the nth red blood cell sample can be obtained to further analyze the integrity and functionality of red blood cells. ES is the reference value of the total energy consumption of red blood cells from maximum deformation to no longer deforming. This reference value can be obtained by testing normal red blood cells during the experiment and taking the average value.

[0044] e is a natural constant, V nmax is the volume of the nth red blood cell sample at the maximum cell deformation, V npost V is the volume of the nth red blood cell sample when the cell recovers from the maximum deformation to no longer deform. npre is the volume of the nth red blood cell sample before the cell begins to deform. The above volume can be obtained by recognizing the image through the deep learning unit. By setting the exponential function, it is beneficial to calculate the volume according to |V npre -V npost |Write R n The value range of is limited to between 0 and 1;

[0045] FT n (t) is the estimated instantaneous membrane tension of the nth red blood cell sample at time t, A n (t) is the surface area of ​​the nth red blood cell sample at time t, which can be obtained by deep learning unit to identify the image. n (t) is the fluid shear force on the nth red blood cell sample at time t, R n (t) is the equivalent radius of the nth red blood cell sample at time t, which can be obtained by deep learning unit to recognize the image, NIAN is the dynamic viscosity of the fluid, that is, the RATE of the red blood cell sample. n is the flow velocity gradient of the channel where the nth red blood cell sample is located, NIAN and RATE n It can be obtained in advance through experiments, F n (t)=NIAN*RATE nis a common formula for calculating fluid shear force; wherein the units of volume and area are preferably cubic micrometers and square micrometers, respectively.

[0046] Patients with blood diseases such as thalassemia have microstructural abnormalities in their red blood cell membranes, such as defects in the α / β chains of the skeleton protein. These changes will lead to a decrease in the elastic and viscous recovery ability of the cell membrane. Under the influence of shear force, these cells do not recover well after deformation, and their dynamic changes are different from those of healthy cells. Conventional morphological indicators are all static indicators that only consider the final deformation results, without considering the dynamic process of deformation. MEMORY is an indicator based on a dynamic process that can reflect the dynamic performance of red blood cells. It is conducive to analyzing the performance of red blood cells from a dynamic level, and helps staff to judge the condition of thalassemia by combining the static performance of red blood cells (morphological indicators) and dynamic performance (red blood cell membrane stress memory parameters).

[0047] like Figure 4 As shown, Figure 4 To estimate the instantaneous membrane tension (unit is 10 -12 N) and the equivalent radius (unit is 10 -6 m) and fluid shear force (unit is 10 -8 N) relationship diagram.

[0048] S5, the display module displays the recognition results and analysis results of the image recognition module.

[0049] The beneficial effects of this solution are as follows: 1. The camera module captures continuous images of red blood cells in the microfluidic chip, and combines this with the image recognition module to extract both the dynamic and static properties of the red blood cells. This allows for simultaneous assessment of the red blood cell's deformability and resilience from both static and dynamic perspectives, improving the accuracy of identifying diseases such as thalassemia.

[0050] 2. The system integrates a microfluidic chip, high-speed imaging, temperature control, image recognition, and display modules to automate the entire process from sample entry to analysis result output. Compared to traditional manual microscopic analysis, it offers faster analysis and more objective results, making it suitable for high-throughput testing needs.

[0051] Example 2: This embodiment should be understood as including all the features of any of the aforementioned embodiments, and is further improved on the basis of the aforementioned embodiments, and also includes a preferred multi-level microchannel and a control method thereof, wherein the preferred multi-level microchannel includes a plastic microchannel, and the plastic microchannel includes an outer wall and an inner wall, the inner wall is composed of PDMS material, and the PDMS material is a conventional material for microchannels, and the outer wall is composed of an azobenzene cross-linked polymer, and the azobenzene cross-linked polymer is a highly transparent material. The azobenzene group of the azobenzene cross-linked polymer can undergo cis-trans isomerization (this process is a reversible process) under irradiation with a specific wavelength (320-450nm), thereby deforming, and the azobenzene group can return to its original state under a wavelength of 450-550nm, and the outer wall and the inner wall are tightly bound by chemical bonding.

[0052] The control method comprises the following steps:

[0053] STEP 1: Initially, a light source with a specific wavelength is used to continuously illuminate the designated position of the plastic microchannel;

[0054] STEP2: The outer wall of the irradiated area deforms, which in turn causes the inner wall to deform.

[0055] Step 3: The microchannel continues to deform. When it reaches the target shape, the irradiation method is changed.

[0056] Specifically, by adjusting the initial illumination time, the microchannel can be deformed into different target shapes, which can be set specifically according to the detection requirements. Different target shapes correspond to different fluid shear forces. By deforming the microchannel into different target shapes, different fluid shear forces can be applied to red blood cells. The fluid shear forces corresponding to different target shapes are obtained by technicians in this field through experiments.

[0057] STEP4: Change the illumination mode to low-frequency intermittent pulse illumination to provide energy to the outer wall intermittently.

[0058] Inhibit its recovery, thus ensuring that the microchannel maintains the target shape;

[0059] STEP 5: After the red blood cell detection is completed, stop the illumination and restore the microchannel to its original state.

[0060] The beneficial effects of this embodiment are as follows: by using azobenzene cross-linked polymers and PDMS materials to form plastic microchannels, it is beneficial to adjust the plastic microchannels through light illumination. By adjusting the illumination-related parameters, plastic microchannels of different target shapes can be obtained, thereby providing different fluid shear forces for red blood cell detection to meet different experimental requirements. Compared with the existing technology, functions that originally required dozens of microchannels to achieve can be integrated into one microchannel, which greatly saves costs, has higher adjustability, and improves the flexibility of the system.

[0061] The above disclosure is only a preferred embodiment of the present invention and does not limit the scope of protection of the present invention. Therefore, any equivalent technical changes made by using the contents of the present invention and the drawings are included in the scope of protection of the present invention. In addition, as technology develops, the elements therein may be updated. The above units are only examples. Those skilled in the art can make different designs and adopt corresponding units according to actual needs when implementing this solution.

Claims

1. A rapid analyzer for thalassemia red blood cell morphology based on a microfluidic chip, characterized in that: It includes a microfluidic chip module, a shooting module, an image recognition module, a temperature control module and a display module. The microfluidic chip module is used to construct a red blood cell flow and deformation channel, the temperature control module is used to control the temperature of the microfluidic chip module, the shooting module is used to shoot the movement and deformation process of red blood cells in the microfluidic chip module, the image recognition module is used to perform image recognition on the captured image and analyze the performance of red blood cells from both dynamic and static directions based on the image recognition results, and the display module is used to display the recognition and analysis results of the image recognition module.

2. The rapid analyzer for thalassemia red blood cell morphology based on a microfluidic chip according to claim 1, characterized in that: The microfluidic chip module includes an inlet chamber, a pre-channel, a multi-level microchannel and an outlet collection pool. The inlet chamber is used to introduce a red blood cell sample. The pre-channel is used to guide the red blood cell sample into the multi-level microchannel and uniformize the sample flow rate. The multi-level microchannel includes microchannels of different widths, heights, bending radii and geometric configurations. The microchannel is used to construct multiple shear force gradient zones and induce red blood cell deformation. The outlet collection pool is used to collect the deformed red blood cell sample and prevent its backflow.

3. The rapid analyzer for thalassemia red blood cell morphology based on a microfluidic chip according to claim 1, characterized in that: The shooting module includes a high-speed camera and a synchronous lighting device. The high-speed camera is used to continuously shoot red blood cell images during the process of red blood cells passing through the microchannel. The synchronous lighting device is used to illuminate during the shooting process to improve image contrast.

4. The rapid thalassemia red blood cell morphology analyzer based on a microfluidic chip according to claim 1, characterized in that: The image recognition module includes a preprocessing unit, a deep learning unit and an analysis unit. The preprocessing unit is used to denoise and enhance the edges of the image. The deep learning unit is used to recognize the image and obtain the morphological indicators of the red blood cells through the deep learning algorithm and the processed image. The analysis unit is used to analyze the dynamic performance of the red blood cells based on the processed image.

5. The rapid thalassemia red blood cell morphology analyzer based on a microfluidic chip according to claim 1, characterized in that: The temperature control module includes a temperature sensor, a heating unit and an adjustment unit. The temperature sensor is used to detect the temperature of the microfluidic chip module, the heating unit is used to heat the microfluidic chip module, and the adjustment unit is used to feedback adjust the heating temperature of the heating unit through a PID adjustment algorithm according to the detection temperature of the temperature sensor.

6. The rapid analyzer for thalassemia red blood cell morphology based on a microfluidic chip according to claim 1, characterized in that: The workflow includes the following steps: S1, the temperature control module controls the temperature of the microfluidic chip module; S2, red blood cells pass through the microfluidic chip module and deform during the process; S3, the shooting module captures images of red blood cells at various moments during the process of passing through the microfluidic chip module; S4, the image recognition module performs image recognition on the captured image and analyzes the performance of the red blood cells based on the image recognition results; S5, the display module displays the recognition results and analysis results of the image recognition module.

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