Hemolysis analyzer based on integrated optical waveguide sensor and preparation method thereof

Abstract

The application discloses a hemolysis analyzer based on an integrated optical waveguide sensor and a preparation method thereof, and comprises an integrated photon waveguide, a nano filter and a micro-fluidic channel, wherein the integrated photon waveguide is a single waveguide with a middle spiral type and coupling gratings on both sides. The hemolysis analyzer fully utilizes the advantages of high sensitivity and small volume of the integrated optical waveguide sensor, and realizes sample-free preparation by combining the methods of instantaneous absorption and hemoglobin filtration, and can quickly and accurately detect hemolysis and the degree of hemolysis.

Description

TECHNICAL FIELD

[0001] The application belongs to the field of sensors, and particularly relates to a hemolysis analyzer based on an integrated optical waveguide sensor and a preparation method thereof. BACKGROUND

[0002] Blood detection plays a core role in modern medical diagnosis and pathological research. The mainstream blood detection methods currently include: direct current resistance impedance method (DC), resistance impedance and radio frequency conductivity combined detection method, multi-angle laser polarized light scattering detection method, and simple cell detection method using image analysis. No matter which detection method, it is started from image processing, and the image is analyzed by pixels to obtain the result, so the detection speed is slow and the computing power is large.

[0003] Among various blood detection methods, whole blood detection is extremely important in diagnosis. However, the detection of whole blood is often disturbed by hemolysis. Hemolysis refers to a phenomenon that red blood cells are ruptured and their contents such as hemoglobin are released into the blood plasma. It is the largest pre-analysis error source of blood analyzers and laboratory blood analysis equipment in medical institutions all over the world, and the number of unqualified samples caused by it accounts for as high as 39-69%, which is about 5 times higher than the second largest reason (error caused by blood sample collection itself).

[0004] The presence of hemolysis interferes with the detection of important parameters such as K and Ca ion concentration in blood. At the same time, hemolysis is also very common in some major surgeries, which brings great trouble to the real-time monitoring of intraoperative vital signs relying on blood detection. During the process of trauma collection and blood sample preparation, traumatic blood drawing and improper sample handling are easy to produce extrinsic hemolysis. It is very important to detect the occurrence of hemolysis in time. At present, the most advanced hemolysis detection scheme needs a centrifugation step to separate blood plasma and cells and needs to collect a large amount of samples, which is time-consuming and resource-consuming, also brings pain to patients, and is also easy to cause additional analysis errors. Moreover, the presence of the centrifugation step makes the whole detection device very complex and inconvenient to use.

[0005] At the same time, in order to meet the requirements of high sensitivity, instantaneity and convenience of blood detection, the method of using light sensor detection is more and more concerned. In particular, silicon-based optical waveguide sensors are focused on because of their potential to greatly reduce costs through large-scale production.

[0006] Based on the above background, the existing hemolysis detection method has the following problems and deficiencies:

[0007] (1) The sample preparation process is complex, which may cause additional analysis errors;

[0008] (2) The detection device is large in size, not mobile, and the operation is complicated;

[0009] (3) The required sample amount is large, and the detection speed is slow. SUMMARY

[0010] In order to solve the problems in the background art, the purpose of the present application is to provide a hemolysis analyzer based on an integrated optical waveguide sensor and a preparation method thereof. The analyzer detects the hemoglobin content by detecting the degree of absorption of light after the light passes through the waveguide covered by the sample, and then determines whether hemolysis occurs.

[0011] The technical solutions adopted by the present application are as follows:

[0012] The present application discloses a hemolysis analyzer based on an integrated optical waveguide sensor, which comprises a quartz substrate, an integrated photon waveguide deposited and attached to the upper surface of the quartz substrate, a nanometer filter wrapped above the integrated photon waveguide, and a microfluidic channel placed closely on the nanometer filter. The integrated photon waveguide comprises a middle spiral waveguide and a coupling grating connected to both sides of the spiral waveguide. The sensing window opened on the microfluidic channel is aligned with the opening opened on the nanometer filter.

[0013] As a further improvement, the spiral waveguide and the coupling grating of the present application are made of the same silicon nitride material, and the spiral waveguide is a single waveguide with equal spacing and width.

[0014] As a further improvement, the nanometer filter of the present application is placed in parallel and closely with the integrated photon waveguide, and the opening part of the nanometer filter corresponds to the spiral waveguide part of the integrated photon waveguide.

[0015] As a further improvement, the nanometer filter of the present application is provided with a plurality of openings, the opening diameter is 150-450nm, and the spacing between the openings is 300-900nm.

[0016] As a further improvement, the opening of the present application is circular or square or rhombic or triangular, and a plurality of openings form a hole array.

[0017] As a further improvement, the size of the sensing window of the microfluidic channel III in contact with the opening of the nanometer filter is 1-2mm, and the surface of the microfluidic channel III is provided with an injection port and a washing liquid outlet, both with a diameter of 0.4-0.7mm.

[0018] As a further improvement, the size of the hemolysis analyzer of the present application is 2-3mm.

[0019] As a further improvement, the microfluidic channel of the present application is placed directly above the opening of the nanometer filter, and the sensing window of the microfluidic channel III is aligned with the opening or hole array on the nanometer filter.

[0020] The application further discloses a preparation method of the hemolysis analyzer based on the integrated optical waveguide sensor.

[0021] 1) cleaning a quartz substrate and depositing a silicon nitride film by using a plasma-enhanced chemical vapor deposition method;

[0022] 2) obtaining a spiral waveguide 1 pattern and a coupling grating 2 pattern structure on photoresist by using ultraviolet exposure;

[0023] 3) selectively etching the silicon nitride by using reactive-ion etching (RIE) to transfer the spiral waveguide pattern and the coupling grating pattern to the silicon nitride film;

[0024] 4) cleaning the remaining photoresist to obtain the spiral waveguide and the coupling grating, and completing the manufacture of the integrated photonic waveguide;

[0025] 5) depositing a silicon oxide upper cladding layer by using a plasma-enhanced chemical vapor deposition (PECVD) method again;

[0026] 6) obtaining an opening pattern of the nanofilter on electron resist at a corresponding position by using electron beam lithography;

[0027] 7) selectively etching the silicon oxide by using reactive-ion etching (RIE) again, cleaning the remaining residual photoresist, and obtaining the nanofilter with the completed opening on the silicon oxide upper cladding layer;

[0028] 8) manufacturing a microfluidic channel and opening a sensing window, a cleaning liquid outlet and an input hole;

[0029] 9) then, testing the hemolysis analyzer itself, that is, introducing an LED and a CCD, inputting a sample, testing the absorption of the plasma to the light field under different hemoglobin concentrations, and after the sensing hemolysis analyzer composite technical indexes are determined, performing electromechanical packaging to complete the preparation of the hemolysis analyzer.

[0030] When hemolysis occurs, the hemoglobin released by hemolysis can absorb the light transmitted in the waveguide, and the degree of hemolysis is represented by detecting how much light is absorbed. The nanofilter (II) can block the interference caused by the contact of large particles of cells with the waveguide. The microfluidic channel (III) enables the whole blood sample to be tested to contact the sensor. Finally, the rapid detection of whether hemolysis occurs and the degree of hemolysis is realized.

[0031] The nanofilter (II) is placed in parallel and close to the integrated photonic waveguide (I), and the opening part of the nanofilter (II) corresponds to the spiral waveguide (1) part of the integrated photonic waveguide (I). The application has the beneficial effect that:

[0032] (1) This invention employs a method combining transient absorption detection and hemoglobin filtration, with hemoglobin filtration achieved through an integrated nanofilter (II). This structure simplifies the sample preparation process, avoids additional errors introduced during sample preparation, and thus improves the accuracy of detection.

[0033] (2) The core of the sensor is the on-chip integrated photonic waveguide (Ⅰ). Based on on-chip photonic integration technology, it is easy to manufacture, the device is small in size, the manufacturing cost is low, and modular assembly can be realized. Compared with existing detection instruments, it solves the problems of large device size, difficult operation, and lack of scalability; at the same time, by utilizing the high sensitivity of the integrated photonic waveguide sensor, the amount of blood collected can be reduced, alleviating the patient's pain.

[0034] (3) The hemolysis analyzer in this project uses a microfluidic channel (Ⅲ), which requires only tens to hundreds of microliters of blood for testing, reducing the sampling volume, increasing the testing speed, and reducing the discomfort of the sampled person. Compared with existing testing methods, it greatly improves the testing efficiency and timeliness, providing more accurate and timely judgment for medical testing, while also improving the comfort of the treatment process.

[0035] This invention is a hemolysis analyzer based on an integrated optical waveguide sensor. It fully utilizes the high sensitivity of on-chip integrated sensors and solves the sample processing problem based on a nano-filter structure. It features simple processing and a compact structure. It can not only meet the requirements of hemolysis analysis, but also serve as a module for large-scale blood analysis equipment, thus having scalability. Attached Figure Description

[0036] Figure 1 This is a top plan view of the integrated photonic waveguide structure involved in this invention;

[0037] Figure 2 This is a three-dimensional schematic diagram of the integrated photonic waveguide structure involved in this invention;

[0038] Figure 3 This is a schematic diagram of the structure of the nanofilter involved in this invention;

[0039] Figure 4 This is a schematic diagram of the microfluidic channel involved in the present invention;

[0040] Figure 5 This is an overall schematic diagram of the hemolysis analyzer involved in the present invention;

[0041] In the figure, 1 is a helical waveguide, 2 is a coupling grating, 3 is a quartz substrate, 4 is an opening, 5 is a sensing window, 6 is an injection port, 7 is a cleaning fluid outlet, Ⅰ is an integrated photonic waveguide, Ⅱ is a nanofilter, and Ⅲ is a microfluidic channel. Methods of implementation

[0042] The technical solutions of the present application are further described below in conjunction with the drawings of the specification through specific embodiments.

[0043] The application discloses a hemolysis analyzer based on an integrated optical waveguide sensor, which comprises an integrated photon waveguide I, a nano filter II and a microfluidic channel III.

[0044] When hemolysis occurs, the hemoglobin released by hemolysis can absorb the light transmitted along the waveguide, and the degree of hemolysis can be represented by detecting how much light is absorbed. The nano filter II can block the interference caused by the contact of large particles of cells with the waveguide. The microfluidic channel III enables the whole blood sample to be tested to be in contact with the sensor. Finally, it realizes the rapid detection of whether hemolysis occurs and the degree of hemolysis.

[0045] The nano filter II is placed in parallel and close to the integrated photon waveguide I, and the opening part of the nano filter II corresponds to the spiral waveguide 1 part of the integrated photon waveguide I.

[0046] The microfluidic channel III is placed directly above the opening 4 of the nano filter II, and the sample inlet sensing window 5 of the microfluidic channel III is aligned with the opening 4 or hole array on the nano filter II.

[0047] The integrated photon waveguide I is deposited and attached to the quartz substrate 3.

[0048] Figure 5 It is the overall schematic diagram of the hemolysis analyzer involved in the present application, which comprises a quartz substrate 3, an integrated photon waveguide I deposited and attached to the upper surface of the quartz substrate 3, a nano filter II wrapped above the integrated photon waveguide I, and a microfluidic channel III placed closely on the nano filter II. The integrated photon waveguide I comprises a middle spiral waveguide 1 and a coupling grating 2 connected to both sides of the spiral waveguide 1. The sensing window 5 opened on the microfluidic channel III is aligned with the opening 4 opened on the nano filter II. The size of the hemolysis analyzer is 2-3mm.

[0049] Figure 1 It is a top view schematic diagram of the integrated photon waveguide structure of the integrated photon waveguide involved in the present application. Figure 2 It is a three-dimensional schematic diagram of the integrated photon waveguide structure involved in the present application, which comprises a spiral waveguide 1 based on an integrated photon waveguide and a coupling grating 2. The coupling grating 2 is connected to the port of the spiral waveguide 1. The optical signal enters from one side of the coupling grating 2, passes through the spiral waveguide 1, and is output from the other side of the coupling grating 2.

[0050] The integrated photon waveguide I is divided into a spiral waveguide 1 and a coupling grating 2, the spiral waveguide 1 and the coupling grating 2 are made of the same silicon nitride material, and the waveguide part is in a strip structure.

[0051] Figure 3 The figure is a structural schematic diagram of the nanometer filter II involved in the present application. The nanometer filter II is placed in parallel and close to the integrated photon waveguide I, and the opening part of the nanometer filter II corresponds to the spiral waveguide 1 part of the integrated photon waveguide I. The nanometer filter II is provided with a plurality of openings 4, the opening 4 has an aperture of 150-450 nm, the spacing between the openings 4 is 300-900 nm, the opening 4 is circular or square or rhombic or triangular, and the plurality of openings 4 form a hole array, the present application is square, and a square array 5*5=25 openings 4 are formed in the middle of the nanometer filter II; the sensing window 5 of the nanometer filter hole II in contact with the lower end of the microfluidic channel III has a size of 2 mm, and the microfluidic channel III is provided with an injection port 6 and a cleaning liquid (waste water) outlet 7 on the surface, and the apertures are all 0.5 mm.

[0052] Figure 4 The figure is a structural schematic diagram of the microfluidic channel involved in the present application. As shown in Figure 3 、 Figure 4 、 Figure 5 The integrated photon waveguide waveguide I and the nanometer filter II are both located on the quartz substrate 3. The sample is injected from the microfluidic channel injection port 6, enters the nanometer filter II after passing through the microfluidic channel sensing window 5, and the filtered material enters the slit of the spiral waveguide 1 below and fully contacts the light field in the waveguide. The microfluidic channel is placed directly above the opening of the nanometer filter, and the microfluidic channel III sample sensing window 5 is aligned with the opening or hole array on the nanometer filter.

[0053] In the present application, the wide-spectrum light enters from one side of the coupling grating 2, passes through the spiral waveguide 1, and is output from the other side of the coupling grating 2, and the degree of absorption of the specified wavelength light is detected to obtain the hemoglobin concentration, and then the hemolytic condition is judged.

[0054] The specific embodiments of the present application and the implementation conditions thereof are as follows:

[0055] First, the waveguide 1 and the coupling grating 2 are simulated and designed. The size of the silicon nitride waveguide 1 is designed by using the finite-difference method (FDM), and the degree of overlap between the light field and the sample (bovine blood is planned to be used for verification) under different waveguide sizes is obtained. As a preliminary preparation, the sample hemoglobin can be deposited on the silicon wafer as a thin film, and the refractive index and absorption coefficient are measured by an ellipsometer. The coupling grating is designed by using the finite-difference time-domain method (FDTD), and the target is to have a coupling efficiency greater than 10% at a wavelength of 420 nm and a bandwidth of not less than 50 nm.

[0056] (Need to be specially noted that in the experimental verification stage, the sample is bovine blood, so there is no ethical approval problem. When the prototype is completed and needs to be verified in the hospital laboratory, the cooperating hospital, Children's Hospital Affiliated to Zhejiang University, will conduct an ethical review and provide permission.)

[0057] Secondly, the optical sensing hemolysis analyzer is processed. The micro-nano processing laboratory provided by the author's unit, Zhejiang University Jiaxing Research Institute, will be used to process and manufacture the hemolysis analyzer. The basic process flow is as follows:

[0058] 1) Clean the quartz substrate 3 and deposit a silicon nitride film on the upper surface of the quartz substrate 3 by using plasma-enhanced chemical vapor deposition (PECVD);

[0059] 2) Use ultraviolet exposure to obtain the spiral waveguide 1 pattern and the coupling grating 2 structure pattern on the photoresist;

[0060] 3) Use reactive ion etching (RIE) to selectively etch the silicon nitride, and transfer the photoresist pattern to the silicon nitride film;

[0061] 4) Clean the remaining photoresist to obtain the spiral waveguide 1 and the coupling grating 2, and complete the manufacture of the integrated photonic waveguide I;

[0062] 5) Again, use plasma-enhanced chemical vapor deposition (PECVD) to deposit a silicon oxide upper cladding layer on the quartz substrate 3, and wrap the completed integrated photonic waveguide I inside.

[0063] 6) Use electron beam lithography to obtain the opening 4 of the nanofilter II and the pattern of its array on the corresponding position of the electron resist.

[0064] 7) Again, the silicon oxide is selectively etched by reactive-ion etching (RIE), the remaining photoresist is cleaned, and the nanofilter II with the array of openings 4 is obtained by coating the silicon oxide.

[0065] 8) The microfluidic channel III is fabricated, and the injection port 6 and the cleaning liquid outlet 7 are formed on the upper surface, and the sensing window 5 is formed on the center of the lower surface.

[0066] 9) The plasma-enhanced chemical vapor deposition, the reactive-ion etching and the ultraviolet exposure are combined in the process of the hemolysis analyzer, so as to improve the production efficiency and the product qualification rate.

[0067] Then, the hemolysis analyzer itself is tested. The LED and the CCD are introduced and placed on the two sides of the grating, respectively, for inputting and receiving the light signal. The sample is injected from the injection port 6 of the microfluidic channel, and the sample enters the surface and the side of the spiral waveguide 1 through the sensing window 5 of the microfluidic channel and the opening 4 of the nanofilter. The absorption of the plasma to the light field under different hemoglobin concentrations is tested. After the composite technical indexes of the sensing hemolysis analyzer are determined, the mechanical and electrical packaging is performed, and the prototype is completed.

[0068] Finally, the prototype is tested by using the human blood in the laboratory of the Children's Hospital Affiliated to Zhejiang University, and the final parameter calibration is completed.

[0069] The above examples are used to explain and illustrate the present application, but not to limit the present application. Any modification and change made to the present application within the spirit and the protection scope of the claims of the present application all fall into the protection scope of the present application.

Claims

1. A hemolysis analyzer based on an integrated optical waveguide sensor, characterized in that: The device includes a quartz substrate (3), an integrated photonic waveguide (Ⅰ) deposited on the upper surface of the quartz substrate (3), a nanofilter (Ⅱ) wrapped above the integrated photonic waveguide (Ⅰ), and a microfluidic channel (Ⅲ) placed close to the nanofilter (Ⅱ). The integrated photonic waveguide includes an intermediate helical waveguide (1) and a coupling grating (2) connected to both sides of the helical waveguide (1). The microfluidic channel (Ⅲ) passes through an opening in the nanofilter (Ⅱ). The spiral waveguide (1) and the coupling grating (2) are made of the same silicon nitride material, and the spiral waveguide (1) is a single waveguide with equal spacing and width; The nanofilter (II) is placed parallel to and close to the integrated photonic waveguide (I), and the opening of the nanofilter (II) corresponds to the spiral waveguide (1) part of the integrated photonic waveguide (I); The microfluidic channel (Ⅲ) is placed directly above the opening (4) of the nanofilter (Ⅱ), and the sensing window (5) of the microfluidic channel (Ⅲ) is aligned with the opening (4) or the pore array on the nanofilter (Ⅱ).

2. The hemolysis analyzer based on an integrated optical waveguide sensor according to claim 1, characterized in that: The nanofilter (II) has several openings (4), the diameter of the openings (4) is 150-450nm, and the spacing between the openings is 300-900nm.

3. The hemolysis analyzer based on an integrated optical waveguide sensor according to claim 2, characterized in that: The opening (4) is circular, square, rhomboid, or triangular, and the plurality of openings (4) form an array of holes.

4. The hemolysis analyzer based on an integrated optical waveguide sensor according to claim 3, characterized in that: The sensing window (5) of the microfluidic channel (Ⅲ) is in contact with the nanofilter (Ⅱ) and its size is 1-2 mm. The upper surface of the microfluidic channel (Ⅲ) is provided with an injection port (6) and a cleaning fluid outlet (7), both with a diameter of 0.4-0.7 mm.

5. The hemolysis analyzer based on an integrated optical waveguide sensor according to claim 1, 2, 3, or 4, characterized in that: The hemolysis analyzer has a size of 2-3 mm.

6. A method for preparing a hemolysis analyzer based on an integrated optical waveguide sensor as described in claim 1, 2, 3, or 4, characterized in that, Includes the following steps: 1) Clean the quartz substrate (3) and deposit a silicon nitride thin film using plasma-enhanced chemical vapor deposition; 2) The spiral waveguide (1) pattern and the coupling grating (2) pattern structure were obtained on the photoresist by ultraviolet exposure; 3) Reactive-ion etching (RIE) is used to selectively remove silicon nitride and transfer the spiral waveguide (1) pattern and the coupling grating (2) pattern onto the silicon nitride thin film. 4) Clean the remaining photoresist to obtain the spiral waveguide (1) and the coupling grating (2), thus completing the fabrication of the integrated photonic waveguide (Ⅰ); 5) Utilizing plasma-enhanced chemical vapor deposition (Plasma-enhanced) again... Chemical vapor deposition (PECVD) to deposit a cladding layer on silicon oxide; 6) Electron beam lithography was used to obtain the opening (4) pattern of the nanofilter (II) on the electron adhesive at the corresponding position; 7) Reactive-ion etching (RIE) is used to selectively remove silicon oxide, clean the remaining residue, and coat the silicon oxide to obtain a nanofilter (Ⅱ) with a complete opening (4); 8) Fabricate a microfluidic channel (Ⅲ) and open an injection port (6), a cleaning fluid outlet (7) and a sensing window (5); 9) Next, the hemolysis analyzer itself is tested, that is, LEDs and CCDs are introduced, samples are passed through, and the absorption of light field by plasma at different hemoglobin concentrations is tested. After the composite technical indicators of the sensor hemolysis analyzer are determined, electromechanical packaging is carried out to complete the preparation of the hemolysis analyzer.

Images