CN121198368A - A flow control chip detection card - Google Patents

Abstract

This project discloses a microfluidic chip technology that enables precise manipulation and detection of trace fluids by constructing microchannels, valves, pumps, and other structures on the chip. In the field of cancer detection, microfluidic chip technology allows for efficient manipulation and detection of trace biological samples. This project's cancer biomarker detection technology, based on microfluidic chip technology, achieves high sensitivity and specificity in cancer biomarker detection by designing unique flow paths and reaction regions, optimizing detection conditions, signal amplification techniques, chip material fabrication, and antibody probe immobilization. This allows for the mixing and reaction of samples with specific biomarker detection reagents within the microfluidic chip, followed by detection of the resulting signals (such as fluorescence and electrochemical signals). This has significant application value in early cancer diagnosis, disease monitoring, and personalized treatment.

Description

Flow control chip detection card

Technical Field

The present specification relates to the field of biomedical detection, and in particular to a technology for manufacturing a cancer detection card based on a microfluidic technology. The technology combines the micro-fluidic chip with the cancer biomarker detection technology, and realizes the rapid, efficient and accurate detection of most cancers. The micro-fluidic chip technology is used as an emerging micro-nano technology, and brings new breakthrough to the field of cancer detection by virtue of the unique fluid control capability and high integration. With the continuous development of biomedical technology and the continuous increase of the demand of people for early cancer discovery, the research and development of a cancer detection card based on a microfluidic chip technology has important significance. At present, a plurality of research institutions and companies at home and abroad are dedicated to research and development in the field, and a series of important achievements are achieved.

Background

Cancer is one of the major diseases that threatens human health worldwide, and early diagnosis thereof is critical for improving survival rate and quality of life of patients. However, the conventional cancer detection methods, such as hematological examination and imaging examination, often have the disadvantages of insufficient sensitivity, low specificity, long detection period, and the like, and are difficult to meet the requirement of early cancer detection. Therefore, developing a high-efficiency, accurate and rapid cancer detection method becomes a problem to be solved urgently in the biomedical field. The micro-fluidic chip technology is used as an emerging biomedical detection platform, and has a broad application prospect in the field of cancer detection by virtue of the unique advantages. The micro-fluidic chip technology realizes accurate control and detection of micro biological samples through a micro-scale channel network and a control element. The technology has the advantages of high sensitivity, high specificity, high flux, rapidness, high efficiency and the like, can finish the detection of various cancer biomarkers in a very short time, and provides powerful support for early detection of cancers. However, current cancer detection cards based on microfluidic chip technology still have some challenges and problems. First, the design and fabrication of microfluidic chips requires a high degree of accuracy and complexity, and requires high requirements for fabrication materials and processing techniques. Second, cancer biomarkers are of a wide variety and different cancers require different biomarkers to be detected, and thus biomarker detection chips for different cancers need to be designed. In addition, the detection sensitivity and specificity of the microfluidic chip are further improved to meet the requirements of clinical detection.

1. Summary of microfluidic chip technology

Microfluidic chip technology is a technology that handles and processes fluids on a micrometer scale. By constructing micro-channel, valve, pump and other structures on the chip, accurate control and detection of micro-fluid are realized. The micro-fluidic chip technology combines the knowledge of multiple subjects such as microelectronics, chemistry, biology and the like, has high integration and customization, and brings new breakthrough to the biomedical detection field. In the field of cancer detection, the microfluidic chip technology can realize efficient control and detection of trace biological samples. By designing unique flow paths and reaction areas, a sample is mixed with a specific biomarker detection reagent and reacts, and then signals (such as fluorescence, electrochemical signals and the like) generated by the reaction are detected, so that high-sensitivity and high-specificity detection of the cancer biomarker is realized. In addition, the microfluidic chip technology can also realize multi-parameter detection, and simultaneously detect various cancer biomarkers, thereby improving the comprehensiveness and accuracy of detection.

2. Cancer biomarker detection technology

Cancer biomarkers refer to specific substances produced during the development and progression of cancer, including proteins, nucleic acids, carbohydrates, and the like. The biomarkers have abnormal contents in biological samples such as blood, urine, tissues and the like of cancer patients and can be used as important indexes for early diagnosis of cancer and monitoring of disease states. The cancer biomarker detection technology based on the micro-fluidic chip technology realizes high-sensitivity and high-specificity detection of the cancer biomarker by fixing a specific antibody or probe on a chip, specifically combining with the cancer biomarker in a sample and then detecting a signal generated after combination. In addition, the sensitivity and the specificity of the detection can be further improved by optimizing the detection conditions and the signal amplification technology.

Disclosure of Invention

There are still some challenges and problems with current cancer detection cards based on microfluidic chip technology. First, the design and fabrication of microfluidic chips requires a high degree of accuracy and complexity, and requires high requirements for fabrication materials and processing techniques. Second, cancer biomarkers are of a wide variety and different cancers require different biomarkers to be detected, and thus biomarker detection chips for different cancers need to be designed. The invention provides a cancer detection card based on a microfluidic chip technology and a manufacturing technology thereof, aiming at realizing rapid screening and diagnosis of most cancers. The detection card comprises a chip substrate, a micro-channel, a valve, a mixer, an antibody or a probe fixed on the chip and other structures. By designing a unique flow path and adopting a high-sensitivity signal detection technology, the technology can realize high-sensitivity and high-specificity detection of the cancer biomarker.

The preparation method of the invention comprises the following steps:

1. Micro-fluidic chip technology, namely, micro-channel, valve, mixer and other structures are manufactured on a chip substrate through micro-processing technology, so that accurate control and detection of micro-fluid are realized.

2. The cancer biomarker detection technology is that specific antibody or probe on a microfluidic chip is utilized to specifically combine with the cancer biomarker, and high-sensitivity and high-specificity detection of the cancer marker is realized through signal amplification and visualization technology.

1. Chip substrate material

The chip substrate is the main body part of the microfluidic chip and carries the structures of the micro-channels, the valves, the mixers and the like. In order to ensure the stability and reliability of the test card, the chip substrate material needs to have good mechanical properties, chemical stability and optical transparency. In the invention, the chip substrate is made of Polydimethylsiloxane (PDMS) material. PDMS has good elasticity, biocompatibility and optical transparency, and is easy to process into various complex microstructures. In addition, the surface of PDMS is easily chemically modified, facilitating immobilization of biomolecules such as antibodies, etc.

2. Microchannel material

The micro-channel is a key component in the microfluidic chip and is used for conveying samples and reagents to be tested. To ensure stable and smooth transport of fluids in the microchannels, the microchannel materials need to have low adsorptivity, good wettability and chemical stability. In the invention, the micro-channel material is also PDMS. The low surface energy and hydrophobicity of PDMS makes the fluid easy to transport in the microchannel and less prone to adsorption and contamination. In addition, the chemical stability of PDMS ensures that the microchannels are not corroded or deformed during long-term use.

3. Valve material

The valve is used for controlling the flow direction and speed of the fluid in the micro-channel, and is an indispensable component in the micro-fluidic chip. In order to ensure the stability and reliability of the valve, the valve material needs to have good mechanical properties, chemical stability and biocompatibility. In the present invention, the valve is made of an elastic material such as rubber or silica gel. These materials have good elasticity and sealing properties, and can ensure that the valve is effectively prevented from fluid leakage when closed. At the same time, they also have good chemical stability and biocompatibility, and do not pollute or interfere with samples and reagents.

4. Mixer material

The mixer is used for uniformly mixing the sample to be detected and the reagent in the micro-channel so as to improve the detection sensitivity. In order to ensure the mixing effect and stability of the mixer, the mixer material needs to have good wettability and chemical stability. In the invention, the mixer is made of PDMS material. The wettability of PDMS makes the sample and reagents easy to mix uniformly in the mixer and is not prone to bubbles or precipitation. Meanwhile, the chemical stability of the PDMS ensures that the mixer cannot be corroded or deformed in the long-term use process.

5. Immobilized probe material

Immobilized probes are used to specifically bind to cancer markers and generate a detectable signal. In order to ensure the stability and specific recognition capability of the probe, the probe material needs to have good biocompatibility, chemical stability and specific recognition capability. In the invention, the immobilized probe is prepared from a silanized glass surface modified by a polymer material with good biocompatibility, such as polyethylene glycol (PEG). The PEG modification can reduce the nonspecific adsorption of the probe surface and improve the specific recognition capability of the probe. Meanwhile, the silanized glass surface has good chemical stability and biocompatibility, and can ensure that the probe keeps stable performance in a long-term use process.

The specific steps for manufacturing the microfluidic chip detection card are as follows:

(1) The shape and size of the chip substrate are designed, and the layout of the microfluidic channels and detection regions is designed. Depending on the type and number of cancer markers, a plurality of detection regions are designed, each detection region performing specific detection for one or more cancer markers.

(2) A suitable transparent material is selected to make a chip substrate, such as Polydimethylsiloxane (PDMS), glass, etc. Micro-fluidic channels and detection areas are etched in the chip matrix by adopting micro-processing technologies such as photoetching and etching.

(3) And coating a layer of antibody or probe which specifically binds to the cancer marker on the detection area. The selection of antibodies or probes should be made according to the kind and nature of cancer markers. The antibody or probe is immobilized on the surface of the detection region by chemical modification or biotin-avidin method.

(4) The signal output module is communicated with the detection area. The signal output module can adopt electrochemical, optical and other detection methods to convert the detection signals into readable results. For example, when an antibody or probe binds to a cancer marker, an electrochemical or fluorescent signal may be generated, read and analyzed by a signal output module.

(5) And packaging and testing the microfluidic chip detection card. The encapsulation process should ensure the tightness and stability of the chip matrix, preventing sample leakage and contamination. The testing process should include performance evaluation and quality control to ensure accuracy and reliability of the test card.

The following is a specific description of the manufacturing method of the present invention:

chip design and manufacture

1. The chip substrate is designed to be rectangular in shape and 5cm multiplied by 3cm in size. A plurality of microfluidic channels and detection regions are designed inside the chip matrix, each detection region performing specific detection for one or more cancer markers. For example, detection regions for common cancers such as lung cancer, breast cancer, liver cancer, and the like can be designed.

2. PDMS is selected as the material for manufacturing the chip matrix. And etching a microfluidic channel and a detection area in the PDMS matrix by photoetching and etching technology. The etching depth and precision should be controlled during the etching process to ensure that the shapes and sizes of the microfluidic channels and the detection areas meet the requirements.

3. And coating a layer of antibody or probe which specifically binds to the cancer marker on the detection area. For example, the detection region for lung cancer may be coated with an antibody against a lung cancer marker such as EGFR or KRAS, and the detection region for breast cancer may be coated with an antibody against a breast cancer marker such as HER2 or CA 15-3. The antibody is immobilized on the surface of the detection area by chemical modification or biotin-avidin method.

4. And communicating the electrochemical signal output module with the detection area. The electrochemical signal output module comprises an electrode, an electrochemical workstation and the like. When the antibody binds to a cancer marker, an electrochemical signal is generated, which is read and analyzed by an electrochemical workstation.

(II) chip Package and test

1. And (3) packaging the chip by adopting a material with good biocompatibility and high chemical stability so as to protect the chip and provide a stable working environment.

2. And (3) chip testing, namely comprehensively testing and verifying the packaged chip, wherein the chip comprises performance indexes such as stability, reliability, sensitivity, specificity and the like. In the test process, an actual detection environment needs to be simulated so as to ensure that the chip has good performance in actual application.

(III) method of use

1. Sample processing, which is to collect a sample to be detected (such as blood, urine, etc.), and perform necessary pretreatment such as centrifugation, filtration, etc.

2. And (3) sample loading and detection, namely injecting the pretreated sample into a microfluidic chip detection card, starting control elements such as a valve and a pump on the chip, so that the sample flows in a micro-channel and is specifically combined with an antibody or a probe fixed on the chip. By detecting signals (such as fluorescence, electrochemical signals and the like) generated on the chip, the high-sensitivity and high-specificity detection of the cancer biomarker is realized.

3. And (3) analyzing the result by using matched detection equipment or software to judge whether the cancer biomarker, the concentration of the marker and other information exist in the sample. Based on the detection results, personalized diagnostic and therapeutic advice may be provided to the patient.

Advantageous effects

The cancer detection card based on the micro-fluidic chip technology and the manufacturing technology thereof realize high-sensitivity and high-specificity detection of the cancer biomarker by designing a unique flow path and adopting a high-sensitivity signal detection technology. The technology has the advantages of rapidness, high efficiency, multi-parameter detection and the like, and has important application value in the aspects of early diagnosis of cancers, disease monitoring, personalized treatment and the like. With the continued development of biomedical technology, it is believed that this technology will be more widely applied and generalized in the future.

The microfluidic chip detection card has the following advantages:

1. The method is efficient and rapid, realizes efficient control and accurate detection of the micro sample by a microfluidic technology, greatly accelerates the detection speed and meets the requirement of early diagnosis.

2. The high sensitivity and high specificity are that an antibody or a probe with strong specificity is adopted to combine with a cancer biomarker, and the signal generated after combination can be processed with high efficiency through a signal output system, thereby realizing the detection with high sensitivity and high specificity.

3. And the multiparameter detection comprises that a plurality of biomarker detection areas are arranged on the microfluidic chip, so that a plurality of cancer biomarkers can be detected simultaneously, and the comprehensiveness and accuracy of detection are improved.

4. The automatic sample introduction, detection and analysis are realized by a microfluidic technology and an automatic control system, so that manual operation is reduced, and the detection efficiency is improved.

Drawings

FIG. 1 shows the overall structure of a microfluidic chip detection card

Fig. 1 shows the overall structure of a microfluidic chip detection card, comprising a chip substrate, a microfluidic channel network, a biomarker detection region and a signal output system. In the figure, it can be seen that the microfluidic channel network forms a complex network structure on the chip substrate, the biomarker detection region is located at a specific position of the channel network, and the signal output system is connected with the detection region.

(II) FIG. 2A partial enlarged view of microfluidic channel network

Fig. 2 shows an enlarged view of a portion of a microfluidic channel network, with the structure and details of the microchannels clearly visible. The channels have the characteristics of high precision and high complexity, and can realize high-efficiency control and accurate detection of micro samples.

(III) FIG. 3 schematic representation of biomarker detection region

FIG. 3 shows a schematic representation of a biomarker detection region, including a specific region to which an antibody or probe is immobilized and an output for a detection signal. During the detection process, the biomarker in the sample specifically binds to the antibody or probe, thereby producing a detectable signal.

(IV) FIG. 4 is a schematic diagram of a signal output system

Fig. 4 shows a schematic diagram of a signal output system that employs fluorescence detection techniques to achieve rapid and efficient processing of the detected signals. In the detection process, whether the cancer biomarker exists in the sample is judged by measuring the intensity or change of the fluorescent signal.

Detailed Description

The invention is further illustrated by the following examples, which are not intended to limit the invention in any way, and any modifications or alterations to the invention, which would be readily apparent to a person of ordinary skill in the art, without departing from the technical solutions of the invention, are intended to fall within the scope of the claims of the invention.

Example 1

Taking bladder cancer as an example

1. The chip substrate is designed to be rectangular in shape and 5cm multiplied by 3cm in size. A plurality of microfluidic channels and detection regions are designed inside the chip matrix, each detection region performing specific detection for one or more cancer markers. Bladder cancer detection area.

2. PDMS is selected as the material for manufacturing the chip matrix. And etching a microfluidic channel and a detection area in the PDMS matrix by photoetching and etching technology. The etching depth and precision should be controlled during the etching process to ensure that the shapes and sizes of the microfluidic channels and the detection areas meet the requirements.

3. And coating a layer of antibody or probe which specifically binds to the cancer marker on the detection area. The detection area for bladder cancer can be coated with antibodies against the nuclear matrix protein 22 and the bladder tumor antigen related antigen, and the antibodies are immobilized on the surface of the detection area by a colloidal gold coated biotin-avidin method.

4. And communicating the electrochemical signal output module with the detection area. The electrochemical signal output module comprises an electrode, an electrochemical workstation and the like. When the antibody binds to a cancer marker, an electrochemical signal is generated, and is read and analyzed by an electrochemical workstation

The specific operation steps are as follows:

1 preparing to take out the detection tube and the sample to be detected, balancing to room temperature, unpacking the aluminum foil bag of the detection tube, and mixing the solid and liquid components uniformly

2 Dripping the substance to be detected into the micro-fluidic chip kit and putting the substance into a detection machine

3 Wait for 2H observations

Judging a detection result:

1. Positive results the instrument showed antibody positivity

2. Negative results the instrument showed that the antibody was negative

Example 2

Collecting urine of 100 patients diagnosed with bladder cancer and 100 persons not suffering from bladder cancer, detecting the yin-yang property of the nuclear matrix protein 22 of the bladder cancer patients and the related antigen of the bladder tumor antigen according to the method of the steps (1) - (3), and detecting the content of the nuclear matrix protein 22 in the urine of 100 persons not suffering from bladder cancer according to the statistical analysis of the steps, wherein the content of the nuclear matrix protein 22 in the urine of 100 bladder cancer patients can be detected while the related antigen of the bladder tumor antigen can be positive, and the content of the nuclear matrix protein 22 in the urine of 100 persons not suffering from bladder cancer can not be detected while the related antigen of the bladder tumor antigen can be negative.

Claims (1)

1. The manufacturing method of the microfluidic chip detection card comprises the following steps:

(1) The shape and size of the chip substrate are designed, and the layout of the microfluidic channels and detection regions is designed. Depending on the type and number of cancer markers, a plurality of detection regions are designed, each detection region performing specific detection for one or more cancer markers.

(2) A suitable transparent material is selected to make a chip substrate, such as Polydimethylsiloxane (PDMS), glass, etc. Micro-fluidic channels and detection areas are etched in the chip matrix by adopting micro-processing technologies such as photoetching and etching.

(3) And coating a layer of antibody or probe which specifically binds to the cancer marker on the detection area. The selection of antibodies or probes should be made according to the kind and nature of cancer markers. The antibody or probe is immobilized on the surface of the detection region by chemical modification or biotin-avidin method.

(4) The signal output module is communicated with the detection area. The signal output module can adopt electrochemical, optical and other detection methods to convert the detection signals into readable results. For example, when an antibody or probe binds to a cancer marker, an electrochemical or fluorescent signal may be generated, read and analyzed by a signal output module.

(5) And packaging and testing the microfluidic chip detection card. The encapsulation process should ensure the tightness and stability of the chip matrix, preventing sample leakage and contamination. The testing process should include performance evaluation and quality control to ensure accuracy and reliability of the test card.

Cancer biomarker detection technology

(1) The method is characterized in that the specific antibody or probe on the microfluidic chip is used for carrying out specific binding with the cancer biomarker according to the claim 1, and the high-sensitivity and high-specificity detection of the cancer marker is realized through a signal amplification and visualization technology.

(2) The method is realized by chip substrate materials, micro-channel materials, valve materials, mixer materials and fixed probe materials.