Application of monoclonal antibodies specific to tissue polypeptide antigens in the preparation of diagnostic reagents for assisting screening of early lung tumors

Abstract

The present invention discloses the use of monoclonal antibodies against tissue polypeptide antigens in the preparation of diagnostic reagents for assisting in the screening of early-stage lung tumors. The invention relates to a diagnostic reagent application, comprising the following steps: S1. Obtaining bronchoalveolar lavage fluid from a patient; S2. Filtering the bronchoalveolar lavage fluid using a double layer of sterile gauze or pre-treating the bronchoalveolar lavage fluid with dithiothreitol to remove mucus from the bronchoalveolar lavage fluid; S3. Centrifuging the mucus-removed bronchoalveolar lavage fluid using a centrifuge to collect the centrifuged bronchoalveolar lavage fluid; S4. Retrieving the bronchoalveolar lavage fluid from a bronchoalveolar lavage fluid collection tank, adding monoclonal antibodies against tissue polypeptide antigens, and measuring the concentration of the tissue polypeptide antigen in the bronchoalveolar lavage fluid using an enzyme-linked immunosorbent assay (ELISA). The bronchoalveolar lavage fluid collection tank used in the present invention is reusable, and the accuracy of screening for early-stage lung tumors is not reduced due to a poor sterile environment caused by repeated use of the bronchoalveolar lavage fluid collection tank.

Description

Technical Field

[0001] The present invention relates to an application of a diagnostic reagent, and in particular to an application of a tissue polypeptide-specific antigen monoclonal antibody in the preparation of a diagnostic reagent for assisting in the screening of early-stage lung tumors. Background Art

[0002] The large number of smokers and workers who work in poor air environments for a long time are mostly suspected of lung cancer. However, the diagnosis of lung cancer is currently difficult. If patients do not receive timely diagnosis and treatment, they will suffer from serious consequences such as continuous deterioration and even life-threatening. Currently, various methods for early lung cancer screening have considerable limitations. In recent years, medical thoracoscopy has been increasingly carried out. Although its diagnostic efficiency is relatively high, many grassroots hospitals do not currently have this technology. Compared with conventional examinations, the examination is expensive, somewhat invasive, and not suitable for patients with weaker physical constitutions, which limits its promotion and application. Therefore, screening for early lung cancer has always been a difficult problem that the medical community has been committed to studying.

[0003] Bronchoalveolar lavage refers to a method in which a bronchoscope is wedged into a bronchial branch, sterile saline is injected through the bronchoscope to clean the alveoli, and as much fluid as possible (i.e., alveolar lavage fluid) is sucked back and absorbed through the working channel of the bronchoscope. The alveolar lavage fluid obtained by this method can be used for subsequent examinations of inflammation, immune cells, and soluble substances. Since this method uses a fiber bronchoscope to infuse and extract saline, the saline can penetrate deep into the patient's lungs. The alveolar lavage fluid is formed by directly cleaning the patient's alveoli, and contains alveolar secretions. It best reflects the pathological characteristics of the patient's lungs and is far more accurate than the pleural effusion detection method outside the lungs. Therefore, as an auxiliary basis for the judgment of early lung tumors, the use of this technology improves the accuracy of screening for early lung tumors. In addition, this method of screening for early lung tumors does not require any surgery, does not cause trauma, is safer and more reliable, and is less expensive. Furthermore, allowing the saline solution to remain in the patient's lungs for 3 to 5 minutes allows for sufficient contact between the saline solution and the patient's alveoli, ensuring that the resulting BALF contains sufficient alveolar secretions, thereby increasing the accuracy of early lung cancer screening. However, existing BALF collection canisters are typically disposable. Because these canisters are easily exposed to air and lack a sterile environment, they can easily deteriorate, compromising the accuracy of early lung cancer screening. Summary of the Invention

[0004] In order to overcome the shortcomings of the existing technology, the purpose of the present invention is to provide the use of tissue polypeptide antigen-specific monoclonal antibodies in the preparation of diagnostic reagents for auxiliary screening of early lung tumors, which can serve as an auxiliary basis for the judgment of early lung tumors, is more accurate for screening of early lung tumors, is non-invasive, and has low cost. During the application process, the accuracy of screening for early lung tumors is not reduced due to the poor sterile environment caused by the reuse of the alveolar lavage fluid collection tank used.

[0005] The purpose of the present invention is achieved by adopting the following technical solutions:

[0006] The use of tissue polypeptide-specific antigen monoclonal antibodies in the preparation of diagnostic reagents for assisting in the screening of early-stage lung tumors comprises the following steps:

[0007] S1. Obtain bronchoalveolar lavage fluid from the patient;

[0008] S2. Using double-layer sterile gauze or filtering the alveolar lavage fluid or adding dithiothreitol to pretreat the alveolar lavage fluid to remove mucus in the alveolar lavage fluid;

[0009] S3. Centrifuge the mucus-removed alveolar lavage fluid using a centrifuge to further remove impurities, collect the centrifuged alveolar lavage fluid, and then store it in a pulmonary lavage fluid collection tank. The pulmonary lavage fluid collection tank includes a cylinder and a lid covering the mouth of the cylinder. The outer layer of the cylinder is a vacuum layer, the inner layer of the cylinder is a liquid nitrogen perfusion layer, and the bottom of the lid is an embedded portion. The embedded portion extends into the interior of the cylinder, and the bottom of the embedded portion is provided with an opening. The opening extends toward the lid to form an activated carbon holding chamber. The pulmonary lavage fluid collection tank also includes a push-pull component, which includes a push-pull rod. The push-pull rod passes through the lid from above, and the bottom end of the push-pull rod is located in the activated carbon holding chamber. The bottom end of the push-pull rod is connected to a tray containing activated carbon. The tray moves downward when the push-pull rod is pushed downward, blocking the opening at the bottom of the embedded portion and closing it. An ultraviolet lamp is provided at the bottom of the interior of the cylinder, and a transparent partition that can transmit ultraviolet light is provided above the ultraviolet lamp.

[0010] S4. Take the alveolar lavage fluid from the alveolar lavage fluid collection tank, add tissue polypeptide-specific antigen monoclonal antibody and tissue polypeptide-specific antigen secondary antibody, and use enzyme-linked immunosorbent assay to measure the concentration of tissue polypeptide-specific antigen in the alveolar lavage fluid, marked as P.

[0011] The enzyme-linked immunosorbent assay (ELISA) used in step S4 is a solid-phase immunoassay method developed from immunoassay technology to detect trace substances in body fluids. It is a special reagent analysis method and a new type of immunoassay technology developed based on immunoenzyme technology. Its basic principles are: ① The antigen or antibody is bound to the surface of a solid phase carrier and its immunoactivity is maintained; ② The antigen or antibody is linked to an enzyme to form an enzyme-labeled antigen or enzyme-labeled antibody. This enzyme-labeled antigen or enzyme-labeled antibody retains both its immunoactivity and enzyme activity. During the assay, the test specimen (the antibody or antigen to be assayed) and the enzyme-labeled antigen or enzyme-labeled antibody are sequentially immunoreacted with the antigen or antibody on the surface of the solid phase carrier. The enzyme-labeled antigen-antibody complex formed on the solid phase carrier is separated from other substances by washing. Finally, the amount of enzyme bound to the solid phase carrier is proportional to the amount of the test substance in the test specimen. After the substrate for the enzyme reaction is added, the substrate is catalyzed by the enzyme to form a colored product. The amount of the product is directly related to the amount of the test substance in the specimen, so qualitative or quantitative analysis can be performed based on the depth of the color reaction. Since the catalytic frequency of the enzyme is very high, the reaction effect can be greatly amplified, thereby making the determination method achieve a very high sensitivity.

[0012] Furthermore, the alveolar lavage fluid from which mucus was removed in step S3 was centrifuged at a rotation speed of 1500 r / min for 10 minutes.

[0013] Furthermore, the transparent partition capable of transmitting ultraviolet rays is one of optical quartz glass, ultraviolet-transmitting black glass, soda-lime-silica short-wave ultraviolet-transmitting glass and soda-lime-ultraviolet-transmitting glass.

[0014] Furthermore, in S2, the mass concentration of dithiothreitol is 0.1%.

[0015] Furthermore, the push-pull component is provided with an external thread at a position where it passes through the cover, and the cover is provided with an internal thread matching the external thread.

[0016] Furthermore, the material of the cylinder is stainless steel or aluminum alloy.

[0017] Furthermore, the top end of the push-pull rod is connected to a handle.

[0018] Furthermore, the cover is provided with a liquid nitrogen vent hole, and the liquid nitrogen vent hole is connected to the liquid nitrogen perfusion layer.

[0019] Compared with the prior art, the present invention has the following beneficial effects:

[0020] The present invention provides an application of a tissue polypeptide-specific antigen monoclonal antibody in the preparation of a diagnostic reagent for assisting in the screening of early-stage lung tumors. Tissue polypeptide-specific antigen (TPS) is a biomarker for early-stage lung tumors. The tissue polypeptide-specific antigen monoclonal antibody used in this method can specifically detect tissue polypeptide-specific antigens in alveolar lavage fluid, and as an auxiliary basis for the judgment of early-stage lung tumors, the screening of early-stage lung tumors is more accurate. In addition, the bronchoalveolar lavage technique of the present invention does not require any surgery to extract alveolar lavage fluid, does not cause trauma, is safer and more reliable, and has lower costs. The fresh alveolar lavage fluid extracted by the present invention is placed in a pulmonary alveolar lavage fluid collection tank. Before use, the pulmonary alveolar lavage fluid collection tank is opened. The ultraviolet lamp at the bottom of the cylinder emits ultraviolet rays that pass through the transparent partition above that can transmit ultraviolet rays to irradiate and sterilize the inside of the cylinder. Then, the push-pull rod outside the lid is pulled upward to separate the tray at the bottom of the push-pull rod and the opening at the bottom of the embedded part. The ozone generated during the ultraviolet sterilization process enters the activated carbon containing chamber from the opening at the bottom of the embedded part. The activated carbon in the tray absorbs the ozone that enters the activated carbon containing chamber and eliminates the ozone inside the cylinder. After the elimination is completed, the alveolar lavage fluid collection tank is used to repeatedly collect new fresh alveolar lavage fluid. The inner layer of the alveolar lavage fluid collection tank is filled with liquid nitrogen. The temperature of the liquid nitrogen is very low, which keeps the inside of the cylinder at a low temperature and allows the alveolar lavage fluid collected in the alveolar lavage fluid collection tank to be preserved for a long time. BRIEF DESCRIPTION OF THE DRAWINGS

[0021] In order to more clearly illustrate the embodiments of the present invention, the following briefly introduces the drawings required for the embodiments. Obviously, the drawings described below are merely embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on the provided drawings without any creative work.

[0022] Figure 1 Schematic diagram of the structure of the alveolar lavage fluid collection tank in Example 1.

[0023] In the figure: 1. Cylinder; 11. Vacuum layer; 12. Liquid nitrogen perfusion layer; 2. Lid; 21. Embedded part; 211. Opening; 3. Activated carbon holding chamber; 4. Push-pull component; 41. Push-pull rod; 42. Tray; 43. Handle; 5. Activated carbon; 6. External thread; 7. Liquid nitrogen vent hole; 10. Ultraviolet lamp; 100. Transparent partition for transmitting ultraviolet rays. DETAILED DESCRIPTION

[0024] The present invention will be further described below in conjunction with specific embodiments. It should be noted that, under the premise of no conflict, the embodiments or technical features described below can be arbitrarily combined to form new embodiments.

[0025] Example 1

[0026] This embodiment provides the use of a tissue polypeptide-specific antigen monoclonal antibody in the preparation of a diagnostic reagent for assisting in the screening of early-stage lung tumors, which is characterized by comprising the following steps:

[0027] S1. Obtain bronchoalveolar lavage fluid from the patient;

[0028] S2. Filter the BALF using double-layer sterile gauze or pretreat the BALF with 0.1% dithiothreitol to remove mucus from the BALF.

[0029] S3. Centrifuge the alveolar lavage fluid from which mucus has been removed using a centrifuge at a speed of 1500 r / min for 10 minutes. Centrifuge the alveolar lavage fluid from which mucus has been removed in step S3 for 10 minutes to further remove impurities. Collect the centrifuged alveolar lavage fluid and store it in a pulmonary lavage fluid collection tank. The pulmonary lavage fluid collection tank includes a cylinder 1 and a lid 2 that covers the mouth of the cylinder 1. The outer layer of the cylinder 1 is a vacuum layer 11, and the inner layer of the cylinder 1 is a liquid nitrogen perfusion layer 12. An embedded portion 21 is provided at the bottom of the lid 2. The embedded portion 21 extends into the interior of the cylinder 1, and an opening 211 is provided at the bottom of the embedded portion 21. , the opening 211 extends toward the cover 2 to form an activated carbon containing chamber 3; the alveolar lavage fluid collection tank further includes a push-pull component 4, which includes a push-pull rod 41. The push-pull rod 41 passes through the cover 2 from above, and the bottom end of the push-pull rod 41 is located in the activated carbon containing chamber 3. The bottom end of the push-pull rod 41 is connected to a tray 42, in which activated carbon 5 is placed. The tray 42 moves downward when the push-pull rod 41 is pushed downward, blocking the opening 211 at the bottom of the embedded portion 21; an ultraviolet lamp 10 is provided at the bottom of the cylinder 1, and a transparent partition 100 that can transmit ultraviolet light is provided on the inner wall of the cylinder 1 above the ultraviolet lamp 10;

[0030] S4. Take the alveolar lavage fluid from the alveolar lavage fluid collection tank, add tissue polypeptide-specific antigen monoclonal antibody and tissue polypeptide-specific antigen secondary antibody, and use enzyme-linked immunosorbent assay to measure the concentration of tissue polypeptide-specific antigen in the alveolar lavage fluid, marked as P.

[0031] like Figure 1As shown, the alveolar lavage fluid collection tank used in step S3 includes a cylinder 1 and a lid 2 covering the bottle mouth of the cylinder 1. The outer layer of the cylinder 1 is a vacuum layer 11, which is used to isolate the low-temperature environment of the inner layer of the cylinder 1 from the external environment temperature. The inner layer of the cylinder 1 is a liquid nitrogen perfusion layer 12, which is perfused with liquid nitrogen. The temperature of the liquid nitrogen is very low, so that the interior of the cylinder 1 maintains a low-temperature environment, so that the alveolar lavage fluid collected in the alveolar lavage fluid collection tank can be stored for a long time. The bottom of the lid 2 is an embedded portion 21, which extends into the interior of the cylinder 1. The bottom of the embedded portion 21 is provided with an opening 211, and the opening 211 extends toward the lid 2 to form an activated carbon receiving chamber 3; the alveolar lavage fluid collection tank also includes The push-pull component 4 includes a push-pull rod 41, which passes through the cover 2 from the top of the cover 2. The bottom end of the push-pull rod 41 is located in the activated carbon holding chamber 3. The bottom end of the push-pull rod 41 is connected to a tray 42, and activated carbon 5 is placed in the tray 42. The tray 42 moves downward in the process of the push-pull rod 41 pushing downward, blocking the opening 211 at the bottom of the embedded part 21 and closing it; an ultraviolet lamp 10 is provided at the bottom of the interior of the cylinder 1, and a transparent partition 100 that can transmit ultraviolet rays is provided above the ultraviolet lamp 10. The transparent partition 100 that can transmit ultraviolet rays can be one of optical quartz glass, ultraviolet-transmitting black glass, soda-lime-silica short-wave ultraviolet-transmitting glass, and soda-lime-transparent ultraviolet glass.

[0032] Preferably, the push-pull component 4 is provided with an external thread 6 at the position where it passes through the cover 2, and the cover 2 is provided with an internal thread matching the external thread 6, so that the push-pull component 4 can move up and down by rotating.

[0033] Preferably, the material of the cylinder 1 is stainless steel or aluminum alloy.

[0034] Preferably, a handle 43 is connected to the top end of the push-pull rod 41 .

[0035] The alveolar lavage fluid collection tank of the present invention contains fresh alveolar lavage fluid obtained from a patient. Before use, the ultraviolet lamp 10 at the bottom of the cylinder 1 is turned on. The ultraviolet rays emitted by the ultraviolet lamp 10 pass through the transparent partition 100 above that is transparent to ultraviolet rays to irradiate and sterilize the interior of the cylinder 1. Then, the push-pull rod 41 on the outside of the lid 2 is pulled upward to separate the tray 42 at the bottom end of the push-pull rod 41 from the opening 211 at the bottom of the embedded portion 21. Ozone generated during the ultraviolet sterilization process enters the activated carbon containing chamber 3 through the opening 211 at the bottom of the embedded portion 21. The activated carbon 5 in the tray 42 absorbs the ozone that enters the activated carbon containing chamber 3, thereby eliminating the ozone inside the cylinder 1. After elimination is completed, the alveolar lavage fluid collection tank is used to repeatedly collect new fresh alveolar lavage fluid. The inner layer of the alveolar lavage fluid collection tank is filled with liquid nitrogen. The temperature of the liquid nitrogen is very low, which maintains a low temperature environment inside the cylinder 1, allowing the alveolar lavage fluid to be stored for a long time.

[0036] The above embodiments are only preferred embodiments of the present invention and cannot be used to limit the scope of protection of the present invention. Any non-substantial changes and replacements made by technicians in this field on the basis of the present invention fall within the scope of protection required by the present invention.

Claims

1. Alveolar lavage fluid collection tank, characterized in that: The alveolar lavage fluid collection tank includes a cylinder and a lid that covers the bottle mouth of the cylinder. The outer layer of the cylinder is a vacuum layer, the inner layer of the cylinder is a liquid nitrogen perfusion layer, and the bottom of the lid is an embedded portion. The embedded portion extends into the interior of the cylinder, and the bottom of the embedded portion is provided with an opening, which extends toward the lid to form an activated carbon storage chamber. The alveolar lavage fluid collection tank also includes a push-pull component, which includes a push-pull rod. The push-pull rod passes through the lid from above, and the bottom end of the push-pull rod is located in the activated carbon storage chamber. The bottom end of the push-pull rod is connected to a tray, in which activated carbon is placed. The tray moves downward when the push-pull rod is pushed downward, blocking the opening at the bottom of the embedded portion from closing. An ultraviolet lamp is provided at the bottom of the interior of the cylinder, and a transparent partition that can transmit ultraviolet light is provided above the ultraviolet lamp. The lid is provided with a liquid nitrogen air vent, which is connected to the liquid nitrogen perfusion layer.

2. The alveolar lavage fluid collection tank according to claim 1, characterized in that: The transparent partition capable of transmitting ultraviolet rays is one of optical quartz glass, ultraviolet-transmitting black glass, soda-lime-silica short-wave ultraviolet-transmitting glass and soda-lime-ultraviolet-transmitting glass.

3. The alveolar lavage fluid collection tank according to claim 1, characterized in that: The push-pull component is provided with an external thread at a position where it passes through the cover, and the cover is provided with an internal thread matching the external thread.

4. The alveolar lavage fluid collection tank according to claim 1, characterized in that: The material of the cylinder is stainless steel or aluminum alloy.

5. The alveolar lavage fluid collection tank according to claim 1, characterized in that: The top end of the push-pull rod is connected with a handle.

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