Method and Application for Detecting Carcinoembryonic Antigen (CEA) without Using Fetal Embryo Antigen
By simplifying the CEA detection process of fetal embryo antigen, the washing steps are reduced from four to two, which solves the problems of cumbersome detection steps and radiation hazards, and achieves efficient and safe detection results.
Patent Information
- Application Number
- CN202111374852.3
- Authority / Receiving Office
- CN · China
- Patent Type
- Patents(China)
- Current Assignee / Owner
- Filing Date
- 2021-11-19
- Publication Date
- 2025-07-25
- Estimated Expiration
- 2041-11-19
AI Technical Summary
The existing fetal embryo antigen CEA detection methods are cumbersome and time-consuming, resulting in high laboratory testing pressure and radioactive radiation hazards.
The detection process is simplified, the washing steps are reduced from four times to two times, the two washing steps after the first incubation are omitted, and the non-diagnostic release-free fetal embryo antigen CEA detection method is adopted.
It improves the detection efficiency, reduces the working intensity of the experimenter, reduces the generation of radioactive waste liquid, reduces the hazards of radiation, and has stable and reliable results.
Smart Images
Figure CN114324868B_ABST
Abstract
Description
Technical Field
[0001] The present invention relates to the technical field of medical testing, and particularly to a non-radioactive method for detecting carcinoembryonic antigen (CEA) of fetal embryo and its application. Background Art
[0002] Carcinoembryonic antigen (CEA) of fetal embryo is one of the important indicators for tumor screening and is widely used in general tumor screening. As a result, a large number of specimens usually need to be detected. Taking this center as an example, the current average daily number of specimens is more than 600, and the number of specimens can reach 2000 - 3000 per day at the peak. However, the detection method for this project has cumbersome steps, long experimental time, tight staffing in the department, and great pressure on laboratory testing. In addition, the non-radioactive detection method uses iodine-125 as a tracer, and it is well known that radioactive iodine-125 is harmful to the human body and the environment due to radiation. Summary of the Invention
[0003] Based on this, in view of the above problems, it is necessary to provide a non-radioactive method for detecting carcinoembryonic antigen (CEA) of fetal embryo for non-diagnostic purposes. Using this method for detecting CEA of fetal embryo simplifies the detection steps, reduces the experimental time, and reduces the working intensity of experimental personnel, thereby improving work efficiency.
[0004] A non-radioactive method for detecting carcinoembryonic antigen (CEA) of fetal embryo for non-diagnostic purposes includes the following steps:
[0005] Preparation: Take a coated tube and number it.
[0006] Sample addition: Add a sample solution in a predetermined amount.
[0007] First incubation: Incubate at 37 ± 2 °C for a first predetermined time, and then aspirate the liquid in the tube.
[0008] Label addition: Directly add a labeled antibody solution to the coated tube that has undergone the first incubation.
[0009] Second incubation: Incubate at 37 ± 2 °C for a second predetermined time, and then aspirate the liquid in the tube.
[0010] Washing: Add a washing solution to the coated tube that has undergone the second incubation for washing, and then aspirate the washing solution.
[0011] Detection: Perform detection on a machine.
[0012] In long-term practice, the inventor found that the conventional method for detecting carcinoembryonic antigen (CEA) in fetal embryos without radioactivity usually uses a commercial kit. According to the instructions of the kit, after the first incubation to allow the analyte to fully react with the coating on the coated tube, washing solution is added and washed four times according to the conventional operation procedure for immunological items (twice after the first incubation and twice after the second incubation). At this time, the purpose of washing is to prevent the HOOK effect caused by too high antigen concentration, but the kit manufacturer did not investigate the rationality and necessity of this operation in the method for detecting CEA in fetal embryos without radioactivity for different detection items.
[0013] With the development of enzyme immunoassay and chemiluminescence, on the one hand, the performance of detection reagents is continuously improving. On the other hand, based on the inventor's in-depth experience in the field of medical testing and the analysis of large clinical sample data, the inventor found that the CEA project has two characteristics: a relatively low positive rate and low positive result values.
[0014] On this basis, the inventor creatively proposed to reduce the original four washes to two washes, omitting the two washing steps after the first incubation in the middle of the experiment. And through experiments, it was confirmed that omitting the washing after the first incubation and reducing the number of washes had no effect on the detection results of CEA in fetal embryos without radioactivity, and the results were stable.
[0015] In one embodiment, in the sample addition step, the sample solution is prepared by the following method: collecting a blood sample, centrifuging to separate the serum, and that is obtained.
[0016] In one embodiment, in the sample addition step, the predetermined quantity is 200 ± 50 μl per tube.
[0017] In one embodiment, in the first incubation step, the first predetermined time is 1 ± 0.5 hr.
[0018] In one embodiment, in the second incubation step, the second predetermined time is 1 ± 0.5 hr.
[0019] In one embodiment, in the labeling step, the labeled antibody is 125 I-labeled anti-CEA monoclonal antibody, and the addition amount is 200 ± 50 μl.
[0020] In one embodiment, in the washing step, the washing step is repeated 2 times.
[0021] In one embodiment, in the washing step, the addition amount of the washing solution is 1000 ± 200 μl per tube each time.
[0022] The present invention also discloses a sample pretreatment device for the detection of carcinoembryonic antigen (CEA) by the non-radioactive method, and the sample is processed according to the following method:
[0023] Preparation: Take the coated tubes and number them.
[0024] Sampling: Add the sample solution in a predetermined amount.
[0025] First incubation: Incubate at 37 ± 2 °C for the first predetermined time, and then aspirate the liquid in the tube.
[0026] Labeling: Directly add the labeled antibody solution into the coated tube that has undergone the first incubation.
[0027] Second incubation: Incubate at 37 ± 2 °C for the second predetermined time, and then aspirate the liquid in the tube.
[0028] Washing: Add the washing solution into the coated tube that has undergone the second incubation for washing, and then aspirate the washing solution; thus, the sample to be loaded onto the machine is obtained.
[0029] The present invention also discloses a non-radioactive method CEA detection device for fetal embryo antigen, comprising:
[0030] A sample processing module, including the above sample pretreatment device;
[0031] A detection module, configured to detect the sample to be loaded onto the machine processed by the sample processing module;
[0032] An output module, configured to output the detection result obtained by the detection module.
[0033] Compared with the prior art, the present invention has the following beneficial effects:
[0034] In the non-diagnostic purpose non-radioactive method for detecting carcinoembryonic antigen (CEA) of the present invention, through a large amount of research and data analysis, it is found that the CEA item has two characteristics: relatively low positive rate and low positive result value. On this basis, the inventor creatively proposes to reduce the original four washings to two washings, and omit the two washing steps after the first incubation in the middle process of the experiment, thus improving the original detection method. And it is confirmed by experiments that there is no obvious difference between the detection process after improvement and the result before improvement, and the result is stable. It is particularly suitable for scientific research on large-scale tumor marker screening in hospital physical examination centers and third-party inspection institutions, etc.
[0035] Taking the central laboratory where the inventor works as an example, calculated according to the specimen volume of 600 in each department, after the method is improved, the number of washings in the experimental process is reduced from the original 4 times to 2 times, and the original 4 times of water aspiration is reduced to 2 times. That is to say, after the department improves the method, it can directly save about 1600 liters of distilled water per day (the washing solution is diluted with distilled water), and the suction device works 30 minutes less (mainly responsible for liquid aspiration work).
[0036] More importantly, during the detection process before improvement, at least 4 people were required to complete the washing step (one person for water injection and three people for water absorption). However, in the improved method, since the number of washing steps and water absorption steps was reduced by 2 times each, one labor force could be saved for the laboratory during the experiment.
[0037] At the same time, the improved method reduces the number of washings by 2 times, greatly reduces the generation of a large amount of radioactive waste liquid, greatly alleviates the storage pressure of radioactive waste liquid, and most importantly, reduces the radiation hazards of radioactive substances to the human body and the environment. Description of the Drawings
[0038] Figure 1 It is a scatter plot drawn according to the Deming regression result and the maximum allowable error in Example 1.
[0039] Figure 2 It is a schematic diagram of the bias of the Deming regression result in Example 1.
[0040] Figure 3 It is a scatter plot drawn according to the linear regression result and the maximum allowable error in Example 1.
[0041] Figure 4 It is a schematic diagram of the bias of the linear regression result in Example 1.
[0042] Figure 5 It is the ratio of the distance between each measured value and the target value to the total error in Example 2. Detailed Implementation Modes
[0043] To facilitate the understanding of the present invention, the present invention will be described more comprehensively below with reference to the relevant drawings. Preferred embodiments of the present invention are shown in the drawings. However, the present invention can be implemented in many different forms and is not limited to the embodiments described herein. On the contrary, these embodiments are provided to make the understanding of the disclosure of the present invention more thorough and comprehensive.
[0044] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by those of ordinary skill in the technical field to which the present invention belongs. The terms used in the description of the present invention herein are only for the purpose of describing specific embodiments and are not intended to limit the present invention. The term "and / or" used herein includes any and all combinations of one or more of the related listed items.
[0045] Unless otherwise specified, the raw materials used in the following examples are all commercially available; unless otherwise specified, the methods used in the following examples can be implemented by conventional methods.
[0046] Example 1
[0047] A method for detecting carcinoembryonic antigen (CEA) of fetal embryo antigen without radioactivity, using iodine 125 I carcinoembryonic antigen immunoradiometric assay kit (manufacturer: Tianjin Xiehe Pharmaceutical Technology Group Co., Ltd., specification: 100 person-times per box), including the following steps:
[0048] 1) Number the coated tubes that have been equilibrated to room temperature;
[0049] 2) Add 200 microliters of sample solution to each tube;
[0050] The sample solution is prepared by the following method: collect blood samples, centrifuge and separate to obtain serum.
[0051] 3) After incubating at 37°C for 1 hour, suck dry the liquid in the tube with a suction device;
[0052] 4) Add 200 microliters of labeled antibody solution to each tube;
[0053] The labeled antibody is 125 I-labeled anti-CEA monoclonal antibody
[0054] 5) After incubating at 37°C for 1 hour, suck dry the liquid in the tube with a suction device;
[0055] 6) Add 1000 microliters of washing solution to each tube and wash with the washing solution;
[0056] The washing solution is prepared by the following method: take 11 ML / bottle of washing solution, dilute it with 20 times the volume of distilled water per bottle, and mix well before use.
[0057] 7) Suck dry the liquid in the tube with a suction device;
[0058] 8) Repeat step 6) once;
[0059] 9) Suck dry the liquid in the tube with a suction device and measure on the machine.
[0060] Comparative Example 1
[0061] A method for detecting carcinoembryonic antigen (CEA) of fetal embryo antigen without radioactivity, using the kit in Example 1 and following the method in the reagent instruction manual, including the following steps:
[0062] 1) Number the coated tubes that have been equilibrated to room temperature;
[0063] 2) Add 200 microliters of sample solution to each tube;
[0064] The sample solution is prepared according to the method in Example 1.
[0065] 3) After incubating at 37°C for 1 hour, suck dry the liquid in the tube with a suction device;
[0066] 4) Add 1000 microliters of washing solution to each tube and wash;
[0067] The washing solution was prepared according to the method of Example 1.
[0068] 5) Use a suction device to dry the liquid in the tube;
[0069] 6) Repeat step 4) once;
[0070] 7) Repeat step 5) once;
[0071] 8) Add 200 μL of labeled antibody solution to each tube;
[0072] 9) After incubating at 37°C for 1 hour, use a suction device to dry the liquid in the tube;
[0073] 10) Add 1000 μL of washing solution to each tube and wash;
[0074] 11) Use a suction device to dry the liquid in the tube;
[0075] 12) Repeat step 10) once;
[0076] 13) Use a suction device to dry the liquid in the tube and measure on the machine.
[0077] Experimental Example 1
[0078] This experimental example verifies and compares the methods in Example 1 and Comparative Example 1.
[0079] 1. Method.
[0080] Randomly select 40 samples and detect them according to the methods of Example 1 and Comparative Example 1 respectively.
[0081] 2. Results.
[0082] 1) Detection data results.
[0083] The detection results are shown in the following table.
[0084] Table 1. Comparison of CEA results
[0085]
[0086]
[0087] It can be seen from the comparison of the detection result data of the two methods before and after the CEA improvement that when the same sample is detected by the two methods of Comparative Example 1 and Example 1 before and after the improvement, omitting the washing step after the first incubation does not have a substantial impact on the detection results.
[0088] 2) Consistency analysis.
[0089] The above comparison data is analyzed by EP Evaluator CLSI EP9, and the results are as Figures 1-4 shown, among which, Figure 1 represents a scatter plot drawn based on the Deming regression result and the maximum allowable error, which can represent the correctness of the result to a certain extent. The results in the figure show that the consistency between the verification method and the method to be verified is relatively good. Figure 2 represents the bias of the Deming regression result; Figure 3 represents a scatter plot drawn based on the conventional linear regression result and the maximum allowable error; Figure 4 represents the bias of the conventional linear regression result.
[0090] The results show that the consistency of the two large groups of data detected in Example 1 and Comparative Example 1 passes, and there is no significant difference.
[0091] Experimental Example 2
[0092] In this experimental example, the washing steps in the CEA method for embryo antigen without a method are adjusted and verified and compared.
[0093] 1. Method.
[0094] Randomly select 29 samples, and refer to the conventional method (i.e., the method of Comparative Example 1), and detect them according to the following group methods respectively.
[0095] Single-washing group: Only retain the single washing process after the second incubation, and omit the washing after the first incubation and the single washing after the second incubation.
[0096] Double-washing group: That is, the method of Example 1.
[0097] Triple-washing group: Only omit the second washing after the first incubation, retain the single washing after the first incubation, and the two washings after the second incubation.
[0098] Quadruple-washing group: That is, the method of Comparative Example 1.
[0099] 2. Results.
[0100] 1) Detection data results.
[0101] The detection results are shown in the following table.
[0102] CEA result comparison
[0103]
[0104]
[0105] From the comparison of the above result data, it can be seen that the main reason for the positive results in the single-washing group may be that the iodine label was not washed clean, and the results are unacceptable.
[0106] 2) Consistency analysis.
[0107] The above comparison data is analyzed by the Multiple Instrument Comparison of EP Evaluator, and the results are as Figure 5 shown, where Figure 5 : It represents the ratio of the distance between each measured value and the target value to the total error. If this value is less than -1 or greater than 1, the result is unacceptable.
[0108] The results show that there is no significant difference in the results of two washes, three washes, and four washes, proving that the improved two - wash method maximally simplifies the detection process and the results are reliable and stable.
[0109] The technical features of the above - described embodiments can be combined arbitrarily. For the sake of brevity of description, not all possible combinations of the technical features in the above - described embodiments are described. However, as long as there is no contradiction in the combination of these technical features, it should be considered as the scope recorded in this specification.
[0110] The above - described embodiments only represent several implementation manners of the present invention. Their descriptions are relatively specific and detailed, but they should not be construed as limiting the scope of the invention patent. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several modifications and improvements can still be made, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the present invention patent should be subject to the appended claims.
Claims
1. A method for detecting carcinoembryonic antigen (CEA) of fetal embryos without using radioactive substances for non-diagnostic purposes, characterized in that, It consists of the following steps: Preparation: Take the coated tubes and number them; Sampling addition: Add the sample solution in a predetermined amount; First incubation: Incubate at 37±2°C for the first predetermined time, and aspirate the liquid in the tube; Label addition: Directly add the labeled antibody solution into the coated tube that has undergone the first incubation; Second incubation: Incubate at 37±2°C for the second predetermined time, and aspirate the liquid in the tube; Washing: Add the washing solution to the coated tube that has undergone the second incubation for washing, and aspirate the washing solution. The addition amount of the washing solution is 1000±200 μl per tube each time, and repeat the washing step 2 times; Detection: Conduct detection on the machine.
2. The method for detecting carcinoembryonic antigen CEA without releasing fetal embryo antigen according to claim 1, characterized in that, In the sampling addition step, the sample solution is prepared by the following method: Collect blood samples, centrifuge and separate to obtain serum; 3. The method for detecting carcinoembryonic antigen CEA without releasing fetal antigen according to claim 2, wherein In the sampling addition step, the predetermined amount is 200±50 μl per tube; 4. The method for detecting carcinoembryonic antigen CEA without releasing fetal embryo antigen according to claim 1, wherein, In the first incubation step, the first predetermined time is 1±0.5 hr; 5. The method for detecting carcinoembryonic antigen CEA by the method of fetal embryo antigen-free release according to claim 1, characterized in that, In the second incubation step, the second predetermined time is 1±0.5 hr.
6. The method for detecting carcinoembryonic antigen CEA without releasing fetal embryo antigen according to any one of claims 1-5, characterized in that, In the labeling step, the labeled antibody is 125 monoclonal anti-CEA antibody labeled with I, and the addition amount is 200 ± 50 μl.