Fluorescence immunochromatography test strip for quantitatively detecting chicken mycoplasma synoviae antibody, preparation method and detection system

Abstract

The invention discloses a fluorescence immunochromatography test strip for quantitatively detecting a chicken mycoplasma synoviae antibody, the fluorescence immunochromatography test strip comprises a bottom plate, a sample pad, a fluorescent pad, a nitrocellulose membrane and an absorption pad are sequentially arranged on the bottom plate, the fluorescent pad is a time-resolved fluorescent microsphere labeled chicken mycoplasma synoviae MSPB protein, and the nitrocellulose membrane is a nitrocellulose membrane. A detection line and a quality control line are arranged on the nitrocellulose membrane, the detection line is chicken mycoplasma synoviae MSPB protein, and the quality control line is goat anti-chicken IgG, the MSPB protein is successfully prepared, can be specifically combined with a chicken mycoplasma synoviae antibody, and does not react with other poultry virus antibodies.

Description

Technical Field

[0001] This invention belongs to the field of biological detection technology, specifically relating to a fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies. Background Technology

[0002] Mycoplasma synoviae (MS) was first discovered in 1954 and classified in 1982 as belonging to the class Malacostraca, order Mycoplasma, family Mycoplasmatidae, and genus Mycoplasma. MS primarily invades the epithelial cells of the chicken's respiratory tract and air sacs, as well as the synovial bursae and tendon sheaths of joints. It can cause subclinical respiratory diseases, exudative synovitis, tenosynovitis, and bursitis. The disease has a long course and is difficult to eradicate once infected. After infection, the disease will spread within the flock for a long time. In laying hens, it manifests as reduced egg production and autoimmune diseases; in broilers, it mainly affects growth retardation, causing significant economic losses to the poultry industry.

[0003] Mycoplasma cells lack lipopolysaccharides and peptidoglycans on their surface, but they possess abundant lipoproteins. Mycoplasma lipoproteins can participate in the adhesion of host cells and influence the host's immune system. Lipoproteins induce host cell damage by stimulating cells to produce pro-inflammatory factors and cytokines. Simultaneously, mycoplasma lipoproteins can evade the surveillance of the host immune system, allowing them to survive long-term within host cells and promoting or aiding the pathogenesis of other diseases. MSPB protein is a lipoprotein on the membrane of Mycoplasma synoviae in chickens and possesses strong immunogenicity.

[0004] Currently, the main methods for detecting Mycoplasma synoviae in chickens are the serum plate agglutination test (SPAT) and enzyme-linked immunosorbent assay (ELISA). There are no fluorescent immunochromatographic test strips for detecting Mycoplasma synoviae antibodies based on fluorescent immunochromatography. Because SPAT cross-reacts with other avian mycoplasma (such as MG) antibodies, further confirmation requires other methods, which is cumbersome. ELISA is complex, requires specialized personnel and equipment, necessitates a professional laboratory, and is time-consuming, making it unsuitable for on-site testing. This invention provides a fluorescent immunochromatographic test strip for Mycoplasma synoviae antibodies that is simple to operate, highly sensitive, highly specific, and quick, and can quantitatively detect MS antibodies. Summary of the Invention

[0005] To address the shortcomings of existing technologies, this invention provides a fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies, along with its preparation method and detection system. This invention successfully prepares MSPB protein, which can specifically bind to Mycoplasma synoviae antibodies and does not react with other avian virus antibodies. Based on this, this invention has researched and designed a fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies.

[0006] To achieve the above objectives, the present invention provides the following technical solution: A fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies includes a base plate, on which a sample pad, a fluorescent pad, a nitrocellulose membrane, and an absorbent pad are sequentially arranged. The fluorescent pad is time-resolved fluorescent microspheres labeled with Mycoplasma synoviae MSPB protein. The nitrocellulose membrane has a detection line and a control line. The detection line is Mycoplasma synoviae MSPB protein, and the control line is goat anti-chicken IgG.

[0007] As a further embodiment of the present invention, the fluorescent microspheres are polystyrene time-resolved fluorescent microspheres, and the particle size of the fluorescent microspheres is 100-300 nm.

[0008] As a further aspect of the present invention, the amount of protein labeled on the fluorescent pad is 20-40 μg / mL.

[0009] As a further aspect of the present invention, the Mycoplasma synoviae MSPB protein specifically binds to positive serum from chicken synoviae.

[0010] As a further embodiment of the present invention, the antibody coating concentration on the quality control line is 0.8-1.0 mg / mL, and the protein coating concentration on the detection line is 1.0-1.2 mg / mL.

[0011] This invention also provides a method for preparing a fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies, comprising the following steps: The sample pad is prepared by treating the first cellulose membrane with a sample pad blocking solution. The sample pad blocking solution is formulated as follows: Tris buffer containing 1% casein, 1% Tween, and 2% BSA. The first glass fiber membrane is then soaked in the sample pad blocking solution and dried to obtain the sample pad. More preferably, the sample diluent is PB buffer containing 0.2% sodium azide. A fluorescent pad was prepared by spraying chicken synoviocyte mycoplasma protein labeled with fluorescent microspheres onto a second glass cellulose membrane. Sheep anti-chicken IgG antibody was coated onto nitrocellulose membrane to form a control line, and chicken synoviocyte mycoplasma MSPB protein was sprayed onto nitrocellulose membrane to form a detection line. The sample pad, fluorescent pad, nitrocellulose membrane, absorbent pad, and base plate were assembled to obtain a fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies. The present invention also mentions a detection system, including an immunofluorescence rapid assay instrument, a fluorescence immunochromatographic test strip, a sample pad blocking solution and a sample diluent, wherein the fluorescence immunochromatographic test strip is provided with an ID card, and the ID card stores a standard curve.

[0012] The present invention has the following beneficial effects: The fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies provided by this invention can directly perform quantitative detection on collected chicken serum samples. The detection results are positively correlated with serum antibody levels, enabling differential diagnosis of antibodies from field bacterial infections and vaccine infections. It has high sensitivity, good specificity, and is easy to operate. The detection results can be quantitatively read in numerical form within 20 minutes, providing a rapid and accurate detection method for Mycoplasma synoviae.

[0013] To more clearly illustrate the structural features and effects of the present invention, the present invention will be described in detail below with reference to the accompanying drawings and specific embodiments. Attached Figure Description

[0014] Figure 1 This is a schematic diagram of the fluorescent immunochromatographic test strip for the Mycoplasma synoviae antibody mentioned in this invention. Figure 1 The sample marked 8 is the sample in the middle.

[0015] Figure 2 This is a dotted distribution diagram of the sample T / C value × 100 mentioned in this invention.

[0016] Figure 3 This is the analysis diagram of the working characteristic curve mentioned in this invention.

[0017] Figure 4 This is a graph showing the sensitivity test results of the fluorescent immunochromatographic test strip for the Mycoplasma synoviae antibody mentioned in this invention. Detailed Implementation

[0018] The present invention will now be further described in conjunction with the accompanying drawings and relevant knowledge, and will be described clearly and completely. Obviously, the described applications are only some embodiments of the present invention, and not all embodiments.

[0019] Reference Figures 1-4As shown, this invention provides a fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies. The test strip uses a double-antibody sandwich method and includes a base plate 1 and a sample pad 2, a fluorescent pad 3, a nitrocellulose membrane 4, and an absorbent pad 5 sequentially mounted on the base plate 1. The fluorescent pad 3 is coated with Mycoplasma synoviae protein labeled with fluorescent microspheres, i.e., purified MSPB protein is coupled to the fluorescent microspheres. The fluorescent microspheres are time-resolved fluorescent microspheres with a particle size of 100–300 nm. The nitrocellulose membrane has an antibody detection line 6 and a control line 7. The detection line 6 is formed by Mycoplasma synoviae MSPB protein coated on the nitrocellulose membrane; the control line 7 is formed by goat anti-chicken IgG antibody coated on the nitrocellulose membrane. The Mycoplasma synoviae antigen protein MSPB mentioned in this invention, as a lipoprotein on the Mycoplasma synoviae membrane, has strong immunogenicity and can be used as an antigen for the detection of Mycoplasma synoviae.

[0020] This invention also provides a method for preparing a fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies, comprising the following steps: The first cellulose membrane was treated with a sample pad blocking solution and dried at 30–60°C to obtain a sample pad. The sample pad blocking solution was a Tris buffer containing 1% casein, 1% Tween, and 2% BSA, and the sample dilution solution was a PB buffer containing 0.2% sodium azide.

[0021] A fluorescent pad was prepared by spraying chicken synovipositor protein labeled with fluorescent microspheres onto a second glass cellulose membrane and drying it.

[0022] Sheep anti-chicken IgG antibodies were coated onto nitrocellulose membranes to form control lines, and chicken synoviocyte mycoplasma protein was sprayed onto nitrocellulose membranes to form detection lines.

[0023] The sample pad, fluorescent pad, nitrocellulose membrane, absorbent pad, and base plate are assembled to obtain a fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies.

[0024] This invention also provides a method for reading test strips to achieve quantitative analysis of Mycoplasma synoviae antibodies in chickens. The method for reading test strips specifically includes the following steps: (1) Develop a standard curve and send it to the instrument; (2) Take 1 drop of sample with a disposable dropper (or 35 µL with a pipette) and add it to the sample processing tube. Mix well and wait for testing. (3) Use a disposable plastic dropper to vertically add 3 drops (or use a pipette to transfer 100 µL) of the diluted sample into the sample well; (4) After adding the sample, place the test strip at a constant temperature of 30-37℃ for 10 min to react; (5) After the reaction is complete, remove the test card, insert it into the card slot of the immunofluorescence rapid test instrument, select “sample test”, and click “start test” to read the quantitative test results.

[0025] Result Interpretation: A detection value ≥ 50 indicates the presence of MS antibodies in the sample; the higher the detection value, the higher the MS antibody content. A detection value < 50 indicates the absence of MS antibodies in the sample, or that the antibody content is below the detection threshold.

[0026] In this invention, the fluorescent immunochromatographic test strip is based on the immunofluorescence double-antibody sandwich method. Its detection principle is as follows: if the sample to be tested contains anti-Mycoplasma synoviae antibodies, the anti-Mycoplasma synoviae antibodies in the sample will bind to the MSPB antigen labeled with fluorescent microspheres in the sample pad and be chromatographically separated along the nitrocellulose membrane by capillary action. When it reaches the detection area, it is captured by the MSPB antigen fixed on the corresponding detection line (T line). The amount of MSPB antigen labeled with fluorescent microspheres bound on the detection line is proportional to the amount of anti-Mycoplasma synoviae antibodies in the sample. The content of anti-Mycoplasma synoviae antibodies in the sample to be tested is calculated by the fluorescence analyzer based on the fluorescence intensity and the standard curve.

[0027] Based on the above detection principle, this invention employs conventional fluorescent immunochromatographic test strip technology. Following standard procedures, fluorescent microspheres are directly labeled with MSPB antigen, and the content of anti-Mycoplasma synoviae antibodies in the test sample is quantitatively analyzed using a fluorescent immunoassay analyzer. Quantitative analysis of the samples allows for the classification of positive samples into strongly positive and weakly positive categories, distinguishing different levels of Mycoplasma synoviae infection. This reduces the false positive rate caused by traditional qualitative analysis and provides a new approach for the prevention and control of Mycoplasma synoviae.

[0028] Unless otherwise specified, the experimental methods used in the following examples are conventional methods, and the experimental materials used are commercially available.

[0029] Example 1: Specific preparation method of Mycoplasma synoviae antigen protein MSPB in chicken synovial fluid. 1.1 Analysis of MSPB protein biological information: The amino acid sequence of the MSPB protein was obtained from the NCBI database (Genbank: AKB11074.1).

[0030] Transmembrane helix analysis was performed using TMHMM-2.0 (https: / / services.healthtech.dtu.dk / services / TMHMM-2.0 / ); signal peptide prediction for the antigen protein was performed using SignalP-6.0, and the signal peptide fragment was removed if present (https: / / services.healthtech.dtu.dk / services / SignalP-6.0 / ); hydrophobicity prediction was performed using a protein hydrophobicity analysis website (https: / / www.novopro.cn / tools / protein-hydrophilicity-plot.html); functional domains of the MSPB protein were predicted using the online software Prosite (https: / / prosite.expasy.org / cgi-bin / prosite / ); and antigenic epitopes of the MSPB protein were predicted using a B-cell epitope prediction website (http: / / tools.iedb.org / bcell / ).

[0031] 1.2 Construction and identification of recombinant plasmids: The codon-optimized MSPB gene sequence was ligated into the pET-28a prokaryotic expression vector to construct a recombinant plasmid, which was then transformed into... E. coli DH5α competent cells were screened for transformants using solid LB agar plates containing Kan resistance. After plating and culturing at 37°C for 12 h, single colonies were picked and added to liquid LB agar containing Kan resistance. The culture was then carried out on a shaker at 37°C and 220 rpm for 10–12 h. A portion of the bacterial culture was then subjected to PCR identification in a clean bench. Positive clones were sent for sequencing. Recombinant bacteria with correct PCR and sequencing results were named pET-28a-MSMSPB-DH5α. A portion of the bacterial culture was mixed with 50% glycerol solution and stored at -80°C. The remaining portion was used to extract plasmids according to the plasmid miniprep kit instructions. After determining the plasmid concentration, the plasmid was named pET-28a-MSMSPB.

[0032] 1.3 Transformation and identification of recombinant plasmids: Transform an appropriate amount of recombinant plasmid pET-28a-MSMSPB into... E. coli After BL21 competent cells were cultured, transformants were screened using solid LB plates containing Kan resistance, and bacterial culture was identified by PCR. Positive clones were sent to the company for sequencing. If the sequencing results were correct, recombinant engineered bacteria were prepared. Recombinant bacteria with correct PCR and sequencing results were named... E. coli BL21-pET-28a-MSMSPB.

[0033] 1.4 Induced expression of MSPB protein: The above obtained E. coli BL21-pET-28a-MSMSPB glycerol bacteria were inoculated at a volume ratio of 1:100 into liquid LB medium containing Kan resistance and activated at 37°C and 220 rpm for 12–16 h using a shaker. The activated bacterial culture was then expanded and cultured under the same conditions for 3–4 h until the OD value of the bacterial culture was determined. 600 The value was between 0.6 and 0.8, and then IPTG was added to induce expression for 6 hours.

[0034] 1.5 Expression and purification of MSPB protein: After induction, the bacterial cells were collected, washed, lysed, and centrifuged. The supernatant and precipitate were collected separately. Whole cells before and after induction, as well as the supernatant and precipitate after lysis, were sampled separately for SDS-PAGE electrophoresis analysis and identification. To ensure that the recombinant protein fused with the His tag and was successfully expressed, all the samples obtained above were simultaneously identified by Western blotting. After the identification was correct, the His-tag protein purification kit was used for purification, and MSPB protein was successfully obtained.

[0035] Example 2: Detection method of fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies in chickens The fluorescent immunochromatographic test strip provided in this embodiment adopts a double-antibody sandwich method, including a base plate and a sample pad, a fluorescent pad, a nitrocellulose membrane, and an absorbent pad sequentially mounted on the base plate. The fluorescent pad is coated with MSPB protein labeled with fluorescent microspheres. The fluorescent microspheres are time-resolved fluorescent microspheres with a particle size of 100-300 nm. The nitrocellulose membrane has a detection line and a control line. The detection line is formed by Mycoplasma synoviae protein coated on the nitrocellulose membrane. The control line is formed by goat anti-chicken IgG antibody coated on the nitrocellulose membrane.

[0036] Example 3: Preparation method of fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies in chickens 1. Prepare the sample pad blocking solution: Tris buffer containing 1% casein, 1% Tween, and 2% BSA.

[0037] 2. Use the sample pad sealing solution to evenly wet the first cellulose membrane, and dry it at 30-60℃ to obtain the sample pad.

[0038] 3. Spray the chicken synovipositor protein containing fluorescent microspheres onto the second glass cellulose membrane and dry it to obtain a fluorescent pad.

[0039] 4. Streak the coating solutions of Mycoplasma synoviae MSPB protein and goat anti-chicken IgG antibody on nitrocellulose membranes, respectively. Then, dry the nitrocellulose membranes in an oven to form test lines and control lines, and place them in a drying oven for later use.

[0040] 5. Assembly of the fluorescent immunochromatographic test strip: The sample pad, fluorescent pad, nitrocellulose membrane, and absorbent pad are sequentially overlapped at specific positions on the base plate. After assembly, the strip is cut to a fixed width to obtain the fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies. See the schematic diagram of the test strip. Figure 1 .

[0041] 6. Prepare sample dilution solution: PB buffer with 0.2% sodium azide.

[0042] Example 4: Reading method of fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies in chickens Develop a standard curve and send it to the instrument. Use a disposable dropper to add 1 drop (or a pipette to add 35 µL) of sample to the sample processing tube, mix well, and use a disposable plastic dropper to vertically add 3 drops (or a pipette to add 100 µL) of diluted sample to the sample well. After adding the sample, place the test strip in a constant temperature environment of 30-37℃ for 10 min to react. After the reaction is complete, insert the test card into the card slot of the immunofluorescence rapid test instrument, select "Sample Test", and click "Start Test" to read the quantitative test results.

[0043] Example 5: Performance evaluation of a fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies in chickens. 5.1 Determination of Critical Values ​​for Positive and Negative Sex Fifty clinically positive serum samples (1#-50#) and fifty clinically negative serum samples (51#-100#) were collected and tested according to the test strip reading method. The T / C value × 100 was used as the reading result. The results are shown in Table 1. ROC curve analysis and dot distribution plots were performed, and the results are as follows. Figure 2 and Figure 3 As shown, Area=1.000 indicates high diagnostic value. A cutoff value of 50 corresponds to a sensitivity of 100% and a specificity of 100%. Therefore, the cutoff value is set at 50. A T / C value × 100 greater than 50 is considered positive, and vice versa.

[0044] Table 1. Results of clinical sample testing using fluorescent immunochromatographic test strips.

[0045] 5.2 Sensitivity Testing MS standard positive serum was serially diluted 2-fold. The 2-fold dilutions were labeled M1, M2, M3, M4, M5, and M6, respectively. M1 through M6 were slowly added to the sample wells, and the tests were performed according to the test strip reading method. The results obtained using a fluorescence immunoassay analyzer are shown in Table 2. These results indicate that the prepared fluorescence immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies in chickens has high sensitivity.

[0046] Table 2. Results of sensitivity testing of the fluorescence immunochromatographic test strips

[0047] 5.3 Specificity Detection The test strips were used to test for positive serum samples of Mycoplasma synoviae (N1), Newcastle disease (N2), avian influenza (N3), infectious bursal disease (N4), and adenovirus (N5). Following the above procedures, the samples were added to the sample pad of the test strip. All results were negative, as shown in Table 3. These results demonstrate that the prepared fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies in chickens has good specificity.

[0048] Table 3. Specific detection results of the fluorescence immunochromatographic test strips

[0049] 5.4 Stability Testing The stability of the quantitative detection strip for Mycoplasma synoviae antibody was verified by placing it at room temperature. Sensitivity and specificity were tested at 0, 3, 6, 9, and 12 months after placement, following the procedures outlined in 5.2 and 5.3 above. The results were recorded for each test, and are shown in Table 4. These results indicate that the prepared quantitative detection strip for Mycoplasma synoviae antibody exhibits good stability.

[0050] Table 4. Results of stability test of fluorescence immunochromatographic test strips

[0051] 5.5 Testing of clinical samples Thirty samples were collected from clinically positive chicken farms for testing. The testing method was as follows: chickens were fixed, blood was collected from the subwing vein, serum was separated, one drop of serum was added to the sample processing solution, mixed thoroughly, and three drops were added to the sample well of the test strip. The strip was placed at a constant temperature of 30-37℃ for 10 minutes. After the reaction, the test card was removed, inserted into the immunofluorescence rapid assay instrument, "sample testing" was selected, and "start testing" was clicked to read the quantitative test results. When the detection value is ≥50, it indicates that the sample contains MS antibodies; the higher the detection value, the higher the MS antibody content. When the detection value is <50, it indicates that the sample does not contain MS antibodies, or the antibody content is lower than the detection threshold. The samples were also tested using the plate agglutination (SPA) method. The test results are shown in Table 5. The SPA detection positivity rate was 18 / 30, and the detection positivity rate of the test strip of this invention was 16 / 30. The concordance rate calculated using the Kappa method was 93.33%, indicating that the test strip has good accuracy.

[0052] Table 5. Detection results of clinical samples using fluorescence immunochromatographic test strips and the SPA method.

[0053] Note: +++ indicates a strong positive agglutination reaction; ++ indicates a positive agglutination reaction; + indicates a weak positive agglutination reaction; - indicates no agglutination reaction.

[0054] 5.6 Correspondence between test strip readings and clinical results Chickens were immunized with a live MS vaccine at 5 weeks of age and with an inactivated MS vaccine at 19 weeks of age. Blood samples were collected from 30 chickens at 19, 23, 27, 31, and 35 weeks of age, and serum was separated. The results were measured using this test strip, and the results are shown in Table 6. The results indicate that the test strip readings conform to the antibody fluctuation pattern after vaccine immunization, and the results can be used to quantitatively detect Mycoplasma synoviae antibodies in chickens.

[0055] Table 6. Detection results of test strips at different time points after immunization.

[0056] In summary, this invention provides a fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae in chickens. It allows for direct detection of chicken serum samples, exhibits high concordance with the SPA method, and enables early detection of MS infection in chickens. The test strip is sensitive, rapid, highly specific, and easy to use, providing results within 20 minutes. It solves the problems of the ELISA method, which requires specialized personnel and equipment, is time-consuming and labor-intensive, and is unsuitable for small and medium-sized poultry farmers.

[0057] The technical principles of the present invention have been described above with reference to specific embodiments, which are merely preferred embodiments of the present invention. The scope of protection of the present invention is not limited to the above embodiments; all technical solutions falling within the scope of the present invention's concept are within its protection scope. Those skilled in the art can conceive of other specific embodiments of the present invention without creative effort, and these embodiments will all fall within the protection scope of the present invention.

Claims

1. A fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies, characterized in that, The device includes a base plate on which a sample pad, a fluorescent pad, a nitrocellulose membrane, and an absorbent pad are sequentially arranged. The fluorescent pad is time-resolved fluorescent microspheres labeled with Mycoplasma synoviae MSPB protein. The nitrocellulose membrane has a detection line and a control line. The detection line is Mycoplasma synoviae MSPB protein, and the control line is goat anti-chicken IgG.

2. The fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies as described in claim 1, characterized in that, The fluorescent microspheres are polystyrene time-resolved fluorescent microspheres with a particle size of 100-300 nm.

3. The fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies as described in claim 2, characterized in that, The amount of protein labeled on the fluorescent pad is 20-40 μg / mL.

4. The fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies as described in claim 3, characterized in that, The MSPB protein of Mycoplasma synoviae specifically binds to positive serum from chicken synoviae.

5. The fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies as described in claim 2, characterized in that, The antibody coating concentration on the quality control line is 0.8-1.0 mg / mL, and the protein coating concentration on the detection line is 1.0-1.2 mg / mL.

6. A method for preparing a fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies, characterized in that, Includes the following steps: The first cellulose membrane was treated with a sample pad sealing solution to prepare the sample pad; A fluorescent pad was prepared by spraying chicken synoviocyte mycoplasma protein labeled with fluorescent microspheres onto a second glass cellulose membrane. Sheep anti-chicken IgG antibody was coated onto nitrocellulose membrane to form a control line, and chicken synoviocyte mycoplasma MSPB protein was sprayed onto nitrocellulose membrane to form a detection line. The sample pad, fluorescent pad, nitrocellulose membrane, absorbent pad, and base plate were assembled to obtain a fluorescent immunochromatographic test strip for the quantitative detection of Mycoplasma synoviae antibodies.

7. The method for preparing a fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies as described in claim 6, characterized in that, The sample pad blocking solution is formulated as follows: Tris buffer containing 1% casein, 1% Tween, and 2% BSA; the sample pad is prepared by impregnating the first glass fiber membrane with the sample pad blocking solution and then drying it.

8. The method for preparing a fluorescent immunochromatographic test strip for quantitative detection of Mycoplasma synoviae antibodies as described in claim 7, characterized in that, The sample diluent was PB buffer containing 0.2% sodium azide.

9. A detection system, characterized in that, The invention includes an immunofluorescence rapid assay instrument, a fluorescence immunochromatographic test strip according to any one of claims 1 to 5, a sample pad blocking solution, and a sample diluent, wherein the fluorescence immunochromatographic test strip is provided with an ID card, and the ID card stores a standard curve.

Images