Application of SMCP in preparation of rheumatoid arthritis diagnosis product

Abstract

The invention discloses application of SMCP in preparation of a rheumatoid arthritis diagnosis product, an anti-SMCP antibody in a biological sample of a patient is used as a marker, and the diagnosis product is used for detecting the content of the anti-SMCP antibody. The SMCP is a polypeptide mixture which is prepared from citrullinated SR-A polypeptide, citrullinated MCV polypeptide, citrullinated caspase14 polypeptide and citrullinated PCOLCE polypeptide. According to the present invention, the anti-SMCP antibody can be subjected to one-time detection so as to achieve the accurate diagnosis of rheumatoid arthritis (RA); besides, the anti-SMCP antibody also has better auxiliary diagnostic value for RA patients with negative anti-CCP antibody, and when the anti-SMCP antibody and the anti-CCP antibody are combined for diagnosis, the RA detection rate can be further improved under the condition of keeping high specificity, and the targets of early diagnosis, early treatment and reduction of disability rate can be more favorably realized.

Description

Technical Field

[0001] The present invention relates to the technical field of disease diagnosis, and in particular to the application of SMCP in the preparation of a product for diagnosis of rheumatoid arthritis. Background Art

[0002] Rheumatoid arthritis (RA) is a chronic rheumatic disease characterized by progressive joint damage and extra-articular manifestations, which can lead to permanent disability. RA is the most common systemic autoimmune disease among rheumatic inflammatory musculoskeletal diseases. Epidemiological studies have shown that the prevalence of RA in my country is approximately 0.28% (Li, R., et al., 中国八种常见风湿性疾病的流行病学:在北京进行的大规模横断面调查 。 北京。 Rheumatology (Oxford), 2012. 51(4): p. 721-9). Early diagnosis and treatment of RA can prevent or significantly slow the progression of joint damage in up to 90% of patients (Aletaha, D. and JS Smolen, 类风湿关节炎的诊断 与管理:综述。 Jama, 2018. 320(13): p.1360-1372), thereby preventing irreversible disability, which is of great significance for improving the quality of life of patients, reducing the economic burden of families, and retaining social labor force.

[0003] Rheumatoid arthritis is a heterogeneous disease that can be subdivided into ACPA-positive and negative RA. Some patients have elevated ACPA levels years before the onset of clinical symptoms. ACPA is present in 50-70% of RA patients (van Delft, MAM and TWJ Huizinga, 类风湿关节炎自身抗体概述 。 J Autoimmun, 2020. 110: p. 102392), is known to recognize a variety of citrullinated antigens, such as alpha enolase, fibrinogen, filaggrin, vimentin, and type II collagen (CII). The identification of citrullinated epitopes as targets led to the development of first- and later second-generation anti-cyclic citrullinated peptide (anti-CCP) antibody assays. Currently, imported ACPA detection kits represented by CCP3 are widely used in clinical practice, but they are expensive and at least 30% of seronegative patients still do not receive timely diagnosis and treatment (Bugatti, S., et al., 自身抗体阴性的类风湿关节炎:对风湿病学家来说仍然是一项挑战 。 Lancet Rheumatol, 2023. 5(12): p. e743-e755).

[0004] New targets for ACPA have successfully revealed ACPA activity in some anti-CCP-negative RA patients, and an algorithm for assessing ACPA activity has shown that up to 50% of patients who test negative for anti-CCP2 or anti-CCP3 show ACPA activity (Savvateeva, E., et al., 用于类风湿关节炎诊断和治疗的多种生物标志物方法 。 Crit Rev Clin Lab Sci, 2021. 58(1): p. 17-28). There may be non-overlapping distributions of these new targets in RA patients, which provides a promising prospect for combined diagnosis of RA. Summary of the invention

[0005] The purpose of the present invention is to address the above-mentioned problems existing in the diagnosis of RA in the prior art, and to provide a polypeptide mixture consisting of citrullinated SR-A polypeptide, citrullinated MCV polypeptide, citrullinated caspase14 polypeptide and citrullinated PCOLCE polypeptide, namely, SMCP, for use in the preparation of a diagnostic product for rheumatoid arthritis. By performing a one-time detection of antibodies to the above four polypeptides in the patient's biological sample, namely, anti-SMCP antibodies, accurate diagnosis of RA can be achieved, thereby improving the detection rate of RA patients.

[0006] The technical solution of the present invention is described in detail as follows: In the first aspect, the present invention provides the use of SMCP in the preparation of a rheumatoid arthritis diagnostic product, wherein SMCP is a polypeptide mixture consisting of a citrullinated SR-A polypeptide, a citrullinated MCV polypeptide, a citrullinated caspase14 polypeptide and a citrullinated PCOLCE polypeptide, and the amino acid sequence is as follows: Citrullinated SR-A polypeptide: KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit (SEQ ID NO: 1); Citrullinated MCV polypeptide: PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP (SEQ ID NO: 2); Citrullinated caspase14 peptide: Cit-GEQ-Cit-DPGETVGGDE (SEQ ID NO: 3); Citrullinated PCOLCE polypeptide: KTLP-Cit-GTAKEGQGPGPK-Cit-G (SEQ ID NO: 4).

[0007] The diagnostic product is used to detect the content of anti-SMCP antibodies. Compared with normal healthy people, patients with systemic lupus erythematosus, Sjögren's syndrome and osteoarthritis, the content of anti-SMCP antibodies in biological samples of rheumatoid arthritis patients is significantly increased.

[0008] Optionally or preferably, the biological sample is blood, plasma, serum or joint fluid.

[0009] Optionally or preferably, the citrullinated SR-A polypeptide, citrullinated MCV polypeptide, citrullinated caspase14 polypeptide and citrullinated PCOLCE polypeptide are all connected to BSA (bovine serum albumin). Specifically, the carrier protein BSA is coupled to the N-terminus of the amino acid sequence through a cysteine ​​residue, and the amino acid sequence after coupling is as follows: Citrullinated SR-A polypeptide: BSA-C-KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit (SEQ ID NO: 5); Citrullinated MCV polypeptide: BSA-C-PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP (SEQ ID NO: 6); Citrullinated caspase14 peptide: BSA-C-Cit-GEQ-Cit-DPGETVGGDE (SEQ ID NO: 7); Citrullinated PCOLCE polypeptide: BSA-C-KTLP-Cit-GTAKEGQGPGPK-Cit-G (SEQ ID NO: 8).

[0010] Optionally or preferably, the diagnostic product is an ELISA test kit.

[0011] In a second aspect, the present invention provides the use of SMCP combined with CCP in the preparation of a rheumatoid arthritis diagnostic product, wherein the SMCP is a polypeptide mixture consisting of a citrullinated SR-A polypeptide, a citrullinated MCV polypeptide, a citrullinated caspase14 polypeptide and a citrullinated PCOLCE polypeptide, and the amino acid sequence is as follows: Citrullinated SR-A polypeptide: KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit (SEQ ID NO: 1); Citrullinated MCV polypeptide: PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP (SEQ ID NO: 2); Citrullinated caspase14 peptide: Cit-GEQ-Cit-DPGETVGGDE (SEQ ID NO: 3); Citrullinated PCOLCE polypeptide: KTLP-Cit-GTAKEGQGPGPK-Cit-G (SEQ ID NO: 4); The combined diagnostic product can be an ELISA kit containing the detection antigens SMCP and CCP (the two are packaged separately), or it can be a separate anti-SMCP antibody ELISA kit and anti-CCP antibody ELISA kit.

[0012] The SMCP is used to detect the content of anti-SMCP antibodies in a patient's biological sample, and the CCP is used to detect the content of anti-CCP antibodies in a patient's biological sample.

[0013] Optionally or preferably, the biological sample is blood, plasma, serum or joint fluid.

[0014] Optionally or preferably, the citrullinated SR-A polypeptide, citrullinated MCV polypeptide, citrullinated caspase14 polypeptide and citrullinated PCOLCE polypeptide are all connected to BSA (bovine serum albumin). Specifically, the carrier protein BSA is coupled to the N-terminus of the amino acid sequence through a cysteine ​​residue, and the amino acid sequence after coupling is as follows: Citrullinated SR-A polypeptide: BSA-C-KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit (SEQ ID NO: 5); Citrullinated MCV polypeptide: BSA-C-PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP (SEQ ID NO: 6); Citrullinated caspase14 peptide: BSA-C-Cit-GEQ-Cit-DPGETVGGDE (SEQ ID NO: 7); Citrullinated PCOLCE polypeptide: BSA-C-KTLP-Cit-GTAKEGQGPGPK-Cit-G (SEQ ID NO: 8).

[0015] The present invention has the following beneficial effects: The present invention can achieve accurate diagnosis of RA by performing a one-time detection of anti-SMCP antibodies in the patient's biological sample. In particular, the sensitivity and specificity of serum as a biological sample are the best, which are 70.19% and 93.54%, respectively, showing good diagnostic value. At the same time, in the serum samples of RA patients with negative anti-CCP antibodies, the positive rate of anti-SMCP antibodies is 60.00%, which also has good supplementary diagnostic value. In addition, the anti-SMCP antibody detection combined with the anti-CCP antibody detection (a diagnostic product prepared by combining SMCP and CCP) can increase the sensitivity of anti-CCP antibody diagnosis of RA by 14.72% while maintaining high specificity. The combined diagnostic sensitivity of the two reached 90.19% and the specificity reached 90.83%, which effectively improved the detection rate of RA and was more conducive to achieving the goal of early diagnosis and treatment and reducing the disability rate. BRIEF DESCRIPTION OF THE DRAWINGS

[0016] Figure 1 The statistical results of the diagnostic value of serum anti-SMCP antibodies for rheumatoid arthritis, where A is the statistical result of anti-SMCP antibody content detection, and B is the ROC curve; in the figure, * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001 (using Kruskal-Wallis test method). [AU value = (test serum OD value / positive serum OD value) * 100, OD value is the detection value of 450 nm of the microplate reader - 570 nm detection value].

[0017] Figure 2 The statistical results of the diagnostic value of plasma anti-SMCP antibodies for rheumatoid arthritis, where A is the statistical result of anti-SMCP antibody content detection, and B is the ROC curve; in the figure, * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001 (using Kruskal-Wallis test method). [AU value = (test plasma OD value / positive serum OD value) * 100, OD value is the detection value of 450 nm - 570 nm of the microplate reader].

[0018] Figure 3 The statistical results of the diagnostic value of anti-SMCP antibodies in synovial fluid for rheumatoid arthritis, where A is the statistical result of anti-SMCP antibody content detection, and B is the ROC curve; in the figure, * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001 (using the Mann-Whitney test method). [AU value = (OD value of synovial fluid to be tested / OD value of positive serum) * 100, OD value is the detection value of 450 nm - 570 nm of the microplate reader].

[0019] Figure 4 The figure is the relationship between the sensitivity and specificity of ELISA detection of different serum antibodies, which reflects the relationship between the sensitivity and specificity of serum anti-CCP antibody (green line) ELISA detection, serum anti-SMCP antibody (blue line) ELISA detection and the combined detection of the two (red line).

[0020] Figure 5 The figure is the relationship between the sensitivity and specificity of ELISA detection of different plasma antibodies, which reflects the relationship between the sensitivity and specificity of plasma anti-CCP antibody (green line) ELISA detection, plasma anti-SMCP antibody (blue line) ELISA detection and the combined detection of the two (red line). DETAILED DESCRIPTION

[0021] In order to make the technical personnel in this field better understand the present application scheme, the present application will be clearly and completely described below in conjunction with the embodiments and drawings. Obviously, the described embodiments are only embodiments of a part of the present application, rather than all embodiments. Based on the embodiments in the present application, all other embodiments obtained by ordinary technicians in this field without making creative work should belong to the scope of protection of this application. The instruments and reagents used in the embodiments are all derived from commercial channels unless otherwise specified.

[0022] Example 1 The citrullinated SR-A peptide, citrullinated MCV peptide, citrullinated caspase14 peptide and citrullinated PCOLCE peptide were synthesized and linked to BSA respectively to better coat the peptides on the ELISA plate (Shanghai Nuoyou Biotechnology Co., Ltd., Zhongpeptide Biochemical Co., Ltd.).

[0023] Citrullinated SR-A peptide: BSA-C-KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit; Citrullinated MCV peptide: BSA-C-PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP; Citrullinated caspase14 peptide: BSA-C-Cit-GEQ-Cit-DPGETVGGDE; Citrullinated PCOLCE peptide: BSA-C-KTLP-Cit-GTAKEGQGPGPK-Cit-G.

[0024] The citrullinated SR-A peptide, citrullinated MCV peptide, citrullinated caspase14 peptide and citrullinated PCOLCE peptide were mixed to obtain the SMCP citrullinated peptide mixture. The anti-SMCP antibodies in the serum, plasma and joint fluid of patients with rheumatoid arthritis, normal healthy people and patients with various rheumatic diseases were detected by enzyme-linked immunosorbent assay (ELISA).

[0025] In addition to the test serum, plasma, and synovial fluid samples, the main reagents used include: coated antigen bound to the ELISA plate (the above-mentioned SMCP citrullinated polypeptide mixture), detection antibody (i.e., enzyme-labeled secondary antibody, which specifically recognizes human IgG and binds to horseradish peroxidase), color substrate (i.e., TMB reagent, which develops color after being catalyzed by horseradish peroxidase, and the test results are presented according to the degree of substrate color development), and standard serum.

[0026] Experimental steps: (1) Plate coating: Dilute the MSCP citrullinated peptide mixture to 10 μg / ml in carbonate solution at 4°C and coat the plate with 100 μl / well at 4°C overnight. (2) Washing: Wash the plate three times with 0.05% Tween 20 PBS (PBS-T), 200 μl / well, 2 min / time; (3) Blocking: Add 3% BSA-PBST, 200 μl / well, and block at room temperature for 2 hours; (4) Washing: Wash the plate three times with 0.05% PBS-T, 200 μl / well, 2 min / time; (5) Primary antibody incubation: dilute the sample serum, plasma or joint fluid at 1:100 in 1% BSA-PBST and add to the 96-well plate in step (4), 100 μl / well, and incubate at 37°C for 1 hour; (6) Washing: Wash the plate 3 times with 0.05% PBS-T, 200 μl / well, 2 min / time; (7) Secondary antibody incubation: Add 100 μl of 1:13,000 diluted goat anti-human IgG (detection antibody) labeled with horseradish peroxidase, incubate at 37°C for 30 min, and wash the plate three times, 200 μl / well*2 min / time; (8) Color development: Add TMB color development solution (color development substrate), 100 μl / well, and react for 10-30 min in the dark; (9) Termination: Terminate the reaction with 2 mol / L sulfuric acid, 100 μl / well, and read the OD values ​​at 450 nm and 570 nm, respectively.

[0027] (10) Calculation results: 1) Select positive control standard serum: select 10 RA sera with OD values ​​greater than 1.5 and mix them in equal volumes; 2) AU value (Arbitrary units), the calculation formula is: AU value = (OD value of the sample to be tested / OD value of the positive serum) * 100, OD value is the detection value of the microplate reader at 450 nm - 570 nm.

[0028] Experimental results: (1) Diagnostic value of anti-SMCP antibodies in RA 1) Serum samples: A total of 265 RA patients (200 anti-CCP antibody positive and 65 anti-CCP antibody negative) were selected, as well as 100 disease controls: systemic lupus erythematosus (SLE), Sjögren's syndrome (SS), osteoarthritis (OA), and 180 healthy controls (HC). Peripheral blood was obtained and serum was separated. Anti-SMCP antibody ELISA was performed according to the above experimental steps to analyze the sensitivity and specificity of ELISA for the diagnosis of RA.

[0029] The results are as follows Figure 1 As shown in the figure, Figure A is the statistical result of the anti-SMCP antibody content detection of peripheral blood serum of different populations using anti-SMCP antibody ELISA method. The results show that the anti-SMCP antibody in RA patients is significantly higher than that in normal people and related disease control groups, and it is statistically significant. Figure B is the ROC curve of serum anti-SMCP antibody ELISA detection for RA patients, which reflects the relationship between the sensitivity and specificity of ELISA diagnosis. The area under the ROC curve (AUC) is 0.9082.

[0030] 2) Plasma samples: A total of 132 RA patients (100 anti-CCP antibody positive and 32 anti-CCP antibody negative), 50 disease controls of systemic lupus erythematosus (SLE), Sjögren's syndrome (SS), and osteoarthritis (OA), and 90 healthy controls (HC) were selected. Their peripheral blood was obtained and plasma was separated. Anti-SMCP antibody ELISA was performed to analyze the sensitivity and specificity of ELISA for the diagnosis of RA.

[0031] The results are as follows Figure 2 As shown in the figure, Figure A is the statistical result of the detection of anti-SMCP antibody content in peripheral blood plasma of different populations by anti-SMCP antibody ELISA method. The results show that the anti-SMCP antibody in RA patients is significantly higher than that in normal people and the control group of related diseases, and it is statistically significant. Figure B is the ROC curve of plasma anti-SMCP antibody ELISA detection for RA patients, which reflects the relationship between the sensitivity and specificity of ELISA diagnosis, and the AUC is 0.8940.

[0032] 3) Joint fluid sample: Thirty-two RA patients and 15 osteoarthritis (OA) disease controls were selected. Their joint fluid was obtained and tested for anti-SMCP antibody ELISA. The sensitivity and specificity of ELISA for RA diagnosis were analyzed.

[0033] The results are as follows Figure 3As shown in the figure, Figure A is the statistical result of anti-SMCP antibody content detection in synovial fluid of different populations using anti-SMCP antibody ELISA method. The results show that anti-SMCP antibodies in RA patients are significantly higher than those in the disease control group, and are statistically significant. Figure B is the ROC curve of synovial fluid anti-SMCP antibody ELISA detection for RA patients, reflecting the relationship between the sensitivity and specificity of ELISA diagnosis, and the AUC is 0.8604.

[0034] Anti-SMCP antibodies are significantly increased in the serum, plasma and joint fluid of RA patients. This detection technology has significant significance for the diagnosis of RA. The sensitivity and specificity of anti-SMCP antibody ELISA for RA diagnosis were further analyzed. The results are shown in Table 1: Table 1. Analysis of the diagnostic value of anti-SMCP antibody ELISA test for RA The results showed that the sensitivity of serum anti-SMCP antibody ELISA for diagnosing RA was 70.19% and the specificity was 93.54%; the sensitivity of plasma anti-SMCP antibody ELISA for diagnosing RA was 65.91% and the specificity was 96.25%; the sensitivity of synovial fluid anti-SMCP antibody ELISA for diagnosing RA was 62.50% and the specificity was 93.33%. Therefore, while ensuring high specificity, anti-SMCP antibody ELISA can achieve a sensitivity comparable to that of anti-CCP antibody, showing good diagnostic value.

[0035] (2) Diagnostic value of anti-SMCP antibodies in RA with negative anti-CCP antibodies Some RA patients are negative for RA-specific antibodies (anti-CCP antibodies), which makes diagnosis difficult for these patients. Therefore, we analyzed the supplementary diagnostic value of anti-SMCP antibodies in RA patients with negative anti-CCP antibodies, as shown in Table 2.

[0036] Table 2. Analysis of the diagnostic value of anti-SMCP antibody ELISA test for RA with negative anti-CCP antibody In the serum samples of RA patients with negative anti-CCP antibodies, the positive rate of anti-SMCP antibodies was 60.00%; in the plasma samples of RA patients with negative anti-CCP antibodies, the positive rate of anti-SMCP antibodies was 50.00%. In summary, anti-SMCP antibodies have a good supplementary diagnostic value in RA patients with negative anti-CCP antibodies.

[0037] (3) The combined diagnostic value of anti-SMCP antibodies and anti-CCP antibodies Considering that anti-SMCP antibodies have good supplementary diagnostic value in anti-CCP antibody-negative RA patients, we conducted a combined diagnostic analysis of anti-SMCP antibody ELISA detection and anti-CCP antibody ELISA detection. The anti-CCP antibody ELISA detection used a commercially available anti-CCP3 antibody ELISA kit (human anti-CCP3 antibody ELISA kit, Inova Diagnostics, USA). The results are shown in Table 3.

[0038] Table 3. Analysis of the combined diagnostic value of anti-SMCP antibodies and anti-CCP antibodies The sensitivity and specificity of anti-SMCP antibodies in serum samples were 70.19% and 93.54%, respectively, and the sensitivity and specificity of anti-CCP antibodies were 75.47% and 97.08%, respectively. The sensitivity and specificity of anti-SMCP antibodies combined with anti-CCP antibodies in diagnosing RA were 90.19% and 90.83%, respectively, which can increase the sensitivity of anti-CCP antibodies in diagnosing RA by 14.72% while maintaining high specificity. Figure 4 As shown in the figure, the AUCs of anti-SMCP antibodies and anti-CCP antibodies were 0.9082 and 0.8970, respectively. The combined diagnosis AUC of anti-SMCP antibodies and anti-CCP antibodies can be increased to 0.9561.

[0039] The sensitivity and specificity of anti-SMCP antibodies in plasma samples were 65.91% and 96.25%, respectively, and the sensitivity and specificity of anti-CCP antibodies were 75.76% and 98.75%, respectively. The sensitivity and specificity of anti-SMCP antibodies combined with anti-CCP antibodies in diagnosing RA were 87.88% and 95.42%, respectively, which can increase the sensitivity of anti-CCP antibodies in diagnosing RA by 12.12% while maintaining high specificity. Figure 5 As shown in the figure, the AUCs of anti-SMCP antibodies and anti-CCP antibodies were 0.8940 and 0.9021, respectively. The combined diagnosis AUC of anti-SMCP antibodies and anti-CCP antibodies can be increased to 0.9589.

[0040] The invention concept is described in detail using specific examples herein, and the description of the above embodiments is only used to help understand the core idea of ​​the invention. It should be pointed out that any obvious modification, equivalent substitution or other improvement made by a person of ordinary skill in the art without departing from the invention concept should be included in the protection scope of the present invention.

Claims

1. Application of SMCP in the preparation of a diagnostic product for rheumatoid arthritis, characterized in that: The SMCP is a polypeptide mixture consisting of citrullinated SR-A polypeptide, citrullinated MCV polypeptide, citrullinated caspase14 polypeptide and citrullinated PCOLCE polypeptide, and the amino acid sequence is as follows: Citrullinated SR-A peptide: KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit; Citrullinated MCV peptide: PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP; Citrullinated caspase14 peptide: Cit-GEQ-Cit-DPGETVGGDE; Citrullinated PCOLCE peptide: KTLP-Cit-GTAKEGQGPGPK-Cit-G.

2. The use according to claim 1, characterized in that: The diagnostic product is used to detect the content of anti-SMCP antibodies. Compared with normal healthy people, patients with systemic lupus erythematosus, Sjögren's syndrome and osteoarthritis, the content of anti-SMCP antibodies in biological samples of rheumatoid arthritis patients is significantly increased.

3. The use according to claim 2, characterized in that: The biological sample is blood, plasma, serum or joint fluid.

4. The use according to claim 1, characterized in that: The N-terminal of the amino acid sequence of the citrullinated SR-A polypeptide, the citrullinated MCV polypeptide, the citrullinated caspase14 polypeptide and the citrullinated PCOLCE polypeptide is also coupled to the carrier protein BSA via a cysteine ​​residue, and the amino acid sequence after coupling is as follows: Citrullinated SR-A peptide: BSA-C-KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit; Citrullinated MCV peptide: BSA-C-PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP; Citrullinated caspase14 peptide: BSA-C-Cit-GEQ-Cit-DPGETVGGDE; Citrullinated PCOLCE peptide: BSA-C-KTLP-Cit-GTAKEGQGPGPK-Cit-G.

5. The use according to claim 1, characterized in that: The diagnostic product is an ELISA detection kit.

6. Use of SMCP combined with CCP in the preparation of a diagnostic product for rheumatoid arthritis, characterized in that: The SMCP is a polypeptide mixture consisting of citrullinated SR-A polypeptide, citrullinated MCV polypeptide, citrullinated caspase14 polypeptide and citrullinated PCOLCE polypeptide, and the amino acid sequence is as follows: Citrullinated SR-A peptide: KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit; Citrullinated MCV peptide: PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP; Citrullinated caspase14 peptide: Cit-GEQ-Cit-DPGETVGGDE; Citrullinated PCOLCE peptide: KTLP-Cit-GTAKEGQGPGPK-Cit-G; The CCP is a cyclized citrullinated polypeptide.

7. The use according to claim 6, characterized in that: The SMCP is used to detect the content of anti-SMCP antibodies in a patient's biological sample, and the CCP is used to detect the content of anti-CCP antibodies in a patient's biological sample.

8. The use according to claim 7, characterized in that: The biological sample is blood, plasma, serum or joint fluid.

9. The use according to claim 6, characterized in that: The N-terminal of the amino acid sequence of the citrullinated SR-A polypeptide, the citrullinated MCV polypeptide, the citrullinated caspase14 polypeptide and the citrullinated PCOLCE polypeptide is also coupled to the carrier protein BSA via a cysteine ​​residue, and the amino acid sequence after coupling is as follows: Citrullinated SR-A peptide: BSA-C-KGD-Cit-GAIGFPGS-Cit-GLPGYAG-Cit; Citrullinated MCV peptide: BSA-C-PG-Cit-VYAT-Cit-SSAV-Cit-L-Cit-SSVP; Citrullinated caspase14 peptide: BSA-C-Cit-GEQ-Cit-DPGETVGGDE; Citrullinated PCOLCE peptide: BSA-C-KTLP-Cit-GTAKEGQGPGPK-Cit-G.

Images