Enzyme immunoassay for the qualitative and quantitative determination of IgG antibodies against U1-RNP in human serum.
Contents of Kit
Microtiter Plate: 1 x 12 x 8 Enzyme Conjugate IgG: 1 x 14 mL Calibrator A - F: 1 x 6 x 2 mL Positive Control: 1 x 2 mL Negative Control: 1 x 2 mL Sample Diluent: 1 x 100 mL TMB Substrate Solution: 1 x 14 mL Wash Buffer, Concentrate (10x): 1 x 100 mL TMB Stop Solution: 1 x 14 mL
Storage
2-8°C
Precision
Detection Range
0 - 60 U/mL, cut-off 4 U/mL
Sensitivity
0.5 U/mL
General Description
Enzyme immunoassay for the qualitative and quantitative determination of IgG antibodies against U1-RNP in human serum. Uracil-rich, small nuclear RNAs (U-snRNA) are physiologically complexed with proteins to ribonucleoprotein particles (U-snRNP) which are involved in the splicing process of pre-mRNA. Different complexes (U1-U6) can be distinguished according to the participating RNA; their protein components are in part common, in part different. For example, U1-snRNA is complexed with the so-called Sm-proteins which are also present in other U-snRNPs, and with the U1-specific proteins A, C and 68 kDa. Autoantibodies directed against these three U1-specific proteins are considered as diagnostic marker for mixed connective tissue disease (MCTD). They are also associated with systemic lupus erythematosus (SLE) but occur with distinctly lower prevalence (about 35%) and mostly in combination with further antinuclear antibodies (ANA). Nowadays, proteins A, C and 68 kDa are regarded as the actual RNP antigen, rather than the complete U1-RNP.
Standard Curve
Citations
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Background
U1-RNP is a nuclear protein-RNA complex that plays an important role in mRNA processing. It consists of U1-RNA, which contains part of the double-stranded secondary structure, and proteins specific for the U1-RNP complex, including U1-A, U1-C, and U1-70kDa. Other proteins may also associate with U1-RNP, such as Sm, but these are not specific to the U1-RNP complex. Autoantibodies are a hallmark of many autoimmune diseases, and researchers have found that high titers of U1-RNP antibodies are associated with mixed connective tissue disease (MCTD), and also occur in systemic lupus erythematosus (SLE) and systemic sclerosis (SSc).
The U1-RNP antibody targets the 70kDa, A, and C proteins that are core components of the U-series small nuclear RNAs involved in pre-messenger RNA splicing. One of the major determinants of the U1-RNP antibody response is U1-70kDa, which is typically produced early in the U1-RNP antibody response and facilitates the production of antibodies against other proteins in the U1-RNP complex through epitope diffusion. One study showed that U1-RNP antibodies were found in 26% of SLE patients before the onset of the disease, and the antibodies frequently appeared within 1 year of the onset of the disease. The type of anti-RNP antibody also varies with the connective tissue disease phenotype; anti-U1-RNP antibodies are predominantly IgM in SLE patients, whereas IgG anti-U1-RNP antibodies are more common in MCTD. This variation in isoform subclasses may be the result of pathogenic mechanisms or may determine the role of antibodies in disease pathogenesis. Apoptosis can lead to changes in self-antigens, which may increase or decrease the affinity of antibodies. Studies have reported that apoptosis results in the formation of different forms of U1-70kDa protein. Recognition of modified U1-RNP (especially U1-70kDa) appears to be associated with specific clinical features. Antibodies to apoptotic U1-70kDa have been shown to be associated with lupus skin disease.
Anti-U1-RNP antibodies are now considered a mandatory laboratory marker in all cases of MCTD, and their sensitivity for diagnosing MCTD is high, approaching 100%. While the specificity depends on the test object, it is about 60% when anti-70kDa U1-RNP antibodies are included, and 20% and 13% when only A and C proteins are tested, respectively.
Figure 1. Laboratory tests for immune differential diagnosis (Source: Dima A, et al. 2018)
References
1. Dima A, et al. The impact of anti-U1-RNP positivity: systemic lupus erythematosus versus mixed connective tissue disease. Rheumatol Int. 2018 Jul;38(7):1169-1178.
2. Stochmal A, et al. Antinuclear Antibodies in Systemic Sclerosis: an Update. Clin Rev Allergy Immunol. 2020 Feb;58(1):40-51.
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References
The diagnostic challenge of patients with anti-U1-RNP antibodies
Rheumatol Int
Authors: Elhani I, Khoy K, Mariotte D, Comby E, Marcelli C, Le Mauff B, Audemard-Verger A, Boutemy J, Maigné G, Martin Silva N, Aouba A, de Boysson H.
Anti-U1-RNP antibodies are necessary for the diagnosis of mixed connective tissue disease (MCTD), but they are also prevalent in other connective tissue diseases, especially systemic lupus erythematosus (SLE), from which distinction remains challenging. We aimed to describe the presentation and outcome of patients with anti-U1-RNP antibodies and to identify factors to distinguish MCTD from SLE. We retrospectively applied the criteria sets for MCTD, SLE, systemic sclerosis (SSc) and rheumatoid arthritis (RA) to all patients displaying anti-U1-RNP antibodies in the hospital of Caen from 2000 to 2020. Thirty-six patients were included in the analysis. Eighteen patients (50%) satisfied at least one of the MCTD classifications, 11 of whom (61%) also met 2019 ACR/EULAR criteria for SLE. Twelve other patients only met SLE without MCTD criteria, and a total of 23 patients (64%) met SLE criteria. The most frequent manifestations included Raynaud's phenomenon (RP, 91%) and arthralgia (67%). We compared the characteristics of patients meeting only the MCTD (n = 7), SLE (n = 12), or both (n = 11) criteria. Patients meeting the MCTD criteria were more likely to display SSc features, including sclerodactyly (p < 0.01), swollen hands (p < 0.01), RP (p = 0.04) and esophageal reflux (p < 0.01). The presence of scleroderma features (swollen hands, sclerodactyly, gastro-oesophageal reflux), was significantly associated with the diagnosis of MCTD. Conversely, the absence of those manifestations suggested the diagnosis of another definite connective tissue disease, especially SLE.
Doubtful Clinical Value of Subtyping Anti-U1-RNP Antibodies Regarding the RNP-70 kDa Antigen in Sera of Patients with Systemic Lupus Erythematosus
Int J Mol Sci
Authors: Ahmad A, Brylid A, Dahle C, Saleh M, Dahlström Ö, Enocsson H, Sjöwall C.
The detection of antinuclear antibodies is central to the diagnosis and prognosis of systemic lupus erythematosus (SLE), primary Sj?gren's syndrome (pSS) and mixed connective tissue disease (MCTD). Anti-U1-RNP and anti-RNP70 antibodies were assayed in the sera of patients with SLE (n = 114), pSS (n = 54) and MCTD (n = 12). In the SLE group, 34/114 (30%) were anti-U1-RNP positive, and 21/114 (18%) were both anti-RNP70 positive and anti-U1-RNP positive. In the MCTD group, 10/12 (83%) were anti-U1-RNP positive, and 9/12 (75%) were anti-RNP70 positive. Only one individual with pSS was antibody positive (for both anti-U1-RNP and anti-RNP70). All anti-RNP70-positive samples were also anti-U1-RNP positive. Anti-U1-RNP-positive subjects with SLE were younger (p < 0.0001); showed lower concentrations of complement protein 3 (p = 0.03); had lower eosinophil (p = 0.0005), lymphocyte (p = 0.006) and monocyte (p = 0.03) counts; and had accrued less organ damage (p = 0.006) than the anti-U1-RNP-negative SLE patients. However, we observed no significant clinical or laboratory parameter differences between the anti-U1-RNP-positive individuals with/without anti-RNP70 in the SLE group. In conclusion, anti-RNP70 antibodies are not exclusive to MCTD but are rarely detected in pSS and healthy individuals. In SLE, anti-U1-RNP antibodies are associated with a clinical phenotype that resembles MCTD, with hematologic involvement and less damage accrual. Based on our results, the clinical value of subtyping anti-RNP70 in anti-U1-RNP-positive sera appears to be of limited value.
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Figure 1. Laboratory tests for immune differential diagnosis
