Clinical Focus
Systemic Sclerosis
Laboratory Markers for Diagnosis and Prognosis
Individuals suitable for testing
Clinical background [return to contents]
Systemic sclerosis (SSc) is a chronic, multisystem, heterogeneous autoimmune disease. Individuals with SSc have a mortality rate approximately 2.8 times that of the general population.1 In the United States, the incidence is approximately 15 cases per 100,000 person-years.2 The disease is characterized by inflammation, vasculopathy, and progressive fibrosis of the skin and internal organs.3 SSc is frequently referred to as scleroderma; however, scleroderma includes SSc and localized forms of scleroderma that affect only the skin.4
The 2 main types of SSc are defined according to the pattern of skin involvement: limited cutaneous SSc (lcSSc) and diffuse cutaneous SSc (dcSSc). In lcSSc, skin thickening is present distal to the elbows and knees, and facial skin thickening may or may not be present. In contrast, dcSSc is characterized by thickening of the skin of the whole extremity, as well as that of the anterior chest, abdomen, and back, with or without facial skin involvement.4,5 Multiple organs, including the heart, lungs, gastrointestinal tract, and kidneys, can be affected in both forms, though organ involvement is generally less severe in lcSSc.4,5 CREST syndrome (calcinosis, Raynaud phenomenon, esophageal dysmotility, sclerodactyly, and telangiectasia) occurs frequently in lcSSC but can also occur in longstanding dcSSc.4
Of patients with SSc, approximately 55% have lcSSc and 35% have dcSSc.4 The remaining 10% of patients with SSc have sine scleroderma or an overlap syndrome.4 Sine scleroderma is a form of SSc that has characteristic clinical features but spares the skin.5 Overlap syndromes occur when features of another autoimmune disease are present along with SSc.5
Early treatment of SSc can improve outcomes, so prompt diagnosis is important. However, diagnosis can be challenging because many patients present with nonspecific symptoms such as Raynaud phenomenon, gastroesophageal reflux, puffy fingers, and fatigue.5 In addition, patients with other autoimmune disorders may present with symptoms suggestive of SSc. Testing for autoantibodies that are associated with SSc assists diagnosis and can help predict organ involvement and severity of disease.
This Clinical Focus discusses the role of autoantibody testing in the diagnosis and categorization of SSc, as well as in the assessment of prognosis. The material is provided for educational purposes only and is not intended as medical advice. Test selection and interpretation, diagnosis, and patient management decisions should be based on the physician’s education, specialization, clinical expertise, and clinical and laboratory assessment of the patient.
Individuals suitable for testing [return to contents]
- Individuals with suspected SSc (ie, those with capillary nailfold changes, Raynaud phenomenon, and/or characteristic skin changes)
- Individuals with unexplained, but suggestive, musculoskeletal, gastrointestinal, cardiac, or pulmonary symptoms
Test availability [return to contents]
Tests available for the diagnosis of SSc and assessment of prognosis are shown in Table 1.
Table 1. Autoantibody Testing Used in the Diagnosis and Prognosis of Systemic Sclerosis [return to contents]
Test code |
Test namea |
Clinical use |
94954b,c |
ANA, IFA, Cascade and Rheumatoid Arthritis Panel 2, With Reflexes Includes ANA screen, IFA, with Reflex to Titer and Pattern and Reflex to Multiplex 11 Antibody Cascade (16814); RF (4418); CCP antibody (IgG) (11173); mutated citrullinated vimentin (MCV) antibody (13238). |
Diagnose autoimmune disease |
36378b |
ANAlyzeR™ ANA, IFA with Reflex Titer/Pattern, Systemic Autoimmune Panel 1 Includes ANA screen, IFA, with Reflex to Titer and Pattern (249); beta-2-glycoprotein I antibodies (IgG, IgA, IgM) (30340); cardiolipin antibodies (IgA, IgG, IgM) (7352); CCP antibody (IgG) (11173); centromere B antibody (16088); chromatin (nucleosomal) antibody (34088); complement component C3c and C4c (5704); DNA (ds) antibody, crithidia IFA with reflex to titer (37092); Jo-1 antibody (5810); mutated citrullinated vimentin (MCV) antibody (13238); scleroderma antibody (Scl-70) (4942); Sjögren antibodies (SS-A and SS-B) (7832); Sm antibody (37923); Sm/RNP antibody (38567); RF (IgG, IgA, IgM) (19705); thyroid peroxidase antibodies (TPO) (5081). |
|
| 249b | ANA Screen, IFA, with Reflex to Titer and Pattern | |
16814b |
ANA Screen, IFA, with Reflex to Titer and Pattern and Reflex to Multiplex 11 Antibody Cascade Includes ANA screen, IFA, with Reflex to Titer and Pattern (249) and reflex to dsDNA (255), Sm/RNP (38567), RNP (19887), Sm (37923), and chromatin antibodies (34088); if all 5 antibodies are negative, reflex to SS-A (38568), SS-B (38569), Scl-70 (4942), and Jo-1 antibodies (5810); if all 4 of these antibodies are negative, reflex to ribosomal P (34283) and centromere B antibodies (16088). |
|
90073b |
ANA Screen, IFA, with Reflex to Titer and Pattern/Systemic Sclerosis Panel 1 Includes ANA screen, IFA, with Reflex to Titer and Pattern (249) and reflex to centromere B antibody (16088); RNA polymerase III (19899); scleroderma antibody (Scl-70) (4942). |
Diagnose SSc |
16088 |
Centromere B Antibody |
Diagnose SSc; determine prognosis |
19899 |
RNA Polymerase III Antibody |
|
4942 |
Scleroderma Antibody (Scl-70) |
|
38567d |
Sm/RNP Antibody |
Determine prognosis of SSc |
94685e |
Systemic Sclerosis (Scleroderma) 12 Antibodies Panel 2 Includes centromere protein (CENP)-A, CENP-B, fibrillarin (U3-RNP), PM/Scl-75, PM/Scl-100, RP11 (RNA polymerase III), RP155 (RNA polymerase III), Scl-70, Th/To, U1-snRNP RNP-70kd, U1-snRNP RNP A, and U1-snRNP RNP C antibodies. |
Diagnose SSc; determine prognosis |
| ANA, antinuclear antibodies; IFA, immunofluorescence assay; SSc, systemic sclerosis. | |
| a | Numbers in parentheses represent test codes for individual components. |
| b | Reflex tests are performed at an additional charge and are associated with an additional CPT code. |
| c | This test was developed and its analytical performance characteristics have been determined by Quest Diagnostics. It has not been cleared or approved by the US Food and Drug Administration. This assay has been validated pursuant to the CLIA regulations and is used for clinical purposes. |
| d | Sm/RNP is also called U1-snRNP and U1-RNP. |
| e | This panel is run as a single line blot; thus, not all components are available as individual tests. |
Test selection and interpretation [return to contents]
Diagnosis and prognosis
Early diagnosis of SSc is important to delay, mitigate, or avoid irreversible end-organ damage. The diagnosis is based primarily on the presence of characteristic clinical findings and autoantibodies. Classification criteria are available from the American College of Rheumatology-European League Against Rheumatism (ACR-EULAR) and have 91% sensitivity and 92% specificity for SSc (Table 2).6 Autoantibody positivity may be present early in the course of the disease and assist in diagnosis of patients with nonspecific signs and symptoms. Autoantibody patterns can also help distinguish the various types of SSc (ie, lcSSc, dcSSc, sine scleroderma, overlap syndrome) and the organ(s) likely to be involved, which is important for determining prognosis and optimal treatment.7,8
Table 2. ACR-EULAR Systemic Sclerosis Classification Criteria6 [return to contents]
Classify a patient as having systemic sclerosis if the sum of points for criteria below is ≥9. |
|
Criterion |
Points |
Skin thickened on fingers of both hands, extending proximal to the metacarpophalangeal joints |
9 |
Skin on fingers thickened (only count highest score) Puffy fingers Sclerodactylya |
2 4 |
Lesions on fingertips (only count highest score) Ulcers on tip of digits Pitting scars on fingertips |
2 3 |
Telangiectasia |
2 |
Abnormal nailfold capillaries |
2 |
Pulmonary arterial hypertension and/or interstitial lung disease (max score is 2) Pulmonary arterial hypertension Interstitial lung disease |
2 2 |
Raynaud phenomenon |
3 |
Presence of 1 or more of the followingb: Centromere antibody Scl-70 antibody RNA polymerase III antibody |
3 |
| ACR-EULAR, American College of Rheumatology-European League Against Rheumatism. | |
| a | Distal to metacarpophalangeal joints but proximal to proximal interphalangeal joints. |
| b | Maximum score is 3. |
The following sections describe the autoantibodies associated with SSc and their relevance for diagnosis and prognosis. A summary of the autoantibodies, their prevalences, and their clinical associations is provided in Table 3. An algorithm showing the use of antibody testing for SSc diagnosis is presented in the Figure.
Table 3. Clinical Characteristics Associated With Autoantibodies Used in the Diagnosis and Classification of Systemic Sclerosisa [return to contents]
Antibody |
Prevalence in SSc, % |
Presence in other autoimmune diseases |
SSc type most likely |
Clinical associations |
Relative SSc prognosis |
ANA9 |
85-95 |
Common |
Does not help differentiate type |
Most common in rheumatic diseases |
Varies |
Centromere A or B7,8,10 |
20-40b |
Uncommon |
lcSSc |
PAH, calcinosis, digital ischemia, intestinal involvement |
Better |
PM/Scl-75 or PM/Scl-1007,8,10-12 |
2-11 |
Common |
SSc-myositis overlap syndrome or lcSSc |
Muscle and joint involvement, calcinosis; GI and lung involvement for those with PM/Scl-75 |
Better |
RNA Polymerase III7,8,10,13,14 |
7-41 |
Rare |
dcSSc |
Renal crisis, progressive skin thickening, GAVE, malignancy |
Poorer |
Scl-707,8,15 |
9.4-71b |
Rare |
dcSSc |
Severe pulmonary fibrosis, cardiac involvement, renal crisis, digital ulcers |
Poorer |
Th/To7,8,10 |
2-10 |
Rare |
lcSSc |
Pulmonary involvement |
Poorer |
U1-snRNP7,10,16 |
2-15 |
Common (90% in patients with MCTD) |
lcSSc |
PAH, arthritis, esophageal dysfunction |
Better |
U3-RNP (Fibrillarin)7,8,10 |
3-20b |
Rare |
dcSSc |
Multiorgan involvement, PAH |
Poorer |
| ANA, antinuclear antibodies; dcSSc, diffuse cutaneous systemic sclerosis; GAVE, gastric antral vascular ectasia; GI, gastrointestinal; lcSSc, limited cutaneous systemic sclerosis; MCTD, mixed connective tissue disease; PAH, pulmonary arterial hypertension; SSc, systemic sclerosis. | |
| a | Data presented in this table have been compiled from the cited references. It is important to note that autoantibody frequency in SSc as well as other autoimmune diseases varies with sex, ethnicity, and population studied.17 |
| b | In the United States, prevalence has been shown to vary across ethnicities.18 |
Antinuclear antibody
Antinuclear antibody (ANA) immunofluorescence assay (IFA) is the initial test for detecting autoantibodies in a patient presenting with nonspecific symptoms, including Raynaud phenomenon. ANA is present in 85% to 95% of patients with SSc; however, ANA is also present in many other autoimmune disorders.9,19 Thus, a positive ANA IFA test should be followed with testing for more specific antibodies such as centromere B, Scl-70 (topoisomerase I), and RNA polymerase III antibodies.20 A negative ANA IFA test decreases the likelihood of SSc but does not rule it out.
Centromere antibodies
Centromere antibodies can be directed against a number of centromere proteins (CENP), including CENP-A, -B, and -C, though CENP-B is thought to be the main target.7,21 These antibodies are found in 20% to 40% of patients with SSc and are associated with the lcSSc subtype.7 Furthermore, centromere antibodies are included in the 2013 ACR-EULAR classification criteria.6 However, these antibodies can occur in other connective tissue diseases, including systemic lupus erythematosus (SLE), primary biliary cholangitis, rheumatoid arthritis, and Sjögren syndrome.7
Presence of centromere antibodies is considered predictive of SSc development in patients with Raynaud phenomenon.21 The antibodies are associated with lower risk of cardiac and renal involvement but higher risk of PAH.7,21 For patients without PAH, centromere antibodies are associated with a better prognosis overall than are other autoantibodies.21
PM/Scl antibodies
PM/Scl antibodies target the PM/Scl exosome complex, and most reactivity is against 2 proteins, PM/Scl-75 and PM/Scl-100. They are present in 2% to 11% of SSc patients and are associated with SSc-myositis overlap syndrome and lcSSc.7,8 PM/Scl antibodies also occur in other autoimmune diseases such as polymyositis and dermatomyositis.
The presence of either PM/Scl-75 or PM/Scl-100 antibody is associated with calcinosis, and the co-occurrence of both antibodies is associated with inflammatory myositis.11,12 However, each antibody may be associated with a distinct clinical phenotype; gastrointestinal symptoms and ILD are common in patients with PM/Scl-75 antibodies but less so in those with PM/Scl-100 antibodies.11,12 Internal organ involvement generally remains mild in patients with any PM/Scl antibodies, and the presence of these antibodies is associated with an overall favorable prognosis.7
RNA polymerase III antibodies
RNA polymerase III antibodies target RNA polymerase epitopes 11 and 155 and are thus also known as anti-RP11 and anti-RP155. These antibodies are found in 7% to 41% of patients with SSc and occur most often in dcSSc.8,14 They are diagnostic for SSc, as they are rarely found in other autoimmune diseases, and are included in the 2013 ACR-EULAR classification criteria.6
The presence of RNA polymerase III antibodies is associated with progressive skin thickening, gastric antral vascular ectasia (GAVE), and renal crisis.7,8 In addition, these antibodies are associated with onset of cancer within a 2-year timeframe before or after onset of SSc skin changes.13 Historically, RNA polymerase III antibodies indicated a poor prognosis, but mortality rates improved after the introduction of ACE inhibitors to treat renal crisis; the prognosis for patients with RNA polymerase III antibodies is now better than for those with Scl-70 or U3-RNP antibodies.8
Scl-70 (topoisomerase I antibody)
Topoisomerase I antibodies were initially named Scl-70 based on immunoblot detection of a 70-kDa protein.21 The prevalence of Scl-70 antibodies in SSc varies widely across geographies and ethnicities, ranging from 9% to 71%.7 These antibodies are strongly associated with dcSSc but also occur in lcSSc.7,8 Furthermore, they are highly specific to SSc and are included in the 2013 ACR-EULAR classification criteria.6,7
The presence of Scl-70 antibodies is associated with pulmonary and cardiac involvement as well as renal crisis and vascular complications.7,8 Therefore, these antibodies indicate a poor prognosis.15 The increased mortality associated with Scl-70 antibodies results primarily from the higher risk for severe ILD.7,8
Th/To antibodies
Th/To antibodies target RNase P and mitochondrial RNase ribonucleoprotein complexes. These antibodies are found in 2% to 5% of SSc patients and are primarily associated with lcSSc.7,8 Th/To antibodies are rarely found in other autoimmune diseases but can occur in patients with localized scleroderma.8
Patients with Th/To antibodies often develop both ILD and PAH and thus have a poorer prognosis than other patients with lcSSc.7,8
U1-snRNP (Sm/RNP) antibodies
U1-snRNP antibodies, also referred to as U1-RNP and Smith (Sm)/RNP, target 3 components of the U1 small nuclear ribonucleoprotein complex: U1-snRNP RNP A, U1-snRNP RNP C, and U1-snRNP RNP-70kd. These antibodies, found in 2% to 14% of SSc patients, are more frequent in lcSSc than in dcSSc.16 The antibodies are also found in patients with other autoimmune diseases, including approximately 90% of patients with mixed connective tissue disease.7,16
Patients with U1-snRNP antibodies tend to have less prominent skin thickness and less renal involvement, but have increased risk of PAH, arthritis, and esophageal dysfunction.8,16 Overall, the presence of U1-snRNP antibodies is associated with a good response to corticosteroids and a favorable prognosis.16
U3-RNP (fibrillarin) antibodies
U3-RNP antibodies target the U3 small nucleolar ribonucleoprotein (U3-RNP) complex, which consists of the protein fibrillarin and U3 RNA.7 These antibodies are found in about 4% to 10% of patients with SSc, and are especially common in African American SSc patients (approximately 30%).7,8 U3-RNP antibodies are rarely found in patients with other autoimmune disorders; thus, the presence of these antibodies supports a SSc diagnosis.8,21 These antibodies occur most often in dcSSc, but they can also occur in lcSSc.7,8
U3-RNP antibodies are associated with multiorgan involvement, including the heart, kidneys, muscle, lungs, and gastrointestinal system. Their presence is an independent risk factor for the development of PAH, and PAH is the most common cause of death in U3-RNP–positive patients.7,8 Therefore, their presence indicates a poorer prognosis.
Supportive testing
Once a diagnosis of SSc is established, and clinical findings and autoantibody testing have established the type, further laboratory testing is performed to support evaluation of end-organ function and overall health. Testing may include examination of liver and kidney function, complete blood count, and inflammation assessment.22 Since SSc is a chronic disease, follow-up assessment at least annually is recommended.22
References [return to contents]
- Lee YH. Overall and sex- and disease subtype-specific mortality in patients with systemic sclerosis: an updated meta-analysis. Z Rheumatol. 2019;78(2):195-201. doi:10.1007/s00393-018-0492-8
- Fan Y, Bender S, Shi W, et al. Incidence and prevalence of systemic sclerosis and systemic sclerosis with interstitial lung disease in the United States. J Manag Care Spec Pharm. 2020;26(12):1539-1547. doi:10.18553/jmcp.2020.20136
- Lee JJ, Pope JE. Diagnosis and management of systemic sclerosis: a practical approach. Drugs. 2016;76(2):203-213. doi:10.1007/s40265-015-0491-x
- Young A, Khanna D. Systemic sclerosis: commonly asked questions by rheumatologists. J Clin Rheumatol. 2015;21(3):149-155. doi:10.1097/RHU.0000000000000232
- Denton CP, Khanna D. Systemic sclerosis. Lancet. 2017;390(10103):1685-1699. doi:10.1016/s0140-6736(17)30933-9
- van den Hoogen F, Khanna D, Fransen J, et al. 2013 classification criteria for systemic sclerosis: an American College of Rheumatology/European League against Rheumatism collaborative initiative. Arthritis Rheum. 2013;65(11):2737-2747. doi:10.1002/art.38098
- Stochmal A, Czuwara J, Trojanowska M, et al. Antinuclear antibodies in systemic sclerosis: an update. Clin Rev Allergy Immunol. 2020;58(1):40-51. doi:10.1007/s12016-018-8718-8
- Hamaguchi Y, Takehara K. Anti-nuclear autoantibodies in systemic sclerosis: news and perspectives. J Scleroderma Relat Disord. 2018;3(3):201-213. doi:10.1177/2397198318783930
- Mahler M, Meroni PL, Bossuyt X, et al. Current concepts and future directions for the assessment of autoantibodies to cellular antigens referred to as anti-nuclear antibodies. J Immunol Res. 2014;2014:315179. doi:10.1155/2014/315179
- Mahler M, Hudson M, Bentow C, et al. Autoantibodies to stratify systemic sclerosis patients into clinically actionable subsets. Autoimmun Rev. 2020;19(8):102583. doi:10.1016/j.autrev.2020.102583
- D'Aoust J, Hudson M, Tatibouet S, et al. Clinical and serologic correlates of anti-PM/Scl antibodies in systemic sclerosis: a multicenter study of 763 patients. Arthritis Rheumatol. 2014;66(6):1608-1615. doi:10.1002/art.38428
- Wodkowski M, Hudson M, Proudman S, et al. Clinical correlates of monospecific anti-PM75 and anti-PM100 antibodies in a tri-nation cohort of 1574 systemic sclerosis subjects. Autoimmunity. 2015;48(8):542-551. doi:10.3109/08916934.2015.1077231
- Shah AA, Hummers LK, Casciola-Rosen L, et al. Examination of autoantibody status and clinical features associated with cancer risk and cancer-associated scleroderma. Arthritis Rheumatol. 2015;67(4):1053-1061. doi:10.1002/art.39022
- Sobanski V, Dauchet L, Lefèvre G, et al. Prevalence of anti-RNA polymerase III antibodies in systemic sclerosis: new data from a French cohort and a systematic review and meta-analysis. Arthritis Rheumatol. 2014;66(2):407-417. doi:10.1002/art.38219
- Pokeerbux MR, Giovannelli J, Dauchet L, et al. Survival and prognosis factors in systemic sclerosis: data of a French multicenter cohort, systematic review, and meta-analysis of the literature. Arthritis Res Ther. 2019;21(1):86. doi:10.1186/s13075-019-1867-1
- Mehra S, Walker J, Patterson K, et al. Autoantibodies in systemic sclerosis. Autoimmun Rev. 2013;12(3):340-354. doi:10.1016/j.autrev.2012.05.011
- Krzyszczak ME, Li Y, Ross SJ, et al. Gender and ethnicity differences in the prevalence of scleroderma-related autoantibodies. Clin Rheumatol. 2011;30(10):1333-1339. doi:10.1007/s10067-011-1751-0
- Nandiwada SL, Peterson LK, Mayes MD, et al. Ethnic differences in autoantibody diversity and hierarchy: more clues from a US cohort of patients with systemic sclerosis. J Rheumatol. 2016;43(10):1816-1824. doi:10.3899/jrheum.160106
- de Almeida Brito F, Maria Eloi Santos S, Aparecida Ferreira G, et al. Diagnostic evaluation of ELISA and chemiluminescent assays as alternative screening tests to indirect immunofluorescence for the detection of antibodies to cellular antigens. Am J Clin Pathol. 2016;145(3):323-331. doi:10.1093/ajcp/aqv083
- Agmon-Levin N, Damoiseaux J, Kallenberg C, et al. International recommendations for the assessment of autoantibodies to cellular antigens referred to as anti-nuclear antibodies. Ann Rheum Dis. 2014;73(1):17-23. doi:10.1136/annrheumdis-2013-203863
- Kayser C, Fritzler MJ. Autoantibodies in systemic sclerosis: unanswered questions. Front Immunol. 2015;6:167. doi:10.3389/fimmu.2015.00167
- Knobler R, Moinzadeh P, Hunzelmann N, et al. European Dermatology Forum S1-guideline on the diagnosis and treatment of sclerosing diseases of the skin, part 1: localized scleroderma, systemic sclerosis and overlap syndromes. J Eur Acad Dermatol Venereol. 2017;31(9):1401-1424. doi:10.1111/jdv.14458
Content reviewed 08/2024
16088 - Centromere B Antibody
16814 - ANA Cascade (ANA, IFA with Reflex to Titer and Pattern, and Reflex to 11 Ab Cascade)
ANA IFA on HEp-2 cells is considered by the American College of Rheumatology as the current gold standard for ANA screening because of its overall high sensitivity for several autoimmune diseases [1]. Knowing the titer and fluorescent staining pattern can be helpful in interpreting positive results. Because ANAs can be present in many other diseases as well as in individuals without autoimmune disease, testing for specific autoantibodies may offer greater specificity and help support the diagnosis.
Individuals with negative results on the ANA IFA usually also have negative results on most of the clinically important specific autoantibodies. Therefore, additional testing by immunoassay is not recommended for individuals without positive ANA IFA results and clinical suspicion of relevant autoimmune disease. However, Jo-1 antibody may be detected in ANA IFA-negative patients with some types of myositis, and SSA antibody may be detected in some ANA IFA-negative patients with lupus or Sjogren syndrome [2].
Note that the cascade stops with the first tier in which a positive antibody result is observed. Therefore, even if an antibody included in a subsequent tier is present, it will not be reported with this test. If the ANA IFA result is positive but all the specific autoantibodies tested in the cascade are negative, the patient may still have an autoimmune disease other than those typically associated with the antibodies tested.
The results of this test should be interpreted in the context of pertinent clinical and family history and physical examination findings.
References
1. Methodology of testing for antinuclear antibodies (position statement). 2009. American College of Rheumatology. Updated December 2019. Accessed May 15, 2023. https://assets.contentstack.io/v3/assets/bltee37abb6b278ab2c/blta48818378bc89445/acr-position-statement-methodology-testing-antinuclear-antibodies.pdf
2. Yazdany J, et al. Arthritis Care Res (Hoboken). 2013;65(3):329-339.
249 - ANA Screen,IFA, with Reflex to Titer and Pattern
This immunofluorescence assay (IFA) is often ordered as part of an initial diagnostic evaluation of individuals with clinical suspicion of autoimmune diseases associated with antinuclear antibodies (ANAs). The American College of Rheumatology (ACR) recommends IFA on human epithelial type 2 (HEp-2) cells as the gold standard method for ANA testing because of its overall high sensitivity [1].
ANAs are associated with several autoimmune diseases, such as systemic lupus erythematosus, systemic sclerosis, mixed connective tissue disease, polymyositis, primary biliary cholangitis, rheumatoid arthritis, juvenile rheumatoid arthritis, Sjogren syndrome, and autoimmune hepatitis. The laboratory evaluation for individuals with clinical suspicion of these autoimmune diseases often begins with an ANA screen.
Knowing the ANA titer can be helpful in interpreting positive ANA results. A titer of at least 1:40 is considered positive, but low-positive titers are not uncommon in healthy individuals. Higher titers are generally associated with greater likelihood of autoimmune disease [2]. When results are positive, various fluorescent staining patterns observed in the nucleus or the cytoplasm can aid in the differential diagnosis and guide selection of further testing for specific autoantibodies. The International Consensus on Antinuclear Antibody Pattern provides guidance on interpretation and reporting of IFA staining patterns with HEp-2 cells [3].
Individuals with negative results on the ANA IFA usually also have negative results on specific ANAs. Therefore, subserology testing is generally not recommended in individuals without positive ANA IFA results and clinical suspicion of relevant autoimmune disease [4]. However, Jo-1 antibody may be detected in ANA IFA-negative patients with some types of myositis, and SSA antibody may be detected in some ANA IFA-negative patients with lupus or Sjogren syndrome [4].
The results of this test should be interpreted in the context of pertinent clinical and family history and physical examination findings.
References
1. Methodology of testing for antinuclear antibodies (position statement). 2009. American College of Rheumatology. Updated December 2019. Accessed May 15, 2023. https://assets.contentstack.io/v3/assets/bltee37abb6b278ab2c/blta48818378bc89445/acr-position-statement-methodology-testing-antinuclear-antibodies.pdf
2. Tozzoli R, et al. Am J Clin Pathol. 2002;117(2):316-324.
3. Chan EK, et al. Front Immunol. 2015;6:412.
4. Yazdany J, et al. Arthritis Care Res (Hoboken). 2013;65(3):329-339.
4942 - Scleroderma Antibody (Scl-70)
94954 - ANA, IFA, Cascade and Rheumatoid Arthritis Panel 2, with Reflexes
Note that the cascade stops with the first tier in which a positive antibody result is observed. Therefore, even if an antibody included in a subsequent tier is present, it will not be reported with this test. If the ANA IFA result is positive but all the specific autoantibodies tested in the cascade antibody tests are negative, the patient may still have an autoimmune disease other than those typically associated with the antibodies tested.
38567 - Sm/RNP Antibody
19899 - RNA Polymerase III Antibody
Antibodies to several different types of RNA Polymerases are found in patients with systemic sclerosis. The immunodominant epitope or RNA Pol III was identified and cloned. The recombinant immunodominant epitope of RNA Pol III can be used in ELISA with high specificity to detect anti-RNA Pol III antibodies in patients with the diffuse cutaneous form of systemic sclerosis, with a high incidence of skin involvement.
90073 - ANA Screen,IFA, with Reflex to Titer and Pattern/Systemic Sclerosis Panel 1
This panel may be used as part of an initial diagnostic evaluation of individuals with clinical suspicion of systemic sclerosis (SSc). This panel starts with an immunofluorescence assay (IFA) to detect antinuclear antibodies (ANAs). A positive result will reflex to 3 individual autoantibodies (antibodies to Scl-70, centromere B, and RNA polymerase III) that are associated with SSc.
The laboratory evaluation for individuals with clinical suspicion of autoimmune diseases often begins with an ANA screen. The classic ANA IFA on human epithelial type 2 cells is considered by the American College of Rheumatology (ACR) as the current gold standard because of its overall high sensitivity for several autoimmune diseases [1]. ANAs can be detected in 90% to 95% of the patients with SSc; a negative ANA IFA result decreases the likelihood of SSc but does not exclude it. Knowing fluorescent staining patterns of an ANA IFA can be helpful in the differential diagnosis and guide selection of further testing for specific autoantibodies [2].
Antibodies to centromere, Scl-70, and RNA polymerase III are included in this panel as reflex tests when the ANA IFA result is positive. The presence of any of these 3 autoantibodies is part of the ACR/European League Against Rheumatism (EULAR) classification criteria for SSc [3]. Identification of certain SSc-associated autoantibodies can help distinguish the various types of SSc and the organ(s) likely to be involved, which is important for determining prognosis and optimal treatment [2].
Several other autoantibodies associated with SSc are less prevalent or less recently characterized and thus are not included in this panel (eg, antibodies to U1-snRNP, Th/To, and fibrillarin). Detecting these autoantibodies may also be helpful in the evaluation of SSc [2].
The results of this test should be interpreted in the context of pertinent clinical and family history and physical examination findings.
References
1. Methodology of testing for antinuclear antibodies (position statement). 2009. American College of Rheumatology. Updated December 2019. Accessed May 15, 2023. https://assets.contentstack.io/v3/assets/bltee37abb6b278ab2c/blta48818378bc89445/acr-position-statement-methodology-testing-antinuclear-antibodies.pdf
2. Stochmal A, et al. Clin Rev Allergy Immunol. 2020;58(1):40-51.
3. van den Hoogen F, et al. Arthritis Rheum. 2013;65(11):2737-2747.
94685 - Systemic Sclerosis (Scleroderma) 12 Antibodies Panel 2
36378 - ANAlyzeR™ ANA, IFA with Reflex Titer/Pattern, Systemic Autoimmune Panel 1
ANAlyzeR ANA, IFA with Reflex Titer/Pattern, Systemic Autoimmune Panel 1
Reference ranges are provided as general guidance only. To interpret test results use the reference range in the laboratory report.
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