INTRODUCTION — The indirect immunofluorescence (IIF) test for antinuclear antibodies (ANA), using the human cell line HEp-2 as substrate, is a commonly used assay to detect human autoantibodies. The results of ANA testing are reported in two parts: the titer of the antibodies and the staining pattern produced by the antibodies. The titer of the antibodies refers to the highest dilution of serum that produces visible fluorescence. The ANA pattern refers to the distribution of staining produced by autoantibodies reacting with antigens in HEp-2 cells. Although the term anti-"nuclear" antibody would imply that only nuclear staining patterns are important, the international consensus is that all staining patterns observed in HEp-2 cells, including nuclear, cytoplasmic, and cell-cycle related patterns, should be reported [1].

This topic review will cover the three broad categories of ANA staining patterns: nuclear, cell cycle-associated, and cytoplasmic. Within each of these categories, individual patterns will be defined and autoantibodies that produce the staining patterns will be identified. The disease associations of autoantibodies producing the staining patterns will be described as well as additional laboratory tests that may be used to further characterize the autoantibodies. The measurement and clinical significance of ANA titer is reviewed elsewhere. (See "Measurement and clinical significance of antinuclear antibodies".)

NOMENCLATURE — There is limited agreement among laboratories as to which antinuclear antibody (ANA) staining patterns should be identified and reported to clinicians. The patterns to be reported are determined by individual laboratory directors. The directors also choose from among HEp-2 cell slides prepared by different companies. These companies may use different fixative and permeabilization techniques. Depending on the preparation of the HEp-2 cell substrate, some autoantibodies may or may not be detected by indirect immunofluorescence (IIF). The clinician should know which staining patterns are recognized and reported by their reference laboratory.

An International Consensus on Antinuclear Antibody Patterns (ICAP) workshop was held in Sao Paolo, Brazil in 2014. The goal of the first and subsequent meetings was to "promote harmonization and understanding of autoantibody nomenclature and thereby optimizing ANA usage in patient care." Members of ICAP reached consensus on the definition and clinical relevance of 29 distinct HEp-2 cell IIF patterns [2,3]. Images of the HEp-2 cell patterns, labeled AC-1 to AC-29, are available at ANApatterns.org.

ICAP workshop participants suggested that there are 11 staining patterns (6 nuclear, 5 cytoplasmic) that "must" be reported by all "competent-level" laboratories. An additional 18 ANA staining patterns "should" be reported by "expert-level" laboratories [2,3]. Results of a survey of members of the Association of Medical Laboratory Immunologists and American Society for Clinical Pathology suggest increasing awareness and acceptance of the ICAP guidelines [4].

ANA STAINING PATTERNS INVOLVING THE NUCLEUS

Homogeneous

●Definition – The homogenous antinuclear antibody (ANA) pattern refers to diffuse staining of the nucleus in resting cells. There is also diffuse staining of the chromosome region in dividing cells. The homogenous staining pattern was reported in 36 percent of more than 9200 ANA-positive serum samples tested at the University Hospitals in Leuven, Belgium [5].

●Targets of antibodies – Histone proteins, double-stranded (ds)DNA, single-stranded DNA.

●Disease association – Homogeneous nuclear staining may be seen in patients with many different systemic autoimmune diseases including systemic lupus erythematosus (SLE), drug-induced lupus, Sjögren's syndrome (SS), scleroderma (SSc), and rheumatoid arthritis (RA), as well as organ-specific autoimmune diseases including Hashimoto's thyroiditis, primary biliary cholangitis (PBC), and autoimmune hepatitis.

Specific antibodies responsible for homogeneous staining, such as anti-dsDNA antibodies, may be detected by indirect immunofluorescence (IIF) using Crithidia luciliae assay, enzyme-linked immunosorbent assay (ELISA), or fluorescent bead assay and may be helpful to support the diagnosis of SLE. Anti-dsDNA antibodies are present in approximately 40 to 60 percent of patients with SLE and are highly specific for the disease (75 to 99 percent, depending on the assay used to detect the antibodies) [6]. In addition, anti-dsDNA antibodies may develop in patients who are treated with inhibitors of tumor necrosis factor (TNF)-alpha [7]. These patients may or may not have symptoms of drug-induced lupus. (See "Clinical manifestations and diagnosis of systemic lupus erythematosus in adults", section on 'Laboratory testing' and "Drug-induced lupus", section on 'Causative drugs' and "Tumor necrosis factor-alpha inhibitors: An overview of adverse effects", section on 'Autoimmunity and autoantibodies'.)

In patients suspected of having drug-induced lupus that is not associated with anti-TNF-alpha therapy, ELISA or fluorescent bead assays (hereafter referred to as "solid phase assays") for anti-histone antibodies may be helpful; a negative test for anti-histone antibodies would argue against a diagnosis of drug-induced lupus. (See "Drug-induced lupus", section on 'Laboratory tests and characteristic autoantibodies'.)

The homogeneous ANA pattern may obscure other staining patterns, such as nuclear speckled. Therefore, additional testing for specific autoantibodies will be required to fully characterize the spectrum of autoantibodies that may be present in serum that produces the homogeneous ANA staining pattern.

●International Consensus on Antinuclear Antibody Patterns (ICAP) code – AC-1 (see 'Nomenclature' above)

Nuclear speckled — Three types of nuclear speckled patterns ("fine," "coarse," and "fine dense") have been described. Individual laboratories may or may not distinguish between these different patterns.

Fine speckled

●Definition – Fine speckled staining refers to hundreds to thousands of dots present throughout the nucleus. Nucleoli may or may not be stained, depending on the specific autoantibody. Except for antibodies directed against Scl70 (topoisomerase I), there is no staining of the chromosome region in dividing cells. The fine nuclear speckled pattern was reported in 20 percent of more than 9200 ANA-positive serum samples [5]

●Targets of antibodies – Ro60, La, Topoisomerase I (also known as Scl70), Ku, Mi-2, transcriptional intermediary factor (TIF)-1gamma, and melanoma differentiation-associated protein-5 (MDA5; also known as CADM-140 and RNA helicase).

●Disease association – As is true with the homogeneous pattern, the fine speckled pattern may be present in patients with a broad spectrum of autoimmune diseases. Additional testing is required to further characterize the antibodies in patient serum that produce this staining pattern.

Antibodies directed against Ro60 are present in approximately 75 percent of patients with subacute cutaneous lupus erythematosus, 60 percent of patients with SS, 30 percent of patients with SLE, 30 percent of patients with PBC, 20 percent of patients with polymyositis (PM), and 15 percent of patients with RA [8]. Anti-Ro60 and/or anti-Ro52 antibodies (see below) are present in nearly all women who have a child with neonatal lupus syndrome (NLS). It is critical to note that anti-Ro60 antibodies may not be detected in Hep-2 cell substrate if the cells are fixed with methanol (as opposed to acetone) [9]. Because anti-Ro60 antibodies are not detected using some HEp-2 cell substrates, if anti-Ro60 is the only autoantibody present in a patient with SLE or SS, the patient will appear to have ANA-negative disease. Anti-Ro60 antibodies may be the only autoantibody present in 25 to 50 percent of all patients with "ANA-negative" SLE [10]. (See "The anti-Ro/SSA and anti-La/SSB antigen-antibody systems" and "Clinical manifestations and diagnosis of systemic lupus erythematosus in adults", section on 'ANA-negative lupus' and "Neonatal lupus: Epidemiology, pathogenesis, clinical manifestations, and diagnosis", section on 'Autoantibodies'.)

Antibodies directed against La are present in approximately 20 percent of patients with SS and are considered relatively specific for this disease. Anti-La antibodies may also be detected in patients with SLE and in women who have a child with NLS. (See "Diagnosis and classification of Sjögren's syndrome", section on 'Antibodies to Ro/SSA and La/SSB' and "Clinical manifestations and diagnosis of systemic lupus erythematosus in adults", section on 'Laboratory testing' and "Neonatal lupus: Epidemiology, pathogenesis, clinical manifestations, and diagnosis", section on 'Autoantibodies'.)

Antibodies directed against Scl70 are present in approximately 20 to 40 percent of patients with diffuse SSc and are rarely detected in patients with other autoimmune diseases or in healthy individuals. In patients with diffuse SSc, anti-Scl70 antibodies are associated with the presence of interstitial pulmonary fibrosis [11]. (See "Clinical manifestations and diagnosis of systemic sclerosis (scleroderma) in adults", section on 'Laboratory testing'.)

Anti-Ku antibodies are rare, with an estimated prevalence of 0.5 percent of all ANA-positive sera. Anti-Ku antibodies are present in approximately 2.5 percent of patients with SSc and are associated with the presence of myositis and arthritis [12]. Anti-Ku antibodies may also be detected in patients with SLE and PM.

Antibodies directed against Mi-2 (a nuclear DNA helicase involved in transcriptional activation) are a rare cause of a speckled ANA. Anti-Mi-2 antibodies are present in approximately 15 percent of patients with DM [13]. In general, patients with these antibodies have skin findings characteristic of DM, including heliotrope rash and Gottron papules, but do not develop lung disease and respond well to corticosteroids [14]. (See "Overview of and approach to the idiopathic inflammatory myopathies", section on 'Myositis-specific autoantibodies'.)

Antibodies directed against TIF-1gamma (as detected by immunoprecipitation) may be present in 15 to 25 percent of patients with dermatomyositis (DM) and may be associated with the presence of an underlying malignancy [15]. (See "Malignancy in dermatomyositis and polymyositis", section on 'Serum autoantibodies'.)

Anti-MDA5 antibodies may produce speckled staining in either the nucleus or cytoplasm of HEp-2 cells. Some HEp-2 cell preparations have little or no detectable MDA5, resulting in a negative test for ANA. When anti-MDA5 antibodies were detected using immunoprecipitation, approximately 25 percent of DM patients were found to have antibodies, and the presence of anti-MDA5 antibodies was associated with amyopathic disease and rapidly progressive interstitial lung disease (ILD) [14,16]. (See "Overview of and approach to the idiopathic inflammatory myopathies", section on 'Myositis-specific autoantibodies'.)

●ICAP code – AC-4 (see 'Nomenclature' above)

Coarse speckled

●Definition – The coarse nuclear speckled pattern refers to hundreds to thousands of nuclear dots, variable in size, but generally larger than the dots seen in the fine speckled pattern. There is no staining of nucleoli or dividing chromosomes.

●Targets of antibodies – Sm, U1 ribonuclear protein (RNP).

●Disease association – Antibodies directed against Sm are relatively specific for the diagnosis of SLE. Antibodies directed against Sm may be detected in 10 to 50 percent of SLE patients, and estimates of the specificity of anti-Sm antibodies for the diagnosis of SLE range from 55 to 100 percent [17-20]. Anti-Sm antibodies may also be detected in patients with mixed connective tissue disease (MCTD), but are rare in other autoimmune diseases.

Antibodies directed against U1RNP antigens are present in all patients with MCTD because the antibodies are a critical component of the diagnosis of this disease. Anti-U1RNP antibodies may also be present in 30 to 40 percent of patients with SLE, as well as some patients with RA, SS, SSc, and PM. (See "Clinical manifestations and diagnosis of systemic lupus erythematosus in adults", section on 'Laboratory testing' and "Definition and diagnosis of mixed connective tissue disease", section on 'Autoantibodies' and "Clinical manifestations and diagnosis of systemic sclerosis (scleroderma) in adults", section on 'Laboratory testing' and "Overview of and approach to the idiopathic inflammatory myopathies", section on 'Myositis-specific autoantibodies' and "Clinical manifestations of mixed connective tissue disease".)

●ICAP code – AC-5 (see 'Nomenclature' above)

Dense fine speckled pattern

●Definition – In the dense fine speckled pattern, the speckles are distributed throughout the nucleus of interphase cells, excluding nucleoli. This pattern differs from the fine and coarse speckled patterns in that the speckles associate with chromosomes in dividing cells. Estimates of the prevalence of the dense fine speckled pattern ranges from 0.8 [21] to 37 percent [22].

●Antibody target – Dense fine speckled, 70 kD (DFS70; also known as lens epithelium-derived growth factor protein 75 kD [LEDGFp75]).

●Disease association – In a study of 597 healthy hospital workers, 20 percent were found to have a positive ANA at a dilution of >1:40. More than half of these ANA-positive individuals had antibodies that produced the dense fine speckled pattern [23]. Other studies have confirmed that patients with antibodies producing this staining pattern have a low prevalence of underlying autoimmune disease [24-26]. Anti-DFS70 antibodies were found to be the only detectable autoantibody in only 1.1 percent of patients with SLE in a large, international cohort. These results further suggest that anti-DFS70 autoantibodies are not associated with systemic rheumatic disease [27]. In a patient population with a low prior probability of having an autoimmune disease, the presence of antibodies producing the DFS70 pattern may be considered a reassuring result, even when these autoantibodies are present in high (>1:640) titer [26]. However, the DFS70 staining pattern may obscure patterns produced by co-occurring autoantibodies. In patients with a high prior probability of autoimmune disease (as determined by history, physical examination, and other laboratory findings), additional testing using solid phase assays may be required to detect disease-associated autoantibodies, including those directed against Ro/SSA, La/SSB, Sm, U1 RNP, Scl70, and histone proteins.

●ICAP code – AC-2 (see 'Nomenclature' above)

Nucleolar

●Definition – Three distinct types of nucleolar staining patterns have been described: "homogeneous," "clumped," and "speckled" or "punctate." However, most clinical laboratories do not distinguish between these subtypes. The prevalence of antinucleolar antibodies in serum samples varies widely, from 1.4 percent of all samples submitted for ANA testing to a hospital in the United Kingdom [28] to 17 percent of more than 9200 ANA-positive sera [5].

●Targets of autoantibodies – Components of the PM/Scl complex (PM-Scl-75, PM-Scl-100), U3RNP (also known as fibrillarin); the 7-2RNP complex (Th/To); RNA polymerases I, II, and III; Topoisomerase I (Scl70).

●Disease association – Antinucleolar antibodies may be detected in patients with SLE, RA, SS, SSc, PM, DM, MCTD, and the Raynaud phenomenon. Antibodies producing the nucleolar staining pattern have been reported in 15 to 40 percent of patients with SSc [11] and the presence of these antibodies has important implications for diagnosis and prognosis. Therefore, in the appropriate clinical setting, additional testing using solid phase or immunoprecipitation assays may be required to identify the autoantibodies that are responsible for the nucleolar staining pattern.

The PM/Scl complex, also known as the exosome, is a multi-protein structure that degrades RNA. Antibodies directed against one or both of two components of the exosome, PM-Scl-75 and PM-Scl-100, are present in 5 to 10 percent of SSc patients and are associated with limited skin disease and decreased risk of pulmonary and renal disease, but increased risk of associated inflammatory myositis [11]. (See "Overview of and approach to the idiopathic inflammatory myopathies", section on 'Myositis-associated autoantibodies'.)

U3-RNP (fibrillarin) is involved in the first step of processing pre-ribosomal RNA. Anti-U3-RNP antibodies are present in 5 to 10 percent of patients with SSc and are associated with diffuse skin disease, pulmonary artery hypertension, pulmonary fibrosis, and myositis [29]. (See "Clinical manifestations, evaluation, and diagnosis of interstitial lung disease in systemic sclerosis (scleroderma)", section on 'Laboratory findings'.)

The Th/To complex functions as a ribonuclease that removes the 5' leader sequences from precursor tRNA molecules. Antibodies directed against components of the Th/To complex are present in 2 to 5 percent of patients with SSc and are associated with limited skin disease but increased risk of pulmonary fibrosis and renal crisis [29]. (See "Clinical manifestations, evaluation, and diagnosis of interstitial lung disease in systemic sclerosis (scleroderma)", section on 'Laboratory findings'.)

Antibodies directed against RNA polymerases are detected in approximately 20 percent of patients with SSc and have a high degree of specificity for this disease. The presence of anti-RNA polymerase antibodies is associated with diffuse skin involvement and increased risk of renal crisis. Pulmonary fibrosis is uncommon in patients with anti-RNA polymerase antibodies [30]. (See "Clinical manifestations and diagnosis of systemic sclerosis (scleroderma) in adults", section on 'Laboratory testing'.)

Antibodies directed against Scl70 (topoisomerase I) produce nuclear speckled (see 'Fine speckled' above) as well as nucleolar staining patterns.

●ICAP codes – AC-8 ("homogeneous"), AC-9 ("clumped"), AC-10 ("speckled" or "punctate") (see 'Nomenclature' above)

Nuclear dot staining patterns — Three types of nuclear dot staining patterns have been described: centromere, PML-Sp100 nuclear body, and Cajal body.

Centromere

●Definition – The centromere staining pattern is characterized by the presence of 30 to 60 discrete, large speckles present in the nucleus of resting cells. The speckles are larger and fewer in number than those seen in the fine and coarse speckled patterns. The speckles align with the chromosome region in dividing cells. Antibodies directed against centromeres were detected in 3 percent of more than 9200 ANA-positive sera [5].

●Targets of antibodies – Centromere proteins A, B, and C.

●Disease association – Anti-centromere antibodies are present in approximately 30 percent of patients with limited SSc. The antibodies are associated with calcinosis and pulmonary hypertension. Anti-centromere antibodies are also present in approximately 15 percent of patients with PBC; such patients tend to have a worse outcome than anti-centromere antibody-negative patients [31]. Anti-centromere antibodies were present in 4 to 15 percent of patients with SS [32,33] and approximately 4 percent of SLE patients [34]. Anti-centromere antibodies may be also detected in patients with isolated Raynaud phenomenon who do not have evidence of a systemic autoimmune disease [35]. (See "Clinical manifestations and diagnosis of systemic sclerosis (scleroderma) in adults", section on 'Laboratory testing' and "Clinical manifestations, diagnosis, and prognosis of primary biliary cholangitis (primary biliary cirrhosis)", section on 'Laboratory tests' and "Clinical manifestations and diagnosis of Raynaud phenomenon".)

●ICAP code – AC-3 (see 'Nomenclature' above)

PML-Sp100 nuclear body

●Definition – The PML-Sp100 nuclear body staining pattern is characterized by the presence of 5 to 20 discrete, large speckles present in the nucleus of resting cells. The pattern can be distinguished from the centromere pattern because there are fewer dots in each cell and there is no staining of chromosomes in dividing cells. Antibodies directed against PML-Sp100 nuclear bodies were detected in 0.2 percent of more than 9200 ANA-positive sera [5].

●Target of antibodies – Speckled-100 kD (Sp100), promyelocytic leukemia protein (PML).

●Disease association – The PML-Sp100 nuclear body is a multi-protein cellular structure that is involved in a variety of cellular functions including gene transcription, cellular apoptosis, cell-cycle control, and DNA repair. Antibodies directed against components of the PML-Sp100 nuclear body (PML, Sp100, or usually both proteins) are present in 20 percent of patients with anti-mitochondrial antibody (AMA)-positive PBC [31]. Anti-PML-Sp100 nuclear body antibodies are also present in approximately 40 percent of PBC patients who do not have AMA as detected by IIF [36] and may be the only antibody marker in some PBC patients. Anti-PML-Sp100 nuclear body antibodies are rarely detected in patients with other autoimmune diseases. Because the PML-Sp100 nuclear body staining pattern is similar to that of the Cajal body staining pattern (see 'Cajal body' below) and because anti-PML-Sp100 nuclear body antibodies are specific for the diagnosis of PBC, it is critical to confirm the presence of these antibodies using a solid phase assay. (See "Clinical manifestations, diagnosis, and prognosis of primary biliary cholangitis (primary biliary cirrhosis)".)

●ICAP code – AC-6 (see 'Nomenclature' above)

Cajal body

●Definition – The Cajal body staining pattern is characterized by the presence of 0 to 8 discrete, large speckles in the nucleus of resting cells. There is no staining of chromosomes in dividing cells. Antibodies directed against Cajal bodies were present in 1.3 percent of more than 9200 ANA-positive sera [5].

●Target of antibodies – p80-coilin.

●Disease association – Autoantibodies directed against Cajal bodies are rarely present in patients with SS and PBC [31]. These antibodies may also be detected in patients with a variety of other autoimmune diseases, including SLE, SSc, and PM [37-39], as well as in healthy individuals. Because antibodies directed against Cajal bodies are rarely detected and are not disease-specific, the clinical significance of these antibodies is uncertain.

●ICAP code – AC-7 (see 'Nomenclature' above)

Nuclear envelope — Two types of nuclear envelope staining patterns have been reported. The first is characterized by smooth, continuous, linear staining of the nuclear envelope; the second by discontinuous linear staining of the nuclear envelope. Autoantibodies directed against components of the nuclear envelope were detected in 0.5 percent of more than 9200 ANA-positive sera [5].

Nuclear envelope pattern

●Targets of antibodies – Nuclear lamin proteins, lamin-associated proteins.

●Disease association – Autoantibodies directed against lamins and lamin-associated proteins have been reported in a broad spectrum of diseases, including SLE, SS, antiphospholipid syndrome (APS), and autoimmune liver disease [40]. Because of the uncertain sensitivity of detecting these antibodies in any autoimmune disease and the lack of disease specificity, the clinical significance of these antibodies is uncertain.

●ICAP code – AC-11 (see 'Nomenclature' above)

Nuclear pore complex pattern

●Targets of antibodies – Glycoprotein-210 kD (Gp210), nucleoporin-62 kD (Nup62).

●Disease association – Antibodies directed against the nuclear pore complex are detected by IIF in approximately 20 percent of patients with AMA-positive PBC [31]. Using an ELISA assay and either recombinant Gp210 or a Gp210 peptide fragment as substrate, anti-Gp210 antibodies were detected in 25 percent of PBC patients and were more than 95 percent specific for this diagnosis. The prevalence of anti-Nup62 antibodies in patients with PBC was approximately 20 percent [41]. (See "Clinical manifestations, diagnosis, and prognosis of primary biliary cholangitis (primary biliary cirrhosis)", section on 'Laboratory tests'.)

Not all clinical laboratories distinguish between nuclear envelope and nuclear pore complex staining patterns. Because of the specificity of anti-Gp210 antibodies for the diagnosis of PBC, it is critical to test for these antibodies by ELISA in patients with autoantibodies reacting with the nuclear envelope.

●ICAP code – AC-12 (see 'Nomenclature' above)

CELL CYCLE-ASSOCIATED STAINING PATTERNS

PCNA

●Definition – Autoantibodies producing the proliferating cell nuclear antigen (PCNA) pattern produce nuclear speckled staining during the G1 phase of the cell cycle, when cells are increasing in size. During S phase, when DNA replication occurs, there is dense homogeneous nuclear staining. There is no staining in dividing cells. Autoantibodies producing the PCNA staining pattern were detected in 0.3 percent of more than 9200 antinuclear antibody (ANA)-positive sera [5].

●Target of antibodies – Delta chain of DNA polymerase.

●Disease association – Autoantibodies producing the PCNA staining pattern were originally identified in a small percentage of patients with systemic lupus erythematosus (SLE) [42]. Subsequent studies reported the presence of anti-PCNA antibodies in a variety of disorders including other autoimmune diseases, viral hepatitis, and malignancies [43]. In a retrospective study, only 1 of 28 patients with antibodies producing the PCNA staining pattern was diagnosed with SLE, while six patients were diagnosed with autoimmune thyroid disease [5]. Based on these results, the clinical significance of antibodies producing the PCNA staining pattern is uncertain.

●ICAP code – AC-13 (see 'Nomenclature' above)

CENP-F

●Definition – The centromere protein-F (CENP-F) staining pattern is characterized by fine speckled staining of the nucleus in resting cells; staining spares the nucleolus. The strongest staining cells are in the G2 phase of cell division, during the "gap" between DNA synthesis and mitosis. There is centromere-associated staining of dividing cells. In addition, during the late stage of cell division, there is staining of the middle region of separating spindles, known as the spindle "midzone." In cells that have progressed further in cell division, there may be staining of a small structure between the two dividing cells known as the mid-body. The CENP-F staining pattern was detected in 0.3 percent of more than 9200 ANA-positive sera [5].

●Target of antibodies – CENP-F.

●Disease association – Approximately half of the patients with autoantibodies producing the CENP-F staining pattern have an underlying malignancy. Associated malignancies include breast, lung, and prostate cancer, as well as lymphoma [44].

●ICAP code – AC-14 (see 'Nomenclature' above)

Mitotic spindle apparatus — Two types of nuclear mitotic apparatus staining patterns have been described: nuclear mitotic apparatus-1 (NuMA1) and NuMa2.

NuMA1 staining pattern

●Definition – The NuMA1 staining pattern is characterized by speckled nuclear staining in resting cells. In dividing cells, there is staining of the spindles near the centrosomes, but the staining does not extend to the centromeres. There is no staining of spindles between the dividing chromosomes. The NuMA1 staining pattern was detected in 0.7 percent of more than 9200 ANA-positive sera [5].

●Target of antibodies – NuMA1.

●Disease association – In a review of the diagnoses in patients whose serum produced the NuMA1 staining pattern, 20 percent had Sjögren's syndrome (SS), 12 percent SLE, and 40 percent other autoimmune diseases [45]. (See "Diagnosis and classification of Sjögren's syndrome".)

●ICAP code – AC-26 (see 'Nomenclature' above)

NuMA2 staining pattern

●Definition – Antibodies producing the NuMA2 staining pattern do not stain the nuclei of resting cells. In dividing cells, there is staining of the spindle fibers that extends from the centrosomes to the centromeres. Later in cell division, there is also staining of spindles between the centromeres. The NuMA2 staining pattern was detected in 0.06 percent of 9200 ANA-positive sera [5].

●Target of antibodies – Human kinesin-like protein (HsEg5; also known as "kinesin family member 11 [KIF11]").

●Disease association – Antibodies that produce the NuMA2 staining pattern are not associated with any specific autoimmune disease [45].

●ICAP code – AC-25 (see 'Nomenclature' above)

CYTOPLASMIC STAINING PATTERNS

Antimitochondrial antibody

●Definition – The anti-mitochondrial antibody (AMA) staining pattern, as seen using the HEp-2 cell substrate, is characterized by the presence of granular and filamentous staining throughout the cytoplasm. On a substrate that is rich in mitochondria, such as rat kidney, intense cytoplasmic staining may be seen in the distal tubular cells. The AMA staining pattern was detected in 74 of 8300 serum samples submitted for testing to the Clinical Immunology Laboratory at the Massachusetts General Hospital in 2012 [46].

●Targets of antibodies – E2 component of pyruvate dehydrogenase complex (also known as dihydrolipoamide S-acetyltransferase [DLAT]), 2-oxo-glutarate dehydrogenase (dihydrolipoamide dehydrogenase [DLD]), and branched-chain 2-oxo-acid dehydrogenase (BCKDC), as well as other mitochondrial components.

●Disease association – Antibodies directed against DLAT, DLD, and BCKDC are highly specific for the diagnosis of primary biliary cholangitis (PBC). Autoantibodies directed against other components of mitochondria may be detected in a broad spectrum of diseases, including infections, myocarditis, systemic lupus erythematosus (SLE), and drug-induced hepatitis [47]. Because only antibodies directed against DLAT, DLD, and BCKDC are specific for PBC, if the AMA staining pattern is detected by indirect immunofluorescence (IIF), then solid phase assays should be used to confirm the presence of antibodies directed against DLAT, DLD, and/or BCKDC [48]. (See "Clinical manifestations, diagnosis, and prognosis of primary biliary cholangitis (primary biliary cirrhosis)".)

●ICAP code – AC-21 (see 'Nomenclature' above)

Cytoplasmic fine speckled

●Definition – In this staining pattern, fine speckles are distributed throughout the cytoplasm of resting HEp-2 cells. Antibodies producing the cytoplasmic fine speckled staining pattern were present in 1.2 percent of more than 9200 antinuclear antibody (ANA)-positive sera [5].

●Target of antibodies – Jo-1 (histidine tRNA synthetase) or any of seven other tRNA synthetases, ribosomal P proteins, Ro52, signal recognition particle (SRP).

●Disease association – Antibodies directed against aminoacyl-tRNA synthetases are present in 25 to 35 percent of patients with dermatomyositis (DM) or polymyositis (PM). These antibodies are rarely detected in patients with other autoimmune diseases. In general, anti-tRNA synthetase antibodies are markers of a subtype of DM or PM characterized by interstitial lung disease (ILD), myositis, nonerosive arthritis, Raynaud phenomenon, and "mechanic's hands." The term "mechanic's hands" refers to hyperkeratosis at the ulnar aspect of the thumb and radial aspect of the index finger. Antibodies directed against Jo-1 (histidyl tRNA synthetase) are the most common anti-tRNA synthetase autoantibody, found in 20 to 30 percent of patients with PM and in 60 to 70 percent of PM patients with ILD [15]. (See "Overview of and approach to the idiopathic inflammatory myopathies", section on 'Myositis-specific autoantibodies'.)

Antibodies directed against ribosomal P antigens are found in 10 to 20 percent of patients with SLE and are rare in other autoimmune diseases. (See "Antiribosomal P protein antibodies".)

Anti-Ro52 antibodies are present in approximately 40 percent of patients with SLE, 75 percent of patients with Sjögren's syndrome (SS), 35 percent of patients with PM, and 20 percent of patients with systemic sclerosis (SSc) [8]. Anti-Ro52 and/or anti-Ro60 antibodies are present in nearly all women who have children with neonatal lupus syndrome (NLS). (See "Clinical manifestations and diagnosis of systemic lupus erythematosus in adults", section on 'Laboratory testing' and "Diagnosis and classification of Sjögren's syndrome", section on 'Antibodies to Ro/SSA and La/SSB' and "Neonatal lupus: Epidemiology, pathogenesis, clinical manifestations, and diagnosis", section on 'Autoantibodies'.)

The signal recognition particle is a cytoplasmic, macromolecular structure that binds to an amino terminal signal sequence that is present in some newly synthesized proteins and directs the proteins to the endoplasmic reticulum. Anti-SRP antibodies are detected in approximately 10 percent of patients with inflammatory muscle disease and identify a subset of patients with rapidly progressive, necrotizing disease [49,50]. (See "Overview of and approach to the idiopathic inflammatory myopathies", section on 'Myositis-specific autoantibodies'.)

●ICAP code – AC-19 and AC-20 (see 'Nomenclature' above)

Cytoskeletal staining

●Definition – The cytoskeletal staining pattern is characterized by staining of a network of fibers in the cytoplasm.

●Targets of antibodies – Actin and actin-associated proteins, cytokeratin, tropomyosin, vimentin.

●Clinical significance – Anti-actin antibodies are relatively specific markers for the diagnosis of Type I autoimmune hepatitis, but may also be detected in patients with viral hepatitis [51]. Antibodies directed against other components of the cytoskeleton are not disease-specific. Therefore, in a patient with suspected autoimmune hepatitis, the presence of anti-actin antibodies should be sought by enzyme-linked immunosorbent assay (ELISA) using purified actin antigen.

●ICAP codes – AC-15, AC-16, and AC-17 (see 'Nomenclature' above)

Golgi apparatus

●Definition – The Golgi apparatus pattern is characterized by irregular staining of the cytoplasm adjacent to the nucleus. The Golgi apparatus staining pattern was detected in 0.4 percent of more than 9200 ANA-positive sera [5].

●Targets of antibodies – A large number of components of the Golgi apparatus have been found to be targets of autoantibodies, including gigantin, golgin 245, golgin 110, and others.

●Clinical significance – Anti-Golgi apparatus antibodies were initially identified using serum from a patient with SS. Antibodies that produce this staining pattern were subsequently detected in patients with SLE, rheumatoid arthritis (RA), sarcoidosis, idiopathic cerebellar ataxia, and viral infections [52-54]. Because anti-Golgi apparatus autoantibodies are rare and there is no consistent disease association, the clinical significance of these antibodies is uncertain.

●ICAP code – AC-22 (see 'Nomenclature' above)

Rods and rings

●Definition – The rods and rings staining pattern is characterized by staining of one or a few circular and/or cylindrical structures in the cytoplasm of HEp-2 cells. This staining pattern can be detected using some, but not all, commercial HEp-2 cell slides [55].

●Targets of antibodies – Inosine monophosphate dehydrogenase 2 (IMPDH2) and cytidine triphosphate synthase 1 (CTPS1).

●Clinical significance – This cytoplasmic staining pattern may be produced by serum from a subset of patients with hepatitis C who have received antiviral therapy with pegylated interferon-alpha and ribavirin. Antibodies directed against rods and rings are rarely detected in patients with autoimmune diseases [56].

●ICAP code – AC-23 (see 'Nomenclature' above)

SOCIETY GUIDELINE LINKS — Links to society and government-sponsored guidelines from selected countries and regions around the world are provided separately. (See "Society guideline links: Antinuclear antibodies".)

SUMMARY

●The indirect immunofluorescence (IIF) test for antinuclear antibodies (ANA), using the human cell line HEp-2 as substrate, is a commonly used assay to detect human autoantibodies. The results of ANA testing are reported in two parts: the titer of the antibodies and the staining pattern produced by the antibodies. The titer of the antibodies refers to the highest dilution of serum that produces visible fluorescence. The ANA pattern refers to the distribution of staining produced by autoantibodies reacting with antigens in the HEp-2 cell nucleus and cytoplasm. (See 'Introduction' above.)

●The International Consensus on Antinuclear Antibody Patterns (ICAP) workshop reached an agreement on the nomenclature, definition, and clinical significance of 29 distinct ANA staining patterns. There is, as yet, limited consensus among laboratories as to which ANA staining patterns should be identified and reported to clinicians. Depending on the preparation of the HEp-2 cell substrate, some autoantibodies may or may not be detected by IIF. The clinician should know which staining patterns are recognized and reported by their reference laboratory. (See 'Nomenclature' above.)

●There are three broad categories of ANA staining patterns: nuclear, cell cycle-associated, and cytoplasmic. Within each of these categories there are individual staining patterns produced by specific autoantibodies. The disease associations, or lack thereof, for each of the staining patterns are discussed above. (See 'ANA staining patterns involving the nucleus' above and 'Cell cycle-associated staining patterns' above and 'Cytoplasmic staining patterns' above.)

●ANA staining patterns involving the nucleus include homogeneous, nuclear speckled, nucleolar, nuclear dot, and nuclear envelope. Three types of nuclear speckled patterns ("fine," "coarse," and "dense fine") have been described. Three types of nuclear dot staining patterns have been described: centromere, PML-Sp100 nuclear body, and Cajal body. (See 'ANA staining patterns involving the nucleus' above and 'Homogeneous' above and 'Nuclear speckled' above and 'Nucleolar' above and 'Nuclear envelope' above.)

●Cell cycle-associated staining patterns include proliferating cell nuclear antigen (PCNA) pattern, centromere protein-F (CENP-F), and mitotic spindle apparatus. Two types of nuclear mitotic apparatus staining patterns have been described: nuclear mitotic apparatus-1 (NuMA1) and NuMa2. (See 'Cell cycle-associated staining patterns' above and 'PCNA' above and 'CENP-F' above and 'Mitotic spindle apparatus' above.)

●Cytoplasmic staining patterns include anti-mitochondrial antibody (AMA), cytoplasmic fine speckled, cytoskeletal staining, Golgi apparatus, and rods and rings. (See 'Cytoplasmic staining patterns' above and 'Antimitochondrial antibody' above and 'Cytoplasmic fine speckled' above and 'Cytoskeletal staining' above and 'Golgi apparatus' above and 'Rods and rings' above.)

ACKNOWLEDGMENT — The UpToDate editorial staff acknowledges Peter Schur, MD, who contributed to an earlier version of this topic review.