Determination Of Antinuclear Antibodies Research Articles

Diagnostic significance of the determination of antinuclear antibodies in children with autoimmune hepatitis

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Determination of antinuclear antibodies (ANA) in systemic rheumatic diseases

Determination of antinuclear antibodies (ANA) in systemic rheumatic diseases

Methodological aspects of anti-nuclear antibodies detection: EFLM, EASI, ICAP AND RAMLD recommendations

This is a review of international and Russian recommendation for the study of anti-nuclear antibodies (ANA) in autoimmune inflammatory rheumatic diseases (AIRD) and autoimmune liver diseases (ALD), including a description of the most important methodological aspects. The main purpose of laboratory diagnostics of AIRD and ALD is to obtain objective information about the presence and immunopathological changes, which is an important tool for early diagnosis, assessment of activity, severity, prognosis of the disease and the effectiveness of therapy. The positive results of ANA determination are the main laboratory markers of AIRD and ALD, being among the diagnostic criteria for diseases. The ‘gold standard’ and primary screening method for determining ANA in serum is the indirect immunofluorescence assay (IFA). Antigen-specific solid phase assays methods are used as confirmatory tests. Standardization of the ANA determination contributes for reducing the intra- and inter-laboratory variability of the results, helps to optimize the interaction between laboratory specialists and clinicians in matters of prescribing and clinical interpretation of ANA tests. Solving the problem of ANA detection standardization is important because of the growing number of laboratories performing these tests and an increased referring for this investigation from rheumatologists and another medical specialist.

A Retrospective Study on Antinuclear Antibody Patterns in Systemic Lupus Erythematosus Patients and Its Correlation With Serological Markers.

Systemic lupus erythematosus (SLE) is a multisystem autoimmune disease. Detection of antinuclear antibodies (ANAs) aids in the diagnosis of SLE. The indirect immunofluorescence (IIF) assay is often used a routine screening test for the detection of ANA. The pathogenic role and significance of various patterns produced in IIF is yet to be explored. This study aimed to detect ANA patterns generated by IIF and correlate these patterns with specific antibodies detected by line immunoassay. We also investigated the significance of each ANA pattern and its association with specific serological SLE markers, such as complement molecules, anti-dsDNA, antiphospholipid antibody, and C-reactive protein (CRP), along with associations with direct Coombs test (DCT). We conducted a retrospective study that included 204 patients newly diagnosed with SLE according to the European Alliance of Associations for Rheumatology/American College of Rheumatology (EULAR/ACR) criteria. The detection and pattern determination of ANA was performed by IIF using HEp-20-10. Furthermore, line immunoassay was performed, and the antibody profile of each sample was obtained.Other immunodiagnostic markers were analyzed, including C3, C4, anti-dsDNA, antiphospholipid antibodies (anti-cardiolipin antibodies, anti-beta-2-glycoprotein I, and lupus anticoagulant), CRP, and DCT. Of the 204 samples, the most frequent ANA pattern observed was nucleus speckled (52.9%), followed by nucleus homogenous (27.5%), mixed (13.7%), and cytoplasm speckled (5.9%). The nucleus homogenous pattern showed the most pathogenic immune profile due to its close association with markers of disease activity, namely, high anti-dsDNA titer, low C3 level, and DCT positivity. Conclusion: This study showed that the most commonpattern associated with SLE is nucleus speckled, followed by the nucleus homogenous pattern. Based on associations with specific serological markers, the nucleus homogenous pattern may be linked to a high disease activity in SLE.

THE SPECTRUM OF IMMUNORESISTANCE DISORDERS AND INFLUENCE OF CYTOTOXICITY AND PATHOGENICITY OF DAMP FRACTIONS ON THE POST-COVID-19 SYNDROME FORMATION AND THE MAIN DОISEASE COMPLICATION IN PATIENTS WITH URGENT SURGICAL PATHOLOGY

Summary. The infectious nature of COVID-19 leads to complications in the immune system. It is relevant to determine the long-term consequences of COVID-19, which can be triggers for the urgent surgical pathology’s development of the thoracic and abdominal organs in patients with certain risk factors associated with characteristic disorders of immunometabolic reactions.
 The aim. To determine the spectrum of immune resistance disorders and to find out the effect of cytotoxicity and pathogenicity of DAMP fractions on the post-COVID-19 syndrome formation and complications of the main disease in patients with urgent surgical pathology.
 Materials and methods. Patients (n = 143) with urgent cardiovascular and abdominal surgical pathology that arose against the background of post-COVID-19 syndrome were examined. We used: spectrophotometry (content of DAMP fractions), immunoturbimetry (concentration of C3 and C4 complement components); luminescent microscopy (phagocytic activity of neutrophils); light microscopy (spontaneous activity of NADPH-oxidase systems of neutrophils); flow cytometry (expression of T lymphocyte activation markers); immunofluorescence (determination of antinuclear antibodies).
 Results. It was found that the predominance of certain DAMP fractions is the cause of the immunopathological patterns formation in the development of post-COVID-19 syndrome. In patients with urgent surgical pathology and the predominance of the DAMP260 oligonucleotide fraction, significant changes in the hemogram, failure of endocytosis and incomplete phagocytosis of neutrophils, increased expression of the T lymphocytes late activation marker (CD3+HLA-DR+) were found. In all examined patients, a high frequency of autoantibodies to nuclear structures (ANA) was found, which depended on the content of DAMP fractions. The maximum repertoire of antibodies was found in patients with the oligonucleotide fraction (DAMP260) predominance.
 Conclusions. In patients with urgent surgical pathology, during the formation of post-COVID-19 syndrome, the chronic immunoinflammatory reaction formation occurs against the background of the activation cascades development of innate and adaptive immunity factors.

Diagnostic value of screening methods for the determination of antinuclear antibodies using indirect immunofluorescence on HEp-2 cells and enzyme immunoassay in autoimmune liver diseases.

Antinuclear antibodies (ANA) are a heterogeneous group of autoantibodies that react with various components of the cell nucleus and cytoplasm. ANA is the main serological marker for autoimmune liver disease (AILD). The aim of the study was to compare the diagnostic value of two methods of screening for the determination of ANA (indirect immunofluorescence reaction on HEp-2 cells (IIF -HEp-2) and enzyme-linked immunosorbent assay (ELISA) in the sera of AILD patients. The sera of 118 patients with AILD (51 with autoimmune hepatitis - AIH, 19 with primary biliary cholangitis - PBC, 48 with overlapping syndrome - OVERLAP), 30 patients with non-alcoholic fatty liver disease (NAFLD) and 30 healthy donors (HD) were studied. Determination of ANA by the IIF-HEp-2 method was carried out by visual assessment of samples under an AXIOSKOP 40 microscope, by ELISA - on an Alegria automatic analyzer. A weak degree of agreement between the positive and negative results of the ANA screening study using IIF-HEp-2 and ELISA (Cohen's kappa coefficient æ=0.4) was noted. Screening determination of ANA in patients with AILD by the IIF-HEp-2 method was distinguished by greater diagnostic sensitivity (DS) (68.6%) and a lower frequency of false negative results (31.4%) compared with ELISA (35.6% and 64.4 % respectively, p<0.05). The overall diagnostic specificity (DS) of the ANA study in IIF-HEp-2 was lower than with ELISA (66.7% and 86.7%, respectively, p<0.05). Both screening methods for determining ANA (IIF-HEp-2 and ELISA) were useful for diagnosing AILD (positive likelihood ratio - LR+: 2.1 and 2.6, respectively). In terms of the negative likelihood ratio (LR-), screening for ANA by the IIF-HEp-2 method, in contrast to ELISA, served as a "useful" test to exclude the diagnosis of AILD (0.5 and 0.8, respectively). The determination of ANA using IIF-HEp-2 is the most sensitive and "useful" screening test for the diagnosis of AILD, and ELISA is classified as a less "useful" screening method due to low diagnostic sensitivity and a high false-negative rate.

Assessment of antinuclear antibodies (ANA): National recommendations on behalf of the Croatian society of medical biochemistry and laboratory medicine.

Antinuclear antibodies (ANA) represent a family of autoantibodies targeting ubiquitous cellular constituents and are a hallmark of systemic inflammatory autoimmune rheumatic diseases named connective tissue diseases (CTD). The gold standard method for ANA determination is indirect immunofluorescence (IIF) on the human laryngeal epidermoid carcinoma cell line type 2 substrate (HEp-2), but with increasing demand for ANA testing, novel methods eased for automation emerged, which allows testing by staff less experienced in this specific field of laboratory diagnostic. In 2016 The working group (WG) for laboratory diagnostics of autoimmune diseases as part of the Committee for the Scientific Professional Development of the Croatian Society of Medical Biochemistry and Laboratory Medicine (CSMBLM) published the data of a survey regarding general practice in laboratory diagnostics of autoimmune diseases in Croatia. Results indicated high diversity in the performance of autoantibody testing as well as reporting of the results and indicated the need of creating recommendations for the assessment of ANA that would help harmonize diagnostics of systemic autoimmune rheumatic diseases in Croatia. This document encompasses twenty-seven recommendations for ANA testing created concerning indications for ANA testing, preanalytical, analytical, and postanalytical issues, including rational algorithm and quality control assurance. These recommendations are based on the relevant international recommendations and guidelines for the assessment of ANA testing and relevant literature search and should help to harmonize the approach in ANA testing and clarify differences in interpretation of the results obtained using different methods of determination.

Prevalence of autoantibodies to cellular cytoplasmic and mitotic antigens in routine antinuclear antibody reporting: Implementation of international consensus on antinuclear antibodies patterns guidelines

Background: The international consensus on antinuclear antibody (ANA) patterns (ICAP) guidelines recommends that non-nuclear patterns should be reported as positive for ANA. Therefore, a prospective study was planned to investigate the frequency of cytoplasmic and mitotic ANA patterns in routine reporting in a high-throughput lab in a tertiary referral center in Northern India. Materials and Methods: Whole blood samples received were centrifuged and serum was separated within 4 hours of collection. Sera were subjected to indirect immunofluorescence (IIF) assay on HEp-2 cells for ANA determination in a dilution of 1:40, per standardized protocol in our laboratory. IIF staining patterns and intensity were evaluated and the results interpreted. Results: Over a 3-month period, of the 3796 serum samples received for ANA testing, 81 (4.9%) samples were positive for cytoplasmic, 7 (0.4%) for mitotic, and 2 (0.1%) for DNA topoisomerase I (Topo I)-like pattern. The most frequent cytoplasmic fluorescence pattern was dense fine speckled (2%) and the most frequent mitotic pattern was NuMA-like (0.2%). The prevalence of total positive ANA in our study population was found out to be 43.6%. Conclusions: Implementation of ICAP guidelines into routine ANA reporting helped us in detecting important cytoplasmic antibodies such as antimitochondrial antibodies, anti-ribosomal P protein antibodies, anti-Jo-1antibodies, anti-Topo I antibodies and allowed us to introduce more specific and confirmatory tests for the same. The new format for ANA reporting also encouraged clinician-laboratory crosstalk.

Debut de lupus eritematoso sistémico en paciente masculino

Introducción: El lupus eritematoso sistémico (LES) es una enfermedad multisistémica inflamatoria crónica y autoinmune que produce anticuerpos antinucleares, misma que es capaz de afectar cualquier órgano, su prevalencia es mayor en las mujeres sobre todo en edad fértil que en hombres con una relación 9:1. Su presentación clínica es insidiosa y variable desde lesiones cutáneas típicas como el eritema malar hasta alteraciones sistémicas cardíacas, pulmonares o renales. Su diagnóstico se enfoca en la clínica más la determinación de Anticuerpos antinucleares y de varios anticuerpos y exámenes de laboratorio, así como de imagen para determinar la presencia de otras complicaciones.&#x0D; Objetivo: Aplicar los conocimientos teóricos del Lupus eritematoso sistémico para analizar el caso clínico del debut de un paciente masculino de 60 años.&#x0D; Material y métodos: Estudio descriptivo retrospectivo, presentación de caso clínico.&#x0D; Resultados: Se presenta un caso clínico de un paciente masculino de 60 años de edad que presentó un síndrome coronario agudo, acompañado de lesiones dérmicas y pérdida de peso que después de varias sospechas diagnósticas se confirmó el diagnóstico de LES.&#x0D; Conclusiones: A pesar de la prevalencia más frecuente del LES en mujeres jóvenes; esta patología puede presentarse en varones mayores de 50 años y su sospecha diagnóstica debe estar presente en pacientes que presenten características clínicas insidiosas de LES ya que su diagnóstico temprano puede evitar las complicaciones sistémicas que esta patología puede desarrollar si no se da el tratamiento adecuado

P781 Prevalence of antinuclear antibodies in inflammatory bowel disease and seroconversion after biological therapy

Abstract Background Higher levels of antinuclear antibodies (ANA) are detected in healthy populations with a prevalence of 6%. In autoimmune disorders it reaches 50–80%. In patients under TNF, the prevalence of positive ANA observed was 40% and occasionally may induce lupus like syndrome (LLS). However, there is a lack of evidence about the global prevalence in inflammatory bowel disease (IBD) and insufficient data about seroconversion of ANA after the beginning of biological therapy. Methods A unicentric transversal study of 879 IBD patients was designed. ANA determination was performed by immunofluorescence assay. IBD patients were classified according to the biological therapy. Presence and of ANA and titres were collected previously to the beginning of anti-TNF therapy. LLS symptoms were collected in a retrospective way. Results A valid result of ANA was obtained in 862 patients. Patients with any autoimmune disease which involve positive ANA were excluded. Positive ANA was reported in 13.62% of IBD patients. The prevalence of ANA was 28.67% in those patients under biological therapy (p &amp;lt; 0.001), especially if the biological treatment was anti-TNF. No significative difference was observed in those patients receiving ustekinumab or vedolizumab therapy. Positive ANA was observed in 4.67% for combined treatment with immunomodulator and biological therapy. In contrast, if the patient received only biological therapy, positive ANA was collected in 24 % (p &amp;lt; 0.001). A reduction in albumin levels and higher levels of IgG and IgM were related to positive ANA. It is important to highlight that a percentage of 53.45% positive ANA had never received biological therapy in the previous 5 years; that means, a 7.28% prevalence in the global cohort. A 48.15% of the patients had ANAs previous to the beginning of biological therapy. ANAs seroconversion was observed in 73.08% of the patients after the treatment. Conclusion IBD patients have a 13.62% prevalence of positive ANA; higher than prevalence in healthy population. The presence of ANA is frequently associated in patients under anti-TNF treatment and decreases with immunomodulator therapy associated to anti-TNF. It is unknown if positive ANA in IBD patients could be conditioned to the future development of other autoimmune disorders. Prospective studies are necessary to clarify these aspects.

Frequency of positive ANCA test in a population with clinical symptoms suggestive of autoimmune disease and the interference of ANA in its interpretation

BackgroundAntibodies against neutrophil cytoplasm (ANCA) are associated with vasculitis. There are different methods to determine their presence. The interference of antinuclear antibodies (ANA) in the differentiation between P-ANCA and C-ANCA patterns has been described. ObjectiveTo determine the frequency of ANCA in a population with manifestations of autoimmune disease, and evaluate the interference of ANA in its interpretation. Materials and methodsRetrospective, descriptive nonexperimental cross-sectional study, including 3330 data. The presumptive diagnosis was autoimmune disease and a test for ANCA was requested. The ANCA and ANA determinations were made by indirect immunofluorescence, L-ANCA® and CytoBead® ANCA. Anti-proteinase 3 and anti-myeloperoxidase were detected by ELISA and CytoBead® ANCA. ResultsANCAs were positive in 10.21% and 12.64% of those positive for ANCA were positive for ANA. The inter-rater agreement statistic (Kappa) for anti-PR3 between CytoBead ANCA and ELISA was 100% (K=1.00; P<.05) and the agreement between anti-myeloperoxidase by ELISA and CytoBead® ANCA was high (K=0.94; P<.05). 30% of those with ANCAs had a diagnosis of a type of vasculitis; 20% of them had an autoimmune disease. ConclusionsThe results suggest an overestimated request for ANCAs as a diagnostic aid in primary care which was not addressed. For an adequate evaluation of ANCAs, the indirect immunofluorescence technique should be implemented for the control and confirmation with the determination of specific antigens for anti-proteinase 3 and anti-myeloperoxidase in any of the confirmatory assays. The high concordance shown by ANCA CytoBeads makes us consider the use of this alternative for the determination of ANCAs and the confirmation. Given the interference of ANAs, the ANA test by IFI in the presence of positive P-ANCA results is recommended in order to minimise “false positives”.

Diagnóstico y manejo de la enfermedad pulmonar intersticial de la esclerosis sistémica en condiciones sanitarias desafiantes

A 51-year-old woman presented with swelling in both hands and a 3-months history of triphasic Raynaud phenomenon. She denied cough and dyspnea. The physical examination was notable for swollen hands, facial telangiectasia and salt and pepper lesions. She had no skin fibrosis. The determination of antinuclear antibodies and antitopoisomerase were positive. A spirometry demonstrated a forced vital capacity of 86% of the predicted. High resolution chest computed tomography revealed bilateral, basal and subpleural ground glass opacities. These findings were consistent with the diagnosis of Scleroderma-Related Interstitial Lung Disease. Moreover, due to the absence of skin fibrosis, a diagnosis of systemic sclerosis sine scleroderma was made. Infusions of endovenous cyclophosphamide were indicated at 4-week intervals, followed by oral azathioprine. This case showed that using PFT as the single screening method for SSc-ILD may cause clinicians to miss a significant number of patients and that the absence of pulmonary symptoms does not exclude lung disease in patients with normal FVC.

ANA as an entry criterion for the classification of SLE.

Systemic lupus erythematosus (SLE) is a prototypic autoimmune disease with highly variable clinical and immunological manifestations. In the classification of patients with this condition, the presence of an antinuclear antibody (ANA) is an important element, with new criteria from the American College of Rheumatology and European League against Rheumatism positioning ANA positivity by an immunofluorescence assay on HEp2-cells (HEp2-IFA) or by an equivalent solid phase assay as the entry criterion. This positioning is based on assumptions about the frequency of ANA positivity in SLE as well as the reliability of the assays. Studies indicate that these assumptions are still a matter of uncertainty since both types of assay show considerable variability and patients with SLE may display negative results in ANA testing. These findings suggest caution in positioning ANA positivity as an entry criterion for classification and point to the value of alternative serological approaches for ANA determinations.

Uncommon patterns of antinuclear antibodies recognizing mitotic spindle apparatus antigens and clinical associations.

Antinuclear antibodies (ANA) are key biomarkers in the evaluation of rheumatic diseases. The prevalence and clinical significance of uncommon or rare patterns, particularly those directed at the mitotic spindle apparatus (MSA), are not well understood. We aimed to investigate the prevalence and clinical significance of anti-MSA patterns in a Colombian population.During 2013 and 2014, 113,491 consecutive determinations of ANA were studied for the presence of uncommon patterns. Clinical and laboratory data of anti-MSA positive patients were retrospectively collected and analyzed.Of the 113,491 patients tested, 60,501 (53%) were positive for ANA, of which 834 (1.3%) were positive for uncommon/rare patterns of ANA (anti-MSA in 592 cases). Of these 592 cases, complete data were available in 329 patients, of whom 116 had an established diagnosis. Anti-MSA antibodies were the only ANA positive test in 81% patients. At least one fine reactivity was identified in 19/116 (16.3%) of ANA-positive patients, of which anti-Ro was the most prevalent (18/116, 15.5%).The most frequent patterns were nuclear mitotic apparatus (NuMA) (56%) and MSA-2 (25%). The NuMA pattern had the highest ANA titers: mean 320 (range 80-2560) and behaved as monospecific antibodies. The most frequent systemic autoimmune diseases were Sjögren syndrome (SS) (18.1%), rheumatoid arthritis (RA) (13.8%), and systemic lupus erythematosus (SLE) (11%). Undifferentiated connective tissue disease (UCTD) was associated with the centrosome (P < .001), NuMA (P < .02) and MSA-2 (P < .45) patterns. Chronic idiopathic urticaria (CIU) was associated with the NuMA pattern (P < .02) and sensorineural hearing loss (SNHL) was associated with the MSA-2 (P < .001), centrosome (P < .68) and CENP-F (P < .38) patterns, previously unreported findings. Malignancies were found in 8 patients (50% were papillary thyroid cancer).In a large cohort of ANA determinations, uncommon patterns were found in around 1% of cases. The most frequent anti-MSA patterns found were NuMA and MSA-2. More than 50% of patients with anti-MSA had an associated CTD, mainly SS, RA and SLE, and anti-MSA behaved as monospecific antibodies. Other entities of presumed autoimmune origin, like CIU and SNHL, might be associated with these patterns.

Assay variation in the detection of antinuclear antibodies in the sera of patients with established SLE

ObjectiveThe expression of antinuclear antibodies (ANA) is considered almost constant in systemic lupus erythematosus (SLE), although recent experience has suggested that many subjects with SLE considered for clinical trials are...

Modern look at the problems of investigation of antinuclear antibodies in systemic lupus erythematosus (literature review).

In the review, topical aspects of the study of antinuclear antibodies (ANA) in systemic lupus erythematosus (SLE) are considered. ANA is the main serological marker of SLE. In the sera of patients with SLE, antibodies to DNA, histones, nucleosomes, extractable nuclear antigens (Sm, U1 ribonucleoprotein, Ro / SSA, La / SSB, ribosomal protein P), nucleolar antigens and other cellular structures are detected. The ANA study using indirect immunofluorescence on HEp-2 cells (IIF-HEp-2) is recommended as a standard screening test for the diagnosis of SLE. The use of automated systems for the interpretation of cellular fluorescent tests contributes to the standardization and improvement of the reproducibility of the IIF. A new international nomenclature of types of nuclear, nucleolar (nucleolar), cytoplasmic and mitotic luminescence of ANA in IIF-HEp-2, including 28 variants of anticell ("Anti-cell" - AC) patterns was developed. In the practice of clinical diagnostic laboratories, high-performance automated methods for the determination of ANA based on ELISA, immunoblot, fluorescent, chemiluminescent and multiplex immunoassay are widely used. New mono- and multiplex methods of solid-phase analysis are expediently used as confirmatory reflex tests for the detection of varieties of antigen-specific ANA in patients with SLE with positive results of IIF-HEp-2. Identification of ANA profiles using multiplex technologies is a useful tool for implementing a personalized approach to diagnosis, evaluation of activity, prognosis, clinical and immunological subtypes, and the effectiveness of SLE therapy. The need for an ANA study not only to confirm the diagnosis of SLE, but also to identify the disease in the early and preclinical stages with the intention to prevent the development of the pathological process is discussed. Detection of monospecific anti-DFS70 antibodies allows to exclude the diagnosis of SLE inANA IIF-HEp-2 positive subjects. Presented is a modern algorithm for testing ANA with SLE.

Rheumatic Disease Autoantibodies in Patients with Autoimmune Thyroid Diseases

Background: Patients with autoimmune thyroid diseases (ATD) such as Graves’ disease (GD) and Hashimoto thyroiditis (HT) may have non-organ specific autoantibodies such as antinuclear antibodies (ANA) and rheumatoid factor (RF). Aim: To study the prevalence of rheumatic autoantibodies in a group of ATD patients without known rheumatic diseases and to evaluate its association with the patients’ epidemiological and treatment profiles. To follow positive non-organ specific autoantibody-positive ATD individuals to investigate whether they will develop a rheumatic disorder. Methods: A sample of 154 ATD patients (70 HT and 84 GD; mean age 45.3 ± 14.2) had determination of ANA by immunofluorescence, using hep-2 cells as substrate, extractable nuclear antigen profile by ELISA kits and RF by latex agglutination. Epidemiological and treatment profiles were obtained through chart review. These patients were followed for the mean period of 5 years, between 2010 and 2015. Results: Positive ANA was found in 17.5% (27/154) of the patients: anti-Ro/SS-A in 4/154 (2.5%); anti-RNP in 4/154 (2.5%), and anti-La/SS-B in 3/154 (1.9%). None had anti-Sm antibodies. RF was detected in 12/154 (7.7%) of ATD patients and was more common in older individuals (p = 0.007). There was a positive association between the presence of RF and ANA (p = 0.03; OR 3.89; 95% CI 1.1–13.3). None of the patients with positive autoantibodies developed clinical rheumatic diseases during the period of observation. Conclusion: We found rheumatic autoantibodies in 17.5% of ATD patients without rheumatic diseases. None of them were associated with the appearance of clinical rheumatic disorder during the period of 5 years.

Recent Approaches To Optimize Laboratory Assessment of Antinuclear Antibodies.

The presence of antinuclear antibodies (ANAs) is a hallmark of a number of systemic autoimmune rheumatic diseases, and testing is usually performed as part of the initial diagnostic workup when suspicion of an underlying autoimmune disorder is high. The indirect immunofluorescence antibody (IFA) technique is the preferred method for detecting ANAs, as it demonstrates binding to specific intracellular structures within the cells, resulting in a number of staining patterns that are usually categorized based on the cellular components recognized and the degree of binding, as reflected by the fluorescence intensity or titer. As a screening tool, the ANA patterns can guide confirmatory testing useful in elucidating a specific clinical diagnosis or prognosis. However, routine use of ANA IFA testing as a global screening test is hampered by its labor-intensiveness, subjectivity, and limited diagnostic specificity, among other factors. This review focuses on current efforts to standardize the nomenclature of ANA patterns and on alternative methods for ANA determination, as well as on recent advances in image-based computer algorithms to automate IFA testing in clinical laboratories.

Biosensor for total antinuclear antibody determination at the point-of-care

Antinuclear antibodies (ANA) are important in diagnosis and follow-up of patients with autoimmune conditions. The current increase in ANA requests is driven by broadening the use of ANA from a test for lupus to a test for diverse autoimmune diseases, but the standard method is protracted, cumbersome and prone to error. We describe an electrochemical method for quantifying total ANA for use as a point-of-care diagnostic aid. In this technology the target autoantigens are derived from a protein/nucleoprotein mixture prepared from an inexpensive source and adsorbed to a porous membrane with high protein binding capacity. Serum is slowly drawn through the membrane comprising the high density autoantigen mixture to induce rapid binding of patient autoantibodies. After rinsing, peroxidase-conjugated anti-IgG is drawn through the membrane followed by rinsing, insertion of an electrode assembly, and addition of the enzyme substrate. Substrate peroxidation is measured by microamperage-level current accompanying electrochemical reduction of the intermediate product. Values are comparable to a standard ANA test but require a total processing time of ~20min. This method has the promise to greatly expand ANA testing in clinical settings for initial patient assessment of autoimmune disease.

Profile of autoantibodies in systemic sclerosis

Objective: to estimate frequency of autoantibodies in a cohort of Russian patients with systemic sclerosis (SS) and to investigate clinical associations of these autoantibodies. Subjects and methods. In 2012 to 2015, the investigation enrolled 300 patients (58 men and 242 women) who fulfilled the American College of Rheumatology/European League Against Rheumatism (ACR/EULAR) SS criteria. All patients underwent immunological examination including determination of antinuclear antibodies, anti-topoisomerase I (anti-Scl-70) antibodies, anti-centromere antibodies (ACA), anti-U1-ribonucleoprotein antibodies (anti-U1 RNP), and anti-RNP III antibodies. Results and discussion. The vast majority of patients were middle-aged females with moderate disease duration. There was a preponderance of patients with limited cutaneous form (55.3%); 37.4% patients had diffuse SS; 5.9% had overlap syndrome; and <1% had visceral and juvenile SS. The vast majority (83.8%) of patients were found to have antinuclear factor. Among the SS-associated antibodies, anti-Scl-70 were most common and were revealed in approximately one-half of the patients. ACA was present in only 44 (14.6%) patients. There was a combination of positivity for ACA and anti-Scl-70 antibodies in 3 patients with limited cutaneous SS, including one with an early form of the disease. 26 patients had anti-U1 RNP antibodies. Among them, there was a preponderance of patients with limited cutaneous SS and overlap syndrome. Anti-RNP III antibodies were found in 5.5% of cases, mainly in those with limited cutaneous SS; these were observed in one patient with diffuse SS and interstitial lung disease. No kidney injury was seen in this patient group. Conclusion. The characteristics of the Russian cohort are the preponderance of the limited cutaneous SS and the frequent detection of anti-Scl-70 antibodies in both diffuse and limited cutaneous SS; no correlation of anti-Scl-70 antibodies with rapid progression of the pathological process, with kidney disease.