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Background/aim: Pneumonitis during durvalumab consolidation therapy after chemoradiotherapy (CRT) is a major cause of treatment discontinuation. Although previous studies have revealed an association between antinuclear antibody (ANA) positivity and the safety and efficacy of immune checkpoint inhibitors in advanced non-small cell lung cancer (NSCLC), there are no reports on durvalumab consolidation therapy. This study investigated the safety and efficacy of durvalumab after CRT in ANA-positive patients., Patients and Methods: We retrospectively reviewed patients with unresectable NSCLC treated with durvalumab after CRT between August 2018 and July 2022 at our institution. We evaluated the association among ANA positivity, treatment-related adverse events (AEs), and survival outcomes., Results: Overall, 80 patients were enrolled, 39 of whom were ANA-positive. Although there were no significant differences in the incidence of each AE of any grade, ANA-positive patients tended to have a higher frequency of pneumonitis of grade 3 to 5 than ANA-negative patients (12.8% vs. 2.4%, p=0.104). ANA-positive patients had a significantly shorter median progression-free survival (PFS) and overall survival (OS) than ANA-negative patients [14.9 months vs. not reached (NR), p=0.005; NR vs. NR, p=0.013]. Multivariate analysis revealed that ANA positivity was an independent predictor of shorter PFS (HR=2.23; 95% CI=1.16-4.29; p=0.016) and OS (HR=2.28; 95% CI=1.01-5.12; p=0.046)., Conclusion: ANA-positive patients receiving durvalumab after CRT tended to have a higher frequency of severe pneumonitis and significantly worse PFS and OS compared with ANA-negative patients., (Copyright © 2024 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Background: Guillain-Barré syndrome (GBS), an acquired immune-mediated autoimmune disorder affecting the peripheral nervous system (PNS), is associated with autoimmunity. The presence of autoantibodies in the blood is an important feature of autoimmune diseases. Herein, we explored the distribution characteristics of the antinuclear antibodies (ANAs) in GBS and the correlation between ANAs and disease severity., Materials and Methods: We retrospectively analyzed the clinical data of 170 GBS patients. According to ANAs, GBS patients were divided into ANAs positive and negative groups. The clinical characteristics of these two groups were compared. The distribution difference was also compared between male and female GBS patients. In addition, all enrolled patients were divided into more severe group and milder group according to whether the Hughes score at nadir ≥ 3 or not. Gender, age, and ANAs were compared between the two groups., Results: In this study, the positive rate of ANAs was 27.1 % in 170 GBS patients, among which anti-SSA-52/Ro52 antibody and antimitochondrial antibody M2 made up the largest proportion. In the ANAs positive group, GBS patients had longer days of hospitalization, more respiratory function involvement, and higher level of CSF IgG than the ANAs negative group. Compared to the ANAs negative group, Medical Research Council (MRC) scores on admission and at nadir were lower, and Hughes functional Grading Scale (HFGS) scores on admission and at nadir were higher in GBS patients with ANAs positive group. Erasmus GBS Respiratory Insufficiency Score (EGRIS) in ANAs positive GBS patients group was significantly higher than ANAs negative group. Gender had no effects on the distribution of ANAs in GBS patients. Moreover, we found that the anti-SSA-60 antibodies and age were positively correlated with GBS severity. In addition, in the anti-SSA-60 antibody positive group, GBS patients had longer days of hospitalization, more respiratory function involvement, higher HFGS scores on admission/at nadir, and lower MRC scores at nadir compared with the anti-SSA-60 antibody negative group., Conclusion: Anti-SSA-52/Ro52 antibody and antimitochondrial antibody M2 were the most common ANAs in GBS patients. Anti-SSA-60 antibodies and age positively correlated with GBS severity. Positive anti-SSA-60 antibodies and age were independent predictors of GBS patient severity., Competing Interests: Declaration of Competing Interest The authors declare no conflicts of interest., (Copyright © 2024. Published by Elsevier B.V.)

Background: Immunity play a pivotal role in the risk of ischemic stroke, and studies have also shown a relationship between ischemic stroke and autoimmune diseases. In light of this we conducted a prospective cohort study to elucidate the impact of antiphospholipid antibodies (aPLs), antinuclear antibodies (ANA), and anti-extractable nuclear antigen autoantibodies (anti-ENA) on the prognosis of ischemic stroke., Methods: 245 stroke patients were recruited in this single-center study and followed up with for 3 years. Autoantibodies, including aPLs (ACA, anti-β2GPI, LA), ANA and anti-ENA were evaluated in recurrent ischemic stroke (RIS) and nonrecurrent ischemic stroke (nonRIS). Stroke severity was judged using the National Institutes of Health Stroke Scale (NIHSS). For preventive treatment, 42 IS patients with positive aPLs + ANA/anti-ENA were randomized 1:1 into a hydroxychloroquine (HCQ) treatment group and a control group, and the prognoses were compared., Results: The positive rate of ACA IgG (p = 0.018), anti-β2GPI IgG (p = 0.047), LA (p = 0.023), and aPLs + ANA/anti-ENA (p = 0.000) were significantly higher in patients with RIS compared to patients with nonRIS, and aPLs + ANA/anti-ENA (HR2.31, 95 % CI1.02-5.25, p = 0.046) and hypertension (HR2.50, 95 % CI1.17-5.35, p = 0.018) were the independent risk factors of recurrence. There were differences in NIHSS at month 36 between those positive and negative for aPLs + ANA/anti-ENA (p = 0.001, Eta 2 = 0.052), anti-ENA (p = 0.016, Eta 2 = 0.030), ANA (p = 0.035, Eta 2 = 0.022), and LA (p = 0.016, Eta 2 = 0.028). Furthermore, the recurrence rate of the HCQ treatment group was lower than that of the control group (p = 0.024)., Conclusions: Co-positivity of aPLs and ANA/anti-ENA is an independent risk factor for RIS. However, HCQ therapy may reduce the recurrence rate of IS for these patients., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

The relevance of the problem of immunoinflammatory rheumatic diseases (IIRD) for modern medicine is determined by their high prevalence in the population, the difficulty of early diagnosis, the rapid development of disability and poor life prognosis. Recent data on the significance of anti-DFS70 have opened up new possibilities for optimizing the step-by-step diagnosis of IIRD. The detection of these antibodies can help in the interpretation of a positive result for antinuclear antibodies (ANA) by indirect immunofluorescence assay on HEp-2 cells (IIFA-HEp-2) in the absence of autoantibodies specific for IIRD. Detection of anti-DFS70 in antinuclear factor (ANF) seropositive patients without clinical and/or serological markers characteristic of a certain disease from the IIRD group can be considered as a potential marker that excludes this group of diseases., (© 2024. Pleiades Publishing, Ltd.)

The pathophysiology of dengue may be influenced by antibodies released during infection. Several autoimmune diseases are accompanied by antinuclear antibodies (ANAs) but 8-10% of the general population have positive ANA tests. To test the hypothesis that an ANA-positive test indicates an immune dysregulated state that modifies the risk for certain clinical disorders in people with or without an autoimmune disease, we examined the various ANA profiles and their relationships to various autoimmune disorders, as well as the severity of these relationships, in patients infected with dengue fever. Enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR) methods were used. Indirect immunofluorescence assay (IIFA) and line immunoassay (LIA) were performed to detect and differentiate the ANAs among dengue infected patients. Out of 135 dengue virus-positive patients, 94.07% were positive by ELISA and 5.93% positive by RT-PCR method. ANAs by IIFA and LIA were detected in 54.8% and 18.5% of the dengue positive patients, respectively, and 10.3% and 7.1% of the 126 dengue negative patients, respectively. This study showed that dengue was associated with an increased risk of autoimmune myositis and mixed connective tissue disease (MCTD), a rare complication of dengue. The risk of other autoimmune diseases did not seem to increase after DENV infection.

Background: The anti-Nucleolar Organizer Region 90 antibodies (NOR90) are rare antinuclear antibodies (ANA) reported in systemic sclerosis (SSc). Especially due to low prevalence, the clinical relevance of NOR90 in SSc remains uncertain., Objectives: To analyze the clinical associations of NOR90 in patients with SSc in a multicentric cohort., Methods: Post-hoc, cross-sectional study of prospectively collected data from the European Scleroderma Trials and Research (EUSTAR) database, with additional information on NOR90. Further, we performed a systematic literature search, using the terms "systemic sclerosis" and "NOR90" across three databases: Medline via PubMed, Scopus, and Thomson Reuters' Web of Science Core Collection, from inception to November 1st, 2023., Results: Overall, 1318 patients with SSc were included (mean age 58.3 ± 13.7 years, 81.3 % female), of whom 44 (3.3 %) were positive for NOR90. Of these, 32 were also positive for one of the SSc-criteria antibodies: 9/44 (20.5 %) for anti-topoisomerase I, 18/42 (42.9 %) for anti-centromere, and 5/40 (12.5 %) for anti-RNA polymerase III. NOR90-positive patients were more frequently female, had lower modified Rodnan skin score (mRSS), and lower prevalence of upper and lower gastrointestinal (GI) symptoms compared to NOR90-negative patients. In multivariable analysis, NOR90 remained significantly associated with lower mRSS and less frequent GI symptoms. The literature search identified 17 articles, including a total number of 87 NOR90-positive out of 3357 SSc patients, corresponding to an overall prevalence of 2.6 %., Conclusion: To our best knowledge, this is the largest SSc cohort tested for NOR90 to date, confirming the NOR90 prevalence in SSc patients is around 3 %., Competing Interests: Declaration of competing interest DIMA Alina: Nothing to disclose. VONK Madelon: Consultancy relationship with and/or research funding from and/or speaker for the following companies in the area of potential treatments for systemic sclerosis and its complications in the last three calendar years: Boehringer Ingelheim, Ferrer, Galapagos, GSK, Janssen, MSD and Novartis. GARAIMAN Alexandru: Nothing to disclose. KERSTEN E Brigit: Nothing to disclose. BECVAR Radim: Nothing to disclose. TOMCIK Michal: Nothing to disclose. HOFFMANN-VOLD Anna-Maria: Speakers bureau: Boehringer Ingelheim, Jannsen, Medscape, Merck Sharp & Dohme and Roche; Consultant: ARXX, Boehringer Ingelheim, Genentech, Jannsen, Medscape, Merck Sharp & Dohme and Roche; Grant/research support: Boehringer Ingelheim, Jannsen CASTELLVI Ivan: Speakers’ bureau: Boehringer, Janssen-Cilag, BMS, Roche, UCB; Consultant: Boehringer, Janssen-Cilag, Topandur. TANDAIPAN JAIME Jose Luis: Nothing to disclose. BRZOSKO Marek: Nothing to disclose. MILCHERT Marcin: Nothing to disclose. KRASOWSKA Dorota: Speakers’ bureau: Novartis, Abbvie, Elli Lilly; Consultant: Boehringer Ingelheim, Novartis, Janssen. MICHALSKA-JAKUBUS Małgorzata: Nothing to disclose. AIRO Paolo: Consulting fees, honoraria for lectures, presentations, or educational events, support for attending meetings and/or travel from: Bristol Myers Squibb, Bohringer Ingelheim, Roche, Novartis, CSL Behring Janssen- Cilag MATUCCI-CERINIC Marco: Speakers’ bureau: Biogen, Sandoz, Boehringer, Lilly, Galapagos; Grant/research support: MSD, Janssen; BRUNI Cosimo: Consultant: Boehringer Ingelheim; Grant/research support: Gruppo Italiano Lotta alla Sclerodermia (GILS), European Scleroderma Trials and Research Group (EUSTAR), Foundation for research in Rheumatology (FOREUM), Scleroderma Clinical Trials Consortium (SCTC); Novartis Foundation for medical-biological research, EMDO Foundation; Educational grants: AbbVie, Wellcome Trust. IUDICI Michele: Nothing to disclose. DISTLER Jorg: Nothing to disclose. GHEORGHIU Ana-Maria: Speakers’ bureau: Sandoz, Boehringer Ingelheim, Ewopharma; Consultant: Boehringer Ingelheim. POORMOGHIM Hadi: Nothing to disclose. MOTTA Francesca: FM had consulting fees from Thermo Fisher. DE SANTIS Maria: Nothing to disclose. PARVU Magda: Nothing to disclose. DISTLER Oliver: OD has/had consultancy relationship with and/or has received research funding from and/or has served as a speaker for the following companies in the area of potential treatments for systemic sclerosis and its complications in the last three calendar years: 4P-Pharma, Abbvie, Acceleron, Alcimed, Altavant, Amgen, AnaMar, Arxx, AstraZeneca, Baecon, Blade, Bayer, Boehringer Ingelheim, Corbus, CSL Behring, Galderma, Galapagos, Glenmark, Gossamer, iQvia, Horizon, Inventiva, Janssen, Kymera, Lupin, Medscape, Merck, Miltenyi Biotec, Mitsubishi Tanabe, Novartis, Prometheus, Redxpharma, Roivant, Sanofi and Topadur. Patent issued “mir-29 for the treatment of systemic sclerosis” (US8247389, EP2331143). Research Grants: BI, Kymera, Mitsubishi Tanabe MIHAI Carina: CM received consulting fees and/or honoraria from Boehringer Ingelheim, Janssen Cilag AG, MED Talks Switzerland, Medbase, Mepha, Novartis, and PlayToKnow AG, and congress support from Boehringer Ingelheim and Roche., (Copyright © 2024 European Federation of Internal Medicine. Published by Elsevier B.V. All rights reserved.)

Primary Sjögren's Syndrome (pSS) falls within the category of connective tissue diseases, characterized by the presence of autoantibodies such as antinuclear antibodies (ANA). However, according to the classification criteria for pSS, some patients may exhibit a negative result for autoantibodies. Patients with a negative result for autoantibodies may lack typical features of connective tissue diseases, and the immunological state as well as the extent of organ involvement and damage may differ from those with positive autoantibodies. This study aims to compare the clinical phenotypes of patients with positive and negative autoantibodies, providing insights for disease classification and treatment selection for clinicians. Patients with pSS were grouped based on the presence and titers of their autoantibodies. Subsequently, differences in organ damage and laboratory indicators were compared between these groups, aiming to analyze the value of autoantibody titers in assessing the condition of pSS. (1) Patients with positive ANA exhibited elevated levels of inflammatory indicators, including ESR, IgG levels, lip gland biopsy pathology grade, and overall organ involvement, in comparison with patients with negative ANA (P < 0.05). Furthermore, ANA-positivity correlated with a higher occurrence of multi-organ damage, particularly affecting the skin, mucous membranes, and the hematological system (P < 0.05). (2) As ANA titers increased, patients demonstrated elevated levels of IgG and an escalation in organ involvement (P < 0.05). (3) Patients in the positive autoantibody group (positive for antinuclear antibodies, anti-SSA, or anti-SSB antibodies) had higher IgG levels compared to the negative group (P < 0.05). (4) Patients with positive anti-SSA and anti-SSB antibodies exhibited higher levels of inflammatory indicators and IgG compared to other patients (P < 0.05); however, no significant differences were observed in terms of organ involvement and organ damage. Patients with positive ANA in pSS typically exhibit higher levels of inflammation and an increased likelihood of experiencing multi-organ damage. Furthermore, as the ANA titers increase, both inflammation levels and the risk of multi-organ damage also escalate. Additionally, the presence of anti-SSA and anti-SSB antibodies may contribute to an elevated risk of increased inflammation levels, but does not increase the risk of organ damage., (© 2024. The Author(s).)

Rare antinuclear antibody (ANA) pattern recognition has been a widely applied technology for routine ANA screening in clinical laboratories. In recent years, the application of deep learning methods in recognizing ANA patterns has witnessed remarkable advancements. However, the majority of studies in this field have primarily focused on the classification of the most common ANA patterns, while another subset has concentrated on the detection of mitotic metaphase cells. To date, no prior research has been specifically dedicated to the identification of rare ANA patterns. In the present paper, we introduce a novel attention-based enhancement framework, which was designed for the recognition of rare ANA patterns in ANA-indirect immunofluorescence images. More specifically, we selected the algorithm with the best performance as our target detection network by conducting comparative experiments. We then further developed and enhanced the chosen algorithm through a series of optimizations. Then, attention mechanism was introduced to facilitate neural networks in expediting the learning process, extracting more essential and distinctive features for the target features that belong to the specific patterns. The proposed approach has helped to obtained high precision rate of 86.40%, 82.75% recall, 84.24% F1 score and 84.64% mean average precision for a 9-category rare ANA pattern detection task on our dataset. Finally, we evaluated the potential of the model as medical technologist assistant and observed that the technologist's performance improved after referring to the results of the model prediction. These promising results highlighted its potential as an efficient and reliable tool to assist medical technologists in their clinical practice., (© The Author(s) 2024. Published by Oxford University Press.)

Introduction: The combination of patterns is a frequent and challenging situation in the daily laboratory routine of autoantibodies testing using HEp-2 cells indirect immunofluorescence assay (HEp-2-IFA). Recently, the Brazilian Consensus on Autoantibodies (BCA) named these combinations as complex patterns (CPs) and organized them into 3 subtypes: multiple, mixed, and composite. This study aimed to describe the most frequent combinations of HEp-2-IIF patterns according to this new nomenclature., Methods: Routine HEp-2-IFA results reported in January and June 2017 were reviewed using the new BCA classification. Visual pattern recognition was performed by experts on HEp-2-IFA readings, using the International Consensus on Antinuclear Antibodies (ANA) Patterns (ICAP) and BCA recommendations., Results: 54,990 serum samples from different patients were tested for ANA-HEp-2, and 11,478 (20.9%) were positive at a titer ≥ 1/80. Among these positive samples, 1,111 (9.7%) displayed CPs, divided into 95 different combinations. A higher proportion of CPs was observed in the pediatric age group. Multiple, mixed, and composite patterns were present in 85.3, 5.4, and 9.5% of the samples, respectively. In the multiple/mixed pattern group (n=1,005), double, triple, and quadruple combinations (ICAP/BCA codes) were observed in 97.7%, 2.2%, and 0.1%, respectively. The double nuclear pattern was the most prevalent combination observed (67.6%). The most common CPs registered were AC-4 (nuclear fine speckled) + AC-6,7 (nuclear discrete dots) (n=264); AC-2 (nuclear dense fine speckled) + AC-6,7 (n=201); AC-4+AC-8,9,10 (nucleolar) (n=129); and AC-3 (centromere)+AC-4 (n=124). All of these combinations were in the multiple subgroup., Conclusion: Almost 10% of positive results in the HEp-2 procedure displayed CPs. Among the 3 subtypes of CPs proposed, the multiple pattern was the most prevalent, especially in the pediatric population. The AC-4, AC-2, and AC-6,7 were the most prevalent single patterns observed in the combinations described in this study. There was a significant association between age and the prevalence of most combined patterns. The AC-4+AC-6,7 combination was the most prevalent complex pattern detected regardless of the age group. The AC-2+AC-6,7 was more prevalent in younger individuals. The concepts involved in the CPs definition should add value to the reading and interpretation of the HEp-2-IIF assay., Competing Interests: Authors WS, AC, DF and NG were employed by company Sabin Medicina and Health. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Santos, Cantuária, Félix, Guimarães and de Melo.)

There are discrepancies across guidelines about whether the dosage of antinuclear antibodies (ANAs) is of use at the diagnosis of primary immune thrombocytopenia (ITP). This review describes the current knowledge about ANA prevalence in patients with primary ITP, and their potential usefulness as biomarkers for ITP evolution, response to treatments and increased risk of subsequent development of systemic lupus and thrombosis., (© 2023 British Society for Haematology and John Wiley & Sons Ltd.)

Introduction: Antinuclear antibodies (ANAs) are regarded as a hallmark of connective tissue diseases (CTDs) and play a key role in their diagnosis, but the value of some particular antibodies in management of patients and the disease prognosis is controversial. The mechanism underlying the production of ANAs in CTDs, other chronic inflammatory conditions and even in healthy people, is not completely elucidated. Anti-DFS70 antibodies connected with the dense fine speckled autoantigen of 70 kD, known as the lens epithelium-derived growth factor p75, are a subgroup of ANAs. Their presence and coexistence with other antibodies and their clinical significance are the matter of debate., Methods: Based on literature data, the authors focused on current knowledge explaining the role of anti-DFS70 antibodies in selected CTDs., Results: However, the literature data is ambiguous and does not fully support the validity of the anti-DFS70 assay for a specific CTD diagnosis. Most researchers claim that the presence of anti-DFS70 as the only one usually exclude the diagnosis of CTD. Nevertheless, its coexistence with other ANAs is not an excluding factor but has predictive value due to more favorable course of CTD. Such situations may also suggest an enhanced risk of the development of a CTD in the future., Conclusions: Although more studies are needed in this field, it seems reasonable to ascertain the presence of anti-DFS70 in routine clinical practice., (© 2023 The Authors. Immunity, Inflammation and Disease published by John Wiley & Sons Ltd.)

Objectives: Detection of antinuclear antibodies (ANA) by indirect immunofluorescence assay using HEp-2 cells (HEp-2 IFA) is used to screen for various autoimmune diseases. HEp-2 IFA suffers from variability, which hampers harmonization., Methods: A questionnaire was developed to collect information on HEp-2 IFA methodology, computer-assisted diagnosis (CAD) systems, training, inter-observer variability, quality assessment, reagent lot change control, and method verification. The questionnaire was distributed to laboratories by Sciensano (Belgium), national EASI groups (Italy, Croatia, Portugal, Estonia, Greece) and ICAP (worldwide). Answers were obtained by 414 laboratories. The results were analysed in the framework of the recent EFLM/EASI/ICAP ANA recommendations (companion paper)., Results: Laboratories used either HEp-2, HEp-2000, or HEp-20-10 cells and most laboratories (80%) applied the same screening dilution for children and adults. The conjugate used varied between laboratories [IgG-specific (in 57% of laboratories) vs. polyvalent]. Sixty-nine percent of CAD users reviewed the automatic nuclear pattern and 53% of CAD users did not fully exploit the fluorescence intensity for quality assurance. Internal quality control was performed by 96% of the laboratories, in 52% of the laboratories only with strongly positive samples. Interobserver variation was controlled by 79% of the laboratories. Limited lot-to-lot evaluation was performed by 68% of the laboratories. Method verification was done by 80% of the respondents., Conclusions: Even though many laboratories embrace high-quality HEp-2 IFA, substantial differences in how HEp-2 IFA is performed and controlled remain. Acting according to the EFLM/EASI/ICAP ANA recommendations can improve the global performance and quality of HEp-2 IFA and nurture harmonization., (© 2023 Walter de Gruyter GmbH, Berlin/Boston.)

Objectives: Antinuclear antibodies (ANA) are important for the diagnosis of various autoimmune diseases. ANA are usually detected by indirect immunofluorescence assay (IFA) using HEp-2 cells (HEp-2 IFA). There are many variables influencing HEp-2 IFA results, such as subjective visual reading, serum screening dilution, substrate manufacturing, microscope components and conjugate. Newer developments on ANA testing that offer novel features adopted by some clinical laboratories include automated computer-assisted diagnosis (CAD) systems and solid phase assays (SPA)., Methods: A group of experts reviewed current literature and established recommendations on methodological aspects of ANA testing. This process was supported by a two round Delphi exercise. International expert groups that participated in this initiative included (i) the European Federation of Clinical Chemistry and Laboratory Medicine (EFLM) Working Group "Autoimmunity Testing"; (ii) the European Autoimmune Standardization Initiative (EASI); and (iii) the International Consensus on ANA Patterns (ICAP)., Results: In total, 35 recommendations/statements related to (i) ANA testing and reporting by HEp-2 IFA; (ii) HEp-2 IFA methodological aspects including substrate/conjugate selection and the application of CAD systems; (iii) quality assurance; (iv) HEp-2 IFA validation/verification approaches and (v) SPA were formulated. Globally, 95% of all submitted scores in the final Delphi round were above 6 (moderately agree, agree or strongly agree) and 85% above 7 (agree and strongly agree), indicating strong international support for the proposed recommendations., Conclusions: These recommendations are an important step to achieve high quality ANA testing., (© 2023 Walter de Gruyter GmbH, Berlin/Boston.)

Antinuclear antibodies tests are of a paramount importance in the diagnosis, classification, prognostic evaluation and management of autoimmune diseases in children. The present study aimed to describe the immuno-clinical profile of antinuclear antibodies tests in a pediatric population in order to guide the clinical practice of biologists and clinicians. Our study enrolled 268 children. Antinuclear antibodies screening was performed using the indirect immunofluorescence assay on HEp-2 cells. Identification of target antigens was conducted using separately or at one timepoint the following techniques: enzyme-linked immunosorbent assay, Immunodot and Chemiluminescence. The average age of patients was 9.6 ± 4.3 years, with a female predominance (sex-ratio = 1.9). Antinuclear antibodies screening was positive in 40.67% of cases. The most frequently observed antinuclear antibodies patterns were speckled (52.3%), homogeneous (13.8%) and mixed homogeneous-speckled (13.8%). Autoantibodies were detected in 4 patients (2.51%) for whom ANA testing using the indirect immunofluorescence assay was negative. Positive antinuclear antibodies specificities were detected in connective tissue diseases (44.03%; n = 48), organ-specific autoimmune diseases (10.09%; n = 11), and in non-autoimmune conditions (inflammatory diseases, infections, hematological diseases, vasculitis and Wilson’s disease) (32.08%; n = 35). Our study revealed a high rate of positive antinuclear antibodies tests in the pediatric population, mainly related to autoimmune diseases (54.12%) besides non-autoimmune conditions (32.08%). Therefore, screening and interpretation of antinuclear antibodies testing in children require the consideration of clinical data and a close collaboration between clinicians and biologists.

Serum anti-Ro60 is one the most frequently encountered autoantibodies in the diagnostic immunopathology laboratory and in clinical practice. A large variety of assays are available to detect this including the popular multiplex line immunoblot (IB) assay. We evaluated the analytical performance of the IB for anti-Ro60 detection, using the counterimmunoelectrophoresis (CIEP) method as the 'gold standard'. We also undertook a survey of international laboratories, who use the IB, about their reporting practices for anti-Ro60. Using the manufacturer's reported cut-off of 15 units, the IB has an analytical sensitivity of 90.9% and specificity of 99.3% for anti-Ro60 detection. The optimal cut-off to balance sensitivity and specificity was determined to be 5 units with a sensitivity of 100% and specificity of 97.4%. Most laboratories use the manufacturer's specified cut-off (15 units) when determining a positive anti-Ro60 result. Whilst the commercial IB generally performs well, laboratorians need to be mindful of the limitations of IB in detecting antibodies that recognise conformational epitopes and what cut-offs they use. A vast majority of laboratories could potentially miss detection of this clinically important autoantibody., (Crown Copyright © 2022. Published by Elsevier B.V. All rights reserved.)

Despite evidence from human and animal studies of pesticide immunotoxicity, little is known about the relationship between agricultural pesticide use and autoimmunity. The objective of the present study was to assess the association between pesticide use and anti-cyclic citrullinated peptide (anti-CCP) antibodies and antinuclear antibodies (ANA) levels. A cross-sectional study including healthy 52 agricultural and 68 non-agricultural workers aged 17-69 years was conducted in the Rio de Janeiro State, Brazil. Serum samples were tested for anti-CCP and ANA by ELISA. Data on pesticide use and covariates were obtained through structured questionnaires. We estimated associations of pesticides and other exposures with log-transformed antibody levels, adjusted for sex, age, education, crops, and fertilizers by multiple linear regression analyzes. Anti-CCP levels were associated with growing certain crops, agricultural tasks, and the use of mancozeb, paraquat, and methomyl. ANA levels were positively associated with azoxystrobin and inversely associated with linuron. These novel findings suggest associations of specific pesticides and fertilizers with anti-CCP antibodies in this healthy agricultural population, supporting the need for larger human studies of preclinical autoimmunity and pesticides., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Objective: To explore the effect of anti-nuclear antibodies (ANAs) on the outcome of in vitro fertilization-embryo transplantation (IVF-ET) and to study the effect of ANAs in follicular fluid (FF) on embryonic development., Methods: The expression of ANAs in FF of patients treated with IVF-ET and healthy group. The patients were divided into ANAs-positive group and ANA-negative group. The age, duration of infertility, body mass index (BMI), basic follicle stimulating hormone (bFSH), anti-Mullerian hormone (AMH), number of retrieved oocytes, portion of metaphase II oocytes (MII), number of embryos in the cleavage stage, bipronuclear (2PN), number of embryos, number of high-quality embryos and the outcome of IVF-ET were compared between the two groups. In vitro, HTR8/SVneo trophoblast cells were cultivated and divided into ANAs-negative group (control group) and ANAs-positive group (ANAs-positive FF was added to cytotrophoblasts). The ANAs titer in the serum and FF of patients who treated with IVF-ET was detected using ELSIA method. CCK-8 assay and flow cytometry (at 24 h and at 48 h) were used to detect the cell proliferation and apoptosis frequency of the two groups, respectively., Results: Among those who underwent IVF-ET treatment, the number of retrieved oocytes, the number of fertilization and the portion of MII oocytes in the FF-positive group were significantly lower than those in the FF-negative group. Furthermore, the implantation rate and the clinical pregnancy rate were decreased, and early miscarriage rate was increased in the FF-positive patients than those in the FF-negative patients. In vitro, the cytotrophoblasts proliferation activity in the ANAs group was significantly lower than that in the control group. Moreover, the cytotrophoblasts apoptosis rate in the ANAs group was significantly higher than that in the control group., Conclusions: Our data suggested that ANAs in FF might become an obstacle to embryonic development through promoting trophoblast apoptosis and inhibiting trophoblast proliferation. ANAs in FF might be an unfavorable factor for the outcome of those who undergo IVF-ET treatment., (© 2022 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Introduction: Anti-Golgi antibodies (AGAs) are rare antibodies that are found as distinct, polarised cytoplasmic staining on the HEp-2 substrate., Methods: We performed a review of patients that demonstrated this autoantibody in a large laboratory cohort in Australia. Over a 24-month period, all patients that had a sample submitted for routine antinuclear antibodies (ANAs) that had AGA staining were retrospectively identified. Medical records were perused to identify clinical associations., Results: There were 23 813 ANAs identified with 34 patients (0.14%) demonstrating AGA staining. AGAs were found in a variety of inflammatory disorders, malignancies and liver diseases. They did not associate with any other significant ANA, and in contrast with previous studies, we did not find any association with systemic autoimmune rheumatic diseases., Conclusions: Anti-Golgi antibodies are rare, non-specific and possibly a bystander phenomenon. Future studies are required to study the origin and longitudinal clinical associations of these autoantibodies., (© 2021 The Scandinavian Foundation for Immunology.)

Objective: We investigated the autoantibody (autoAb) profiles in ANA+ individuals lacking systemic autoimmune rheumatic disease (SARD) and early SARD patients to determine the key differences between these groups and identify factors that are associated with an increased risk of symptomatic progression within the next 2 years in ANA+ individuals., Methods: Using custom antigen (Ag) microarrays, 144 IgM and IgG autoAbs were surveyed in 84 asymptomatic and 123 symptomatic (48 UCTD and 75 SARD patients) ANA+ individuals. AutoAbs were compared in ANA+ individuals lacking a SARD diagnosis with ≥2 years follow-up (n = 52), including all those who demonstrated progression (n = 14) during this period, with changes over time assessed in a representative subset., Results: We show that ANA+ individuals have autoAb to many self-Ags that are not being captured by current screening techniques and very high levels of these autoAbs are predominantly restricted to early SARD patients, with SLE patients displaying reactivity to many more autoAgs than the other groups. In general, the symptoms that developed in progressors mirrored those seen in SARD patients with similar patterns of autoAbs. Only anti-Ro52 Abs were found to predict progression (positive predictive value 46%, negative predictive value 89%). Surprisingly, over 2 years of follow-up the levels of autoAbs remained remarkably stable regardless of whether individuals progressed or not., Conclusion: Our findings strongly argue that development of assays with an expanded set of auto-Ags and enhanced dynamic range would improve the diagnostic and prognostic ability of autoAb testing., (© The Author(s) 2021. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

Purpose: The detection of antinuclear antibodies (ANA) in serum of patients with inflammatory bowel disease (IBD) has been associated with a worse response to anti-TNF therapy and the development of cutaneous or arthritic manifestations. The aim of this study was to investigate a possible association of serum ANA with infliximab (IFX) and adalimumab (ADA) trough levels (TLs) and anti-drug antibodies in IBD patients treated with IFX or ADA., Methods: Consecutive IBD patients under maintenance therapy with IFX or ADA in whom there was at least one available measurement of anti-TNF TLs, antibodies to IFX or ADA, and ANA in serum were included. The correlation of ANA positivity with demographics, clinical characteristics, treatment, TLs and anti-drug antibodies, of all patients was analyzed., Results: One hundred two IBD patients under maintenance therapy with IFX or ADA were enrolled. Of these, 53 (52%) were ANA positive with 28 (27.5%) positive also to anti-ds-DNA in serum. In the univariate analysis ANA positivity was found to be correlated with age (P = 0.008), female gender (P = 0.03), duration of treatment (P = 0.06), arthralgias (P = 0.04) and TLs (P = 0.005). However, in multivariate logistic regression analysis only age and TLs remained significantly associated with the presence of ANA positivity (P = 0.04 and P =  = 0.006, respectively). No significant association of ANA positivity with the development of cutaneous or rheumatological manifestations was found., Conclusions: In IBD patients under maintenance therapy with anti-TNF ANA positivity is associated with lower TLs. The clinical significance of this finding remains to be defined in future larger prospective studies., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Objective: Eltrombopag remains a prominent option in the treatment of steroid-dependent or steroid-refractory immune thrombocytopenia (ITP) patients. Unfortunately, not all patients respond to eltrombopag. Antinuclear antibody (ANA) positivity can be seen at rates of up to 30% in ITP patients. Despite being widely used, more markers to predict the response to eltrombopag are still needed. In the present study, we aimed to show the association between ANA positivity and eltrombopag response in ITP patients., Materials and Methods: Patients who were diagnosed with ITP in the Trakya University Faculty of Medicine’s Department of Hematology and who underwent eltrombopag treatment due to their resistance to steroids and other treatments were included in our study. ANA measurement was performed by indirect fluorescent antibody method and titers of 1:160 and above were considered positive. ANA measurements were made before starting eltrombopag., Results: Forty-five patients were included in our study, 33 being women and 12 men. The mean age of the patients was 45.73 years. There were 14 patients with ANA positivity and 31 patients were found to be ANA-negative. Response rates were higher in ANA-negative patients compared to ANA-positive patients in the 1 st and 6 th months of eltrombopag treatment (p<0.05)., Conclusion: ANA positivity in ITP may indicate unresponsiveness to eltrombopag treatment, a finding that should be further supported by prospective studies involving more patients.

The detection of antinuclear autoantibody (ANA) is dependent on many factors and varies between the populations. The aim of the study was first to assess the prevalence of ANA in the Polish adult population depending on age, sex and the cutoff threshold used for the results obtained. Second, we estimated the occurrence of individual types of ANA-staining patterns. We tested 1731 patient samples using commercially available IIFA using two cutoff thresholds of 1:100 and 1:160. We found ANA in 260 participants (15.0%), but the percentage of positive results strongly depended on the cutoff level. For a cutoff threshold 1:100, the positive population was 19.5% and for the 1:160 cutoff threshold, it was 11.7%. The most prevalent ANA-staining pattern was AC-2 Dense Fine speckled (50%), followed by AC-21 Reticular/AMA (14.38%) ANA more common in women (72%); 64% of ANA-positive patients were over 50 years of age. ANA prevalence in the Polish population is at a level observed in other highly developed countries and is more prevalent in women and elderly individuals. To reduce the number of positive results released, we suggest that Polish laboratories should set 1:160 as the cutoff threshold., (© 2021. The Author(s).)

Introduction: The indirect immunofluorescence assay on HEp-2 cells (HEp-2/IFA) is used worldwide for screening for autoantibodies to cellular antigens. Cell culture and fixation methods influence the cell distribution of autoantigens and the preservation of epitopes. Therefore, discrepancy of results obtained using different HEp-2/IFA kits (interkit nonreproducibility) is a common phenomenon in the clinical laboratory routine., Objective: This study evaluated the interkit nonreproducibility of HEp-2/IFA results using samples from patients with systemic autoimmune disease (SAD), nonautoimmune diseases (NAD), and healthy blood donors (HBD)., Methods: Serum from 275 SAD patients, 293 NAD patients, and 300 HBD were processed at 1:80 dilution using four HEp-2 kits according to the manufacturers' instructions. Interkit reproducibility was determined for positive/negative results and patterns. The agreement of positive/negative results among kits for each sample was determined as the reactivity agreement score (RAS). The pattern reproducibility score (PRS) in each sample was calculated as a function of the number of kits showing equivalent patterns. Qualitative variables and ordinal variables were analyzed by the Chi-square and Mann-Whitney U tests, respectively., Results: A total of 402 samples were nonreactive in all kits and were considered devoid of autoantibodies. Further analysis included the 466 reactive samples (238 SAD, 119 NAD, 109 HBD). Reactivity to the nucleus had the highest interkit reproducibility (RAS = 83.6), followed by the metaphase plate (RAS = 78.9), cytoplasm (RAS = 77.4), and nucleolus (RAS = 72.4). Interkit reproducibility was higher in SAD (RAS = 78.0) than in NAD (RAS = 70.6) and HBD (RAS = 71.3) groups. Samples with strong reactivity (++++/4 and +++/4) had higher interkit reproducibility than those with weak reactivity (+/4). In the SAD group, RAS for nuclear reactivity was 87.5% for strongly reactive samples as opposed to 4.4% for weakly reactive samples, and the same was observed for NAD and HBD samples. The most robust patterns were the centromere AC-3 (PRS = 78.4), multiple nuclear dots AC-6 (PRS = 73.6), nuclear coarse speckled AC-5 (PRS = 71.3), nuclear homogeneous AC-1 (PRS = 67.9), and the reticular cytoplasmic AC-21 (PRS = 68.6)., Conclusion: Interkit nonreproducibility in HEp-2/IFA is prevalent and occurs with the highest frequency with weakly reactive samples. International initiatives with the engagement of in vitro diagnostic industry are encouraged to promote the harmonization of the properties and performance of HEp-2/IFA commercial kits., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Silva, Dellavance, Baldo, Rodrigues, Grecco, Prado, Agustinelli and Andrade.)

The establishment of the International Consensus on ANA Patterns (ICAP) in 2014-2015 was welcomed by members of the medical community as a significant improvement in guiding harmonization of ANA test interpretation and reporting. In the subsequent years, several itinerant meetings and continuous interaction with the community contributed to disseminate the ICAP harmonization on the immunofluorescence patterns observed in the indirect immunofluorescence assay on HEp-2 cells (HEp-2 IFA) and to promote progressive improvement in the classification of HEp-2 IFA patterns. The 6th ICAP Workshop was held in person on September 6, 2021 as a satellite meeting of the 15th Dresden Symposium on Autoantibodies. This article summarizes the major discussions at the meeting as well as outlining the current plans for the ICAP committee., (© American Association for Clinical Chemistry 2021.)

Background: Antiextractable nuclear antigens (anti-ENAs) are regarded as diagnostic tests with no established value for serial monitoring. We therefore sought to establish the stability over time of anti-ENAs in a large diagnostic immunopathology laboratory., Methods: A retrospective review of all patients who had a serial anti-ENA ordered at the Westmead Hospital (Sydney, Australia) was performed over 24 months. Anti-ENA characterization was performed using line immunoassay, and historical data were available from 2013 onward. The earliest available densitometry readings were compared with the latest available to examine for a change in quantitation or qualitative (serostatus) result (from negative to positive, and vice versa). Medical records were examined for clinical correlations., Results: A total of 283 patients (24.1%) had serial testing of anti-ENA in the audit period, with each patient having an average of 3.9 ± 2.9 tests each. Most patients were diagnosed with systemic lupus erythematosus or primary Sjögren's syndrome. About 25% and 58% of patients had a qualitative and quantitative change, respectively, in at least 1 anti-ENA in the study period. Changes in anti-ENA levels correlated with erythrocyte sedimentation rate and disease activity. Increasing duration between serial tests increased the probability of observing a change in anti-ENA levels., Conclusion: Certain anti-ENAs are dynamic autoantibodies that may have significance for monitoring disease activity. Laboratories may consider reporting quantitative results. Further disease- and autoantibody-specific studies are required to determine the clinical significance of changes in anti-ENAs., (© American Association for Clinical Chemistry 2021. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

As iconic and important diagnostic autoantibodies, anti-Ro60 and anti-Ro52/tri-partite motif-containing 21 (TRIM21) make a common appearance in a number of systemic autoimmune disorders such as systemic lupus erythematosus (SLE). These autoantibodies often co-exist together; yet despite their close relationship, there is no evidence that they are physically linked and probably reflect a convergence of separate processes of failed immunological tolerance. Confusingly, they are sometimes classed together as the "SSA" or "Ro" autoantibody system without clear distinction between the two. In this Short Communication, we discuss the diagnostic merits for separate detection and reporting of these two autoantibodies, and discuss avenues for future research. Indeed, further insight into their fascinating origins and pathogenic roles in autoimmunity will surely shed light on how we can prevent and treat devastating autoimmune disorders., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Introduction: The morphological patterns in indirect immunofluorescence assay on HEp-2 cells (HEp-2 IFA) reflect the autoantibodies in the sample. The International Consensus on ANA Patterns (ICAP) classifies 30 relevant patterns (AC-0 to AC-29). AC-4 (fine speckled nuclear pattern) is associated to anti-SS-A/Ro, anti-SS-B/La, and several autoantibodies. Anti-SS-A/Ro samples may contain antibodies to Ro60 and Ro52. A variation of AC-4 (herein designated AC-4a), characterized by myriad discrete nuclear speckles, was reported to be associated with anti-SS-A/Ro. The plain fine speckled pattern (herein designated AC-4b) seldom was associated with anti-SS-A/Ro. This study reports the experience of four expert laboratories on AC-4a and AC-4b., Methods: Anti-Ro60 monoclonal antibody A7 was used to investigate the HEp-2 IFA pattern. Records containing concomitant HEp-2 IFA and SS-A/Ro tests from Durand Laboratory, Argentina ( n = 383) and Fleury Laboratory, Brazil ( n = 144,471) were analyzed for associations between HEp-2 IFA patterns and disease-associated autoantibodies (DAA): double-stranded DNA, Scl-70, nucleosome, SS-B/La, Sm, and U1-RNP. A total of 381 samples from Dresden Technical University (TU-Dresden), Germany, were assayed for HEp-2 IFA and DAA., Results: Monoclonal A7 recognized Ro60 in Western blot and immunoprecipitation, and yielded the AC-4a pattern on HEp-2 IFA. Analyses from Durand Laboratory and Fleury Laboratory yielded compatible results: AC-4a was less frequent (8.9% and 2.7%, respectively) than AC-4b (26.1% and 24.2%) in HEp-2 IFA-positive samples. Reactivity to SS-A/Ro occurred in 67.6% and 96.3% of AC-4a-pattern samples against 23% and 6.8% of AC-4b pattern samples. Reciprocally, AC-4a occurred in 24% and 47.1% of anti-SS-A/Ro-positive samples, and in 3.8% and 0.1% of anti-SS-A/Ro-negative samples. Data from TU-Dresden show that the AC-4a pattern occurred in 69% of 169 anti-SS-A/Ro-monospecific samples (62% of all anti-SS-A/Ro-positive samples) and in 4% of anti-SS-A/Ro-negative samples, whereas anti-SS-A/Ro occurred in 98.3% of AC-4a samples and in 47.9% of AC-4b samples. In all laboratories, coexistence of anti-SS-B/La, but not other DAA, in anti-SS-A/Ro-positive samples did not disturb the AC-4a pattern. AC-4a was predominantly associated with anti-Ro60 antibodies., Conclusions: This study confirms the association of AC-4a pattern and anti-SS-A/Ro in opposition to the AC-4b pattern. The results of four international expert laboratories support the worldwide applicability of these AC-4 pattern variants and their incorporation into ICAP classification under codes AC-4a and AC-4b, respectively. The AC-4 pattern should be maintained as an umbrella pattern for cases in which one cannot discriminate AC-4a and AC-4b patterns. The acknowledgment of the AC-4a pattern should add value to HEp-2 IFA interpretation., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Röber, Dellavance, Ingénito, Reimer, Carballo, Conrad, Chan and Andrade.)

Background: The diagnosis of systemic autoimmune rheumatic diseases (SARD) is based on the detection of serum antinuclear antibodies (ANA) for which indirect immunofluorescence (IIF) is the golden standard. New solid-phase immunoassays have been developed to be used alone or in combination with the detection of extractable antinuclear antibodies (ENA) to improve SARD diagnosis. The purpose of this study was to compare the clinical performances of different ANA screening methods alone or in combination with ENA screening methods for SARD diagnosis., Methods: A total of 323 patients were screened for ANA by IIF, EliA™ CTD Screen, and ELISA methods. Agreements were calculated between the methods. Then, EliA™ CTD Screen positive samples were screened for ENA by line immunoassay (LIA) and fluorescence enzyme immunoassay (FEIA)., Results: The diagnostic accuracy of EliA™ CTD Screen (79% sensitivity and 91% specificity) was better than that of ELISA or IIF. The combination of EliA™ CTD plus IIF had the highest sensitivity (93%). ENA determination revealed that Ro52 and Ro60 were the most prevalent specificities. The use of IIF alone was not able of detecting up to 36% of samples positive for Ro52, and 41% for Ro60., Conclusions: EliA™ CTD Screen has a better diagnostic performance when compared to IIF and ELISA. The combined use of EliA™ CTD Screen and IIF clearly improves the rate and accuracy of SARD diagnosis. The use of EliA™ CTD Screen as first-line screening technique allows the detection of antibodies, which could not be detected by IIF alone., (© 2021 The Authors. Journal of Clinical Laboratory Analysis published by Wiley Periodicals LLC.)

Objectives: ANA are the most extensively used test for the diagnosis of systemic autoimmune diseases. However, testing by indirect immunofluorescence assays (IIFAs) on HEp-2 cells, the gold standard test, is time-consuming and needs expertise. Thus there is a trend to replace it with other automated solid-phase assays directed against specific ANA. Nonetheless, the Hep-2 cell is an autoantigen array and ANA have been classified into 29 types, some of them with no clear association with a specificity to be detected. It is especially in these uncommon patterns where no clinical relationship is found and no antigenic specificity is detected. Here we retrospectively collected clinical data from patients with confirmed uncommon HEp-2 IIFA patterns to search for an associated clinical condition., Methods: We conducted an observational retrospective study including 608 patients with organ-specific and non-organ-specific autoimmune diseases (OSADs and NOSADs, respectively) with a confirmed rare pattern of ANA detected by IIFA on HEp-2 cells in the routine practice of the Spanish European Autoantibodies Standardization Initiative laboratories. Inclusion criteria are the existence of a minimum follow-up of 2 years and the availability of clinical data., Results: Nuclear patterns were more frequent in SLE (P = 0.001) and SS (P = 0.001), whereas the cytoplasmic ones were significantly higher in SSc (P = 0.022) and inflammatory myositis (P = 0.016). Mitotic patterns did not show any preferences for a specific disease and 62.7% of them corresponded to the nuclear mitotic apparatus pattern (AC-26). The most frequent NOSADs in patients with the AC-26 pattern were SLE (28.6%), SS (11.9%) and RA (11.9%). The cytoplasmic HEp-2 IIFA patterns were equally distributed in both groups of patients. In the OSAD patients there was no predominant pattern, except for AC-6 in primary biliary cholangitis due to Sp-100 antibodies (P < 0.001)., Conclusion: Detection of infrequent ANA might be a unique finding with no disease-associated specificities and could lead to the suspicion of an autoimmune disease., (© The Author(s) 2021. Published by Oxford University Press on behalf of the British Society for Rheumatology. All rights reserved. For permissions, please email: journals.permissions@oup.com.)

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., Competing Interests: Potential conflict of interest None declared., (Croatian Society of Medical Biochemistry and Laboratory Medicine.)

Introduction: The clinical significance of common antinuclear antibody (ANA) patterns, such as nuclear homogenous and nuclear speckled patterns with their corresponding specific antibodies, has already been established. However, the clinical relevance of these uncommon ANA patterns have not been well elucidated and these patterns are therefore not reported by most clinical laboratories. We herein report some retrospective data analysis linking patients' clinical status to several uncommon ANA patterns., Methods: We retrieved and assessed the patient records for ANA reports generated in our hospital over a period of two years. All testing had been performed using the gold standard Indirect Immunofluorescence Assay., Results: Records of 1235 consecutive patients tested for ANA were reviewed. ANA was positive in 330 of these patients with 6.39% found to have uncommon nuclear, cytoplasmic or mitotic sub-patterns. The mitotic spindle (0.89%), cytoplasmic anti-mitochondrial antibodies (0.80%), followed by discrete nuclear dots-multiple (0.72%) were the dominating patterns, with a higher prevalence in females than in males. Systemic lupus erythematosus and rheumatoid arthritis were the two most common autoimmune disorders associated with mitotic spindle fibers and nuclear centromere and nuclear large/coarse speckled ANA patterns., Conclusion: The prevalence of these relatively uncommon ANA patterns was higher than expected. Further evaluation of these patterns along with their corresponding antibodies and their clinical utility must be encouraged. We trust this endeavour will provide diagnostic information in autoimmune and other disease conditions., (Copyright © 2021 The Canadian Society of Clinical Chemists. Published by Elsevier Inc. All rights reserved.)

Background: A previous study have evaluated that antinuclear antibodies (ANA) negativization is linked to low lupus disease activity., Aim: To describe a lupus patient who evolved with negativization of ANA, anti-dsDNA, and anti-chromatin antibodies after vitamin D supplementation., Method: Case report., Results: A 56-year-old female patient, diagnosed with systemic lupus erythematosus since 2015 characterized by typical malar erythema, photosensitivity, polyarthritis, leucopenia, positive antinuclear antibody, anti-dsDNA, and anti-chromatin antibody. She received hydroxychloroquine and prednisone. After 1 year, corticotherapy was tapered off, and no clinical evidence of lupus activity was registered (SLEDAI = 0). However, ANA remained positive with a titer of 1:640 with a homogeneous pattern, and positive anti-dsDNA 1/20 and anti-chromatin 97 Units (normal range: <20 Units) remained all-time positive. Treatment with vitamin D 25,000 IU/day was initiated, and during follow-up, anti-chromatin and anti-dsDNA disappeared. In 2019, the patient was asymptomatic, keeping SLEDAI = 0, negative anti-dsDNA and anti-chromatin, and surprisingly the ANA turned negative, which was confirmed on several occasions until now., Conclusion: This case adds knowledge to the understanding that negative antinuclear antibodies appear to be associated with a better prognosis in lupus patients. Furthermore, the use of vitamin D seems to be a complementary therapeutic tool for this purpose., (Copyright © 2021 European Society for Clinical Nutrition and Metabolism. Published by Elsevier Ltd. All rights reserved.)

Purpose: The aim of this study was to investigate the possible link between different types of systemic sclerosis-specific antinuclear antibodies, adipokines and endothelial molecules which were recently found to have a pathogenic significance in systemic sclerosis., Materials/methods: Serum concentration of adiponectin, resistin, leptin, endothelin-1, fractalkine and galectin-3 were determined in the sera of patients with systemic sclerosis (n ​= ​100) and healthy controls (n ​= ​20) using ELISA., Results: The following associations between antinuclear antibodies and increased serum concentrations were identified: anticentromere antibodies with endothelin-1 (p ​< ​0.0001; mean level in patients 2.21 vs control group 1.31 ​pg/ml), anti-topoisomerase I antibodies with fractalkine (p ​< ​0.0001; 3.68 vs 1.68 ​ng/ml) and galectin-3 (p ​= ​0.0010, 6.39 vs 3.26 ​ng/ml). Anti-RNA polymerase III antibodies were associated with increased resistin (p ​< ​0.0001; 15.13 vs 8.54 ​ng/ml) and decreased adiponectin (p ​< ​0.0001; 2894 vs 8847 ​ng/ml)., Conclusion: In systemic sclerosis metabolic and vascular factors may serve as mediators between immunological abnormalities and non-immune driven clinical symptoms., Competing Interests: Declarations of competing interest The authors declare no conflict of interests., (Copyright © 2020 Medical University of Bialystok. Published by Elsevier B.V. All rights reserved.)

Objectives: Currently available computer-aided diagnosis (CAD) systems for the detection of anti-nuclear antibodies (ANA) by indirect immunofluorescence (IIF) assay enable a standardized measurement of system-specific fluorescent intensity (FI) measures. We aimed to evaluate an internal quality control (iQC) program that controls the total ANA IIF process in routine practice., Methods: In addition to the kit iQC materials, supplemental quality indicators were integrated in a total quality assurance (QA) program: patient-derived iQC's samples (negative, 1/160 fine speckled and 1/160 homogeneous), median sample FI per run and percentage of ANA IIF positive samples per run. Analytical rejection criteria were based on the imprecision of the positivity index (PI) measure of the Zenit PRO system (Menarini). Clinical rejection criteria were based on changes in FI that correspond to a change in ANA IIF titer of ≥2. To evaluate the QA program, different artificial errors were introduced during the ANA IIF process. After every run, quality indicators were evaluated and compared to the pre-set target values., Results: Rescanning the ANA IIF slides five times, using an old conjugate and a needle obstruction resulted in analytically and even clinically relevant errors in ANA IIF results. All errors were correctly detected by the different defined quality indicators. Traditional Westgard rules, including analytically (and clinically) defined rejection limits were useful in monitoring quality indicators., Conclusions: The integration of a total process iQC program in CAD systems, based on the specific FI measurands and performance criteria of the system, adds value to QA., (© 2021 Walter de Gruyter GmbH, Berlin/Boston.)

Background & Aims: The simplified criteria for the diagnosis of autoimmune hepatitis (AIH) include immunofluorescence testing (IFT) of antinuclear and smooth muscle autoantibodies (ANA and SMA) on rodent tissue sections. We aimed to establish scoring criteria for the implementation of ANA IFT on human epithelioma-2 (HEp-2) cells and ELISA-based testing., Methods: ANA and SMA reactivity of 61 AIH sera and 72 non-alcoholic fatty liver disease controls were separately assessed on tissue sections and HEp-2 cells to compare the diagnostic value at increasing titers. A total of 113 patients with AIH at diagnosis and 202 controls from 3 European centers were assessed by IFT as well as 3 different commercially available ANA ELISA and 1 anti-F-actin ELISA., Results: ANA assessment by IFT on liver sections had 83.6% sensitivity and 69.4% specificity for AIH at a titer of 1:40. On HEp-2 cells, sensitivity and specificity were 75.4% and 73.6%, respectively, at an adjusted titer of 1:160. Area under the curve (AUC) values of ANA ELISA ranged from 0.70-0.87, with ELISA coated with HEp-2 extracts in addition to selected antigens performing significantly better. SMA assessment by IFT had the highest specificity for the SMA-VG/T pattern and anti-microfilament reactivity on HEp-2 cells. ELISA-based anti-F-actin evaluation was a strong predictor of AIH (AUC 0.88) and performed better than SMA assessment by IFT (AUC 0.77-0.87)., Conclusion: At adjusted cut-offs, both ANA IFT using HEp-2 cells and ELISA-based autoantibody evaluation for ANA and SMA are potential alternatives to tissue-based IFT for the diagnosis of AIH., Lay Summary: Autoantibodies are a hallmark of autoimmune hepatitis and are traditionally tested for by immunofluorescence assays on rodent tissue sections. Herein, we demonstrate that human epithelioma cells can be used as a reliable substrate for immunofluorescence testing. ELISA-based testing is also a potentially reliable alternative for autoantibody assessment in autoimmune hepatitis. We propose the implementation of these testing methods into the simplified criteria for the diagnosis of autoimmune hepatitis., Competing Interests: Conflict of interest Weiler-Normann C reports speaker's fees from Euroimmun and Werfen (Inova) to her institution. All other authors declare no conflict of interest with respect to this study. Please refer to the accompanying ICMJE disclosure forms for further details., (Copyright © 2020 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.)

Background: Indirect immunofluorescence (IIF) is the most prevalent screening antinuclear antibody test for systemic autoimmune rheumatic disease (SARD). Certain IIF patterns have known antibody and disease associations, but the dense fine speckled (ANA-DFS) pattern has no confirmed clinical associations. Our objective was to determine the prevalence of SARD among a group of ANA-DFS positive individuals and to identify final diagnoses among non-SARD individuals in order to determine possible clinical associations with the ANA-DFS pattern., Methods: A retrospective study of 425 patients from a university health care system with a positive ANA-DFS pattern consecutively between August 2017 and September 2018. Sera samples underwent ANA testing by IIF on HEp-2 cell substrates (Euroimmun, Germany). Clinical information was retrieved from electronic health records and stored in a de-identified database., Results: The prevalence of SARD was 24%. Undetermined diagnosis (17%), skin disorders (12.1%), and fibromyalgia/chronic pain syndrome/chronic fatigue syndrome (11.8%) were the most common non-SARD diagnoses. Taking into account past medical history, the most common non-SARD were atopic disorders (21.2%), fibromyalgia/chronic pain syndrome/chronic fatigue syndrome (17.6%), and skin disorders (16.7%)., Conclusions: The ANA-DFS pattern may be indicative of an underlying antigen-antibody interaction that plays a role in either the initiation or propagation of immunologic reactions. DFS70/LEDGF is a transcription factor involved in cell survival and stress protection, and autoantibodies may inhibit its function. It is likely that there are other antibodies producing the ANA-DFS pattern besides anti-DFS70/LEDGF, and more research is necessary to identify additional antibody specificities. The ANA-DFS pattern may be an indicator of a proinflammatory microenvironment given the high frequency of symptomatic patients and disease processes with an immunologic basis (including SARD)., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Introduction: A proportion of patients with drug-induced liver injury (DILI) present with autoantibodies, which has led to the current concept of autoimmune-like DILI. However, no standardized definition exists and the clinical relevance has not been studied in detail yet., Methods: 143 patients with DILI enrolled in a prospective study were analyzed. DILI diagnosis was based on the monocyte-derived hepatocyte-like cell test and supported by Roussel Uclaf Causality Assessment Method (RUCAM) and expert adjudication. Testing for antinuclear antibodies (ANA) and antimitochondrial antibodies (AMA) was performed using immunofluorescence. ANA titers ≥1:100 were considered positive and ≥1:400 clinically relevant; AMA positivity was considered at titers ≥1:100., Results: 67% exhibited ANA ≥1:100 and 29% ANA ≥1:400; 10% were AMA positive. There was no significant correlation between the ANA titers and the causative drug, while AMA positive patients had taken nonsteroidal anti-inflammatory drugs more frequently. No difference was seen regarding clinical characteristics or laboratory parameters in patients with ANA ≥1:400, while patients with positive AMA presented with higher aminotransferases, bilirubin, and international normalized ratio. Significantly higher proportions of patients with ANA ≥1:400 or AMA positivity exhibited elevated immunoglobulin G levels. AMA positivity but not elevated ANA titers correlated with a higher proportion of Hy's law positivity., Conclusion: A closer look in a causality proven DILI cohort provided no evidence that presence of ANA titers is specific for DILI by a certain medication. AMA rather than ANA positivity was related to a more pronounced liver injury., (© 2020 S. Karger AG, Basel.)

Background: Indirect immunofluorescence assay (IIFA) is viewed as a preliminary standard to assess antinuclear antibodies (ANAs). Our aim was to explore ANA positivity rate, titers, and patterns in patients with systemic autoimmune rheumatic diseases (SARD), including systemic lupus erythematosus (SLE), rheumatoid arthritis (RA), primary Sjögren's syndrome (pSS), systemic sclerosis (SSc), and mixed connective tissue disease (MCTD), compared with healthy controls (HC)., Methods: Assess antinuclear antibody titers and patterns were retrospectively identified and compared by IIFA using human epithelial cells (HEp-2) and primate liver tissue substrate according to international consensus in SARD. Serum complement 3 (C3), C4, and immunoglobulin G were compared among subgroups with different ANA titers. The positive predictive values (PPV) for different ANA titers were calculated., Results: There were a total of 3510 samples, including 2034 SLE, 973 RA, 155 SSc, 309 pSS, and 39 MCTD cases. There was no difference in age between HC and SARD, excluding RA. ANA positivity rate in SARD and HC was 78.7% and 12.2%, respectively. A titer of ≥1:320 revealed a PPV of 84.0% in SARD. SLE patients with ANA titers ≥1:320 had significantly lower levels of C3 and C4. AC-4 (31.2%) was the major pattern in patients with SARD, followed by AC-5 (23.9%) and AC-1 (18.8%). SLE mostly presented with AC-4 (30.3%). Several mixed patterns provided a significant hint for SSc and SLE. The major pattern in HC was AC-2 (12.2%)., Conclusions: Assess antinuclear antibody positivity, titers, and patterns display differences in various SARD, contributing to the classification of SARD., (© 2020 The Authors. Journal of Clinical Laboratory Analysis Published by Wiley Periodicals LLC.)

Combined immunotherapy and chemotherapy is a promising standard treatment in patients with advanced non-small cell lung cancer (NSCLC). This study aimed to evaluate the relationship between the combined therapy and pretreatment serum antinuclear antibody (ANA) levels as a prognostic indicator in patients with NSCLC. We retrospectively analyzed patients with advanced NSCLC who were treated with combinatorial immunotherapy and chemotherapy between January and December 2019 at six institutions in Japan. Relationship between ANA status and patients' characteristics were reviewed. A total of 77 patients with advanced NSCLC were enrolled in the study. Patients were divided into ANA-positive (ANA ≥ 1:160) and ANA-negative (ANA < 1:160) groups. The ANA-positive group tended to have a shorter progression-free survival and significantly shorter overall survival in univariate (hazard ratio [HR], 2.11, 95% confidence interval [CI] 0.88-5.07, p = 0.093; and HR 3.11, 95% CI 1.14-8.49, p = 0.027, respectively) and multivariate (HR 1.90, 95% CI 0.77-4.68, p = 0.16; and HR 3.37, 95% CI 1.15-9.86, p = 0.027, respectively) analyses than ANA-negative group. The incidence of discontinuation of all treatment components due to severe adverse events was significantly higher in the ANA-positive than in ANA-negative group (50% vs. 15.9%, p = 0.042). The study showed that the presence of antinuclear antibodies may result in a poor prognosis in patients treated with combinatorial immunotherapy and chemotherapy, although further prospective investigations are needed.

Background: Antibodies 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., Objective: To determine the frequency of ANCA in a population with manifestations of autoimmune disease, and evaluate the interference of ANA in its interpretation., Materials and Methods: Retrospective, descriptive nonexperimental cross-sectional study, including 3,330 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., Results: ANCAs 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., Conclusions: The 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 minimize "false positives"., (Copyright © 2018 Elsevier España, S.L.U. and Sociedad Española de Reumatología y Colegio Mexicano de Reumatología. All rights reserved.)

Problem: The mechanism(s) by which antinuclear antibodies (ANA) induce implantation failure are not clear, and little information regarding the function of autoantibodies in reproductive tissues is available., Methods of Study: A total of 380 patients who underwent in vitro fertilization and embryo transfer (IVF-ET) were divided into control, serum positive, and follicular fluid (FF) positive groups based on the results of indirect immunofluorescence assay for ANA in the serum and FF. Immunofluorescence assay was performed to evaluate the existence of ANA in granular cells and endometrial tissues. Presence in FF of soluble apoptotic markers, including Bcl-2, Caspase-3, cleaved PARP, Cytochrome C, GAPDH, and p53, was assessed using magnetic bead based assays., Results: The patients in the FF positive group had the lowest numbers of retrieved oocytes, fertilizations, and high-quality embryos. The fertilization rate, and the proportion of two pronuclear (2PN) embryos in patients in the FF positive group were significantly lower than those in the other two groups. The FF positive group also had the lowest clinical pregnancy rate, and the highest early miscarriage rate. Granulosa cells and endometrial tissues in patients in the FF positive group were ANA positive. High levels of BCL-2, Caspase-3, Cytochrome C, GAPDH, and p53 were found in the FF of patients in the FF positive group., Conclusions: Antinuclear antibodies in FF and endometrial tissues may cause imbalanced apoptosis, resulting in poor IVF-ET treatment outcomes. Local autoimmunity and cell apoptosis in reproductive tissues could be considered new therapeutic targets for improving IVF-ET treatment efficacy., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Connective tissue diseases (CTD) are autoimmune diseases highly associated with the presence of antinuclear antibodies (ANA). Since ANA and musculoskeletal symptoms are not uncommon in the general population, differential diagnostic challenges frequently occur for the treating physician. Dense fine speckled antibodies (DFS70) were recently discovered but their presence appeared to be rare in CTD., Methods: In this cross-sectional study a total of 270 patients treated in the Rheumatism Center of the Ruhr Area (Rheumazentrum Ruhrgebiet) were preferentially included, when they were ANA+ (≥1:80). Other autoantibodies and DFS70 antibodies were also investigated. The diagnosis of CTD was confirmed by rheumatologists. The sensitivity, specificity and the positive predictive value of DFS70 antibodies were differentially evaluated for various ANA titers., Results: In 91 (33.7%) of the ANA+ patients (33.7%) the diagnosis of CTD was confirmed and in 84 (92.3%) the ANA titer was ≥1:160. The DFS70 antibodies were detected in 17 out of 130 ANA+ patients without CTD (13.1%) versus 2 ANA+ patients (2.2%) with CTD (p = 0.027). None of the patients with ANA 1:80 had DFS70 antibodies. The specificity of DFS70 antibodies to detect the absence of CTD was 97.6%, the sensitivity was 13.1% and the positive predictive value was 89.5%. There was almost no effect of the strength of the different ANA titers., Conclusion: It was found that DFS70 antibodies are rarely present in ANA+ patients with CTD but the diagnosis of CTD cannot be reliably ruled out solely by the presence of DFS70 antibodies; however, the high specificity of DFS70 antibodies can be of clinical importance in unclear situations and in cases of anxious patients.

Background: Narrowband UVB (NBUVB) phototherapy is the cornerstone treatment for vitiligo. Before its initiation, some experts recommend antinuclear antibody (ANA) screening out of concern for either photosensitivity to NBUVB or autoimmune disease exacerbation during treatment. As vitiligo is considered an autoimmune disorder, ANAs can be positively found in the disease without any clinical importance. The necessity for ANA investigations for pre-phototherapy vitiligo patients is therefore questioned., Methods: We conducted a retrospective study to investigate vitiligo patients who had been checked for ANA before commencing NBUVB phototherapy. Demographic data-including vitiligo type and age of onset-were collected. Samples of ANA, anti-thyroglobulin, and anti-thyroid peroxidase were obtained. The phototherapy treatment protocol and cutaneous reactions to the phototherapy were also recorded., Results: Among 85 Thai vitiligo patients, the ANA prevalence was 35.3%. The speckled ANA pattern was the most common, and the large majority of patients (80%) had a titer of ≤1:100. Factors associated with positive ANA were female gender and positive anti-thyroglobulin. There were no statistical differences between the phototoxic reactions or phototoxic doses of NBUVB of the ANA-positive vitiligo and ANA-negative vitiligo groups. No cases of SLE were detected in ANA-positive group., Conclusions: ANA positivity was not correlated with the incidence or dose of phototoxic reaction in phototherapy treated vitiligo, and it may not a predictive factor for SLE diagnosis in vitiligo. ANA might therefore not need to be routinely checked in pre-phototherapy in vitiligo, unless there are clinical suspicions of an autoimmune disease. However, ANA might be involved in part of the cutaneous photoadaptation response to phototherapy., (© 2020 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Introduction: Antinuclear antibody (ANA) testing using indirect immunofluorescence assay (IIFA) is a common and economical method which contributes to detect systemic autoimmune diseases (SARD) and autoimmune liver diseases (AILD). The primary aim of our study was to investigate ANA positivity and their patterns in multiple liver diseases, including primary biliary cirrhosis (PBC), autoimmune hepatitis (AIH), hepatitis B virus infection (HBV), hepatitis C virus infection (HCV), and hepatic carcinoma (HCC). Besides, we also compared the ANA titers and patterns in patients with liver disease, SARD, and healthy controls (HC)., Methods: A total of 2537 patients with SARD, 137 PBC cases, 57 AIH cases, 3420 HBV cases, 769 HCV cases, 268 HCC cases, and 1073 HC were retrospectively assessed. The titers and patterns of ANA were detected with the IIFA method., Results: ANA positivity rate was considerably discernible between these diseases, which is 90.1% in SARD, 93.4% in PBC, 49.1% in AIH, 19.1% in HBV, 13.9% in HCV, and 23.5% in HCC. Moreover, only 4.9% of HCC cases, 2.5% of HBV patients, and 1.6% of HCV patients had an ANA titer ≥ 1:320. The mixed pattern which composed of at least two patterns majorly lied in PBC. AC-15 and AC-21 was frequently related to liver diseases; the former pattern was more frequently found in AIH (84.2%) and PBC (8.8%), and the latter pattern was easily seen in PBC (62.2%) and HCC (22.6%). The positive rate of ANA in HC was 12.2%, and its major pattern was AC-2., Conclusions: There are differences in ANA positivity among patients with SARD and various liver diseases. Some mixed patterns may provide important evidence for the diagnosis of PBC. Clinicians should pay attention to ANA patterns and titer during the interpretation of this test. Key Points • Defining the clinical relevance of antinuclear antibody (ANA) using indirect immunofluorescence assay in the context of diseases can be an important tool for the clinician in the diagnostic work-up of patients with liver diseases. • The mixed pattern of ANA is majorly found in primary biliary cirrhosis (PBC). ANA patterns including AC-15 and AC-21 are frequently related to liver diseases. AC-15 is more often found in autoimmune hepatitis (AIH) (84.2%) and PBC (8.8%), and AC-21 is easily found in PBC (62.2%, and hepatic carcinoma (HCC) (22.6%). • ANA positivity can be seen in 19.1% of hepatitis B virus infection (HBV) cases, 13.9% of hepatitis C virus infection (HCV) cases, and 23.5% of HCC cases. Only 2.5% of HBV patients, 1.6% of HCV patients, and 4.9% of HCC cases have an ANA titer ≥ 1:320.

Purpose: Immune checkpoint inhibitors (ICIs) show promising clinical activity in advanced cancers. However, the safety and efficacy of PD-1/PD-L1 blockade in patients with preexisting antinuclear antibodies (ANA) are unclear., Methods: 191 patients treated with nivolumab, pembrolizumab, atezolizumab, or durvalumab for unresectable advanced cancers between September 2014 and December 2018 were identified retrospectively. Patients were divided into positive (ANA titers ≥ 1:160) and negative ANA groups (ANA titers < 1:160). Development of immune-related adverse events (irAEs), the overall response rate (ORR), and disease control rate (DCR) were monitored., Results: Positive ANA titers were seen in 9 out of 191 patients. Four patients in the positive ANA group and 69 patients in the negative group developed irAEs of any grade without a significant difference between the groups. The development of endocrine, pulmonary, and cutaneous irAEs was not significant, whereas positive ANA was significantly higher in patients who developed colitis (2/9) than in patients who did not (3/182, P = 0.0002). DCR in the positive and negative ANA group was 37.5% and 67.5%, respectively, and was not statistically significant, but had better efficacy in patients without ANA (P = 0.08). ANA-related autoimmune diseases such as SLE, Sjögren's syndrome, MCTD, scleroderma, dermatomyositis, and polymyositis was not induced in either group. However, one patient with preexisting dermatomyositis had a flare up after initiation of atezolizumab., Conclusion: Further studies to identify predictive factors for the development of irAEs are required to provide relevant patient care and maximize the therapeutic benefits of ICIs.

Background: The indirect immunofluorescence assay (IFA) using HEp-2 cell substrates is the preferred method by some for detecting antinuclear antibodies (ANA) as it demonstrates a number of characteristic staining patterns that reflect the cellular components bound as well as semi-quantitative results. Lack of harmonized nomenclature for HEp-2 IFA patterns, subjectivity in interpretation and variability in the number of patterns reported by different laboratories pose significant harmonization challenges. The main objectives of this study were to assess current practice in laboratory assessment of HEp-2 IFA, identify gaps and define strategies to improve reading, interpretation and reporting., Methods: We developed and administered a 24-item survey based on four domains: educational and professional background of participants, current practice of HEp-2 IFA testing and training, gap assessment and the perceived value of International Consensus on Antinuclear Antibody Patterns (ICAP) and other factors in HEp-2 IFA assessment. The Association of Medical Laboratory Immunologists (AMLI) and American Society for Clinical Pathology administered the survey from April 1 to June 30, 2018, to members involved in ANA testing. This report summarizes the survey results and discussion from a dry workshop held during the 2019 AMLI annual meeting., Results: One hundred and seventy-nine (n = 179) responses were obtained where a significant number were clinical laboratory scientists (46%), laboratory directors (24%), supervisors (13%) or others (17%). A majority of respondents agreed on the need to standardize nomenclature and reporting of HEp-2 IFA results. About 55% were aware of the ICAP initiative; however, among those aware, a significant majority thought its guidance on HEp-2 IFA nomenclature and reporting is of value to clinical laboratories. To improve ICAP awareness and further enhance HEp-2 IFA assessment, increased collaboration between ICAP and the clinical laboratory community was suggested with emphasis on education and availability of reference materials., Conclusions: Based on these suggestions, future efforts to optimize HEp-2 IFA reading, interpretation and reporting would benefit from more hands-on training of laboratory personnel as well as continuous collaboration between professional organizations, in vitro diagnostic manufacturers and clinical laboratories.

Introduction: Antinuclear antibody (ANA) tests are widely used for the diagnosis of autoimmune diseases, but ANAs are also commonly found in patients with various infections. This retrospective study aimed to investigate the relationship between infections and ANA status. Methods: Patients that visited the Department of Infectious Diseases at Inha University Hospital between January 2007 and July 2018 were investigated. We analysed their ANA test results and reviewed rheumatic and infectious diagnoses of patients with positive ANA findings. Results: Of the 9,320 patients during the study period, 1,111 underwent ANA testing and 110 tested positive. Seven of the 110 patients were previously diagnosed with ANA-positive disease, and 21 were diagnosed with autoimmune disease during the present study. Of the remaining 82 patients, 43 were confirmed with infectious disease. The most common pathogen was Mycobacterium tuberculosis ( n  = 10), followed by Treponema pallidum ( n  = 5), Orientia tsutsugamushi ( n  = 5), Escherichia coli ( n  = 5), Bartonella henselae ( n  = 3), and human immunodeficiency virus ( n  = 3). Of the 39 patients without a confirmed pathogen, 7 were seropositive for O. tsutsugamushi , B. henselae , or Rickettsia spp. Patients were observed at an average of 24 weeks in our hospital. One patient developed systemic lupus erythematosus after being diagnosed with Epstein-Barr virus-induced infectious mononucleosis, and another patient developed adult-onset Still's disease after being diagnosed with scrub typhus. Conclusion: This study showed that various relationships exist between infections and rheumatic diseases. In particular, several patients with a positive ANA test result were found to have intracellular infections such as mycobacterial infections, syphilis, or scrub typhus.

The study aimed to retrospectively investigate the clinical significance of anti-rods and rings (anti-RR) antibodies in antinuclear antibodies (ANAs) test samples of western China. Between January 2016 and November 2018, the laboratory data and clinical details of patients with positive anti-RR antibodies were collected and analysed. The results showed that total of 197 227 patients tested, 109 453 patients presented with positive ANAs (55.50%), but only 107 patients with positive anti-RR antibodies (0.10%), including 51 females and 56 males. Diagnose were established in 51 of 107 patients: 25 were hepatopathy (HCV 8/25, HBV 12/25); 13 were autoimmune diseases (AID); and 7 were renal insufficiency; 6 were chronic obstructive pulmonary disease (COPD). We make the conclusions that anti-RR antibodies have a low prevalence, and there is no gender difference. Anti-RR antibodies exist other diseases besides hepatitis C, such as HBV, some autoimmune diseases, renal insufficiency and COPD, which we need further investigation., (© 2019 The Scandinavian Foundation for Immunology.)