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INTRODUCTION The antinuclear antibody (ANA) test is a screening test for systemic autoimmune rheumatic disease (SARD). We hypothesised that the presence of anti-DFS70 in ANA-positive samples was associated with a false-positive ANA test and negatively associated with SARD. METHODS A retrospective analysis of patient samples received for ANA testing from 1 January 2016 to 30 June 2016 was performed. Patient samples underwent ANA testing via indirect immunofluorescence method and anti-DFS70 testing using enzyme-linked immunosorbent assay. RESULTS Among a total of 645 ANA-positive samples, the majority (41.7%) were positive at a titre of 1:80. The commonest nuclear staining pattern (65.5%) was speckled. Only 9.5% of ANA-positive patients were diagnosed with SARD. Anti-DFS70 was found to be present in 10.0% of ANA-positive patients. The majority (51/59, 86.4%) of patients did not have SARD. Seven patients had positive ANA titre > 1:640, the presence of anti-double stranded DNA and/or anti-Ro60. The presence of anti-DFS70 in ANA-positive patients was not associated with the absence of SARD (Fisher’s exact test, p = 0.245). CONCLUSION The presence of anti-DFS70 was associated with a false-positive ANA test in 8.6% of our patients. Anti-DFS70 was not associated with the absence of SARD.

Background Time and cost constraints lead to majority of clinical laboratories deviating away from an ideal practice of checking for antinuclear antibodies (ANAs) by indirect immunofluorescence (IIF) at multiple dilutions. Usage of screening dilution of 1:40 recommended by most manufacturers of commercial ANA kits results in numerous false positive-tests and misdiagnosis of connective tissue disorders (CTDs). We sought to study the ideal screening dilution for ANA by IIF for a diagnosis of ANA-related CTDs. Methods Serum samples of patients with ANA-related conditions (n = 233) and healthy controls (n = 154) were evaluated by IIF using Immuno Concepts Hep-2000 ® ANA kits at dilutions from 1:40 to 1:640. Accuracy for diagnosis of CTDs for each serum dilution was assessed by receiver operating curve (ROC) analysis. Results Antinuclear antibodies (ANA) positivity was observed in 19.5%, 10.4%, 4.55%, 0.65%, and 0% of healthy controls at dilutions of 1:40, 1:80, 1:160, 1:320, and 1:640, respectively. ANA positivity at 1:40 dilution was observed among 26.4% cases with mimics of CTDs. Prevalence of ANA positivity in ANA-related CTDs was 97.3%, 96.4%, 89.3%, 83.9%, and 71.4% at dilutions of 1:40, 1:80, 1:160, 1:320, and 1:640, respectively. ROC analysis revealed best test performance for distinction between healthy and ANA-related CTD populations at a serum dilution of 1 in 80. Conclusions Antinuclear antibodies (ANA) positivity at low titers (1:40) is highly prevalent in healthy population (19.5%) as well as amongst mimics of CTD (26.4%). Our study suggests a higher screening dilution of 1:80 for ANA by IIF for diagnosis of CTD maybe better. Combination of 1:80 and 1:160 dilutions provides optimum sensitivity and specificity for diagnosis of ANA-related disorders.

Objectives To evaluate the performance characteristics of a line immunoassay (LIA) for the detection of Mi-2 antibodies associated with dermatomyositis (DM). Methods In total, 432 consecutive patient specimens were tested for Mi-2 antibodies concurrently by LIA (Mi-2α or Mi-2β) or immunoprecipitation (IP) test and antinuclear antibody by indirect immunofluorescence assay using HEp-2 substrate. Following antibody evaluation, results for patients positive in any of the assays for Mi-2 antibody had a retrospective chart review for diagnostic categorization. The performance of all tests was evaluated based on the extracted clinical data. Results Forty patients were positive in at least one of the Mi-2 assays. The frequency of Mi-2β antibody by LIA was highest (75.0%), followed by Mi-2 by IP (35.0%) and Mi-2α by LIA (20.0%), respectively. Mi-2 by IP had the best total percent agreement for DM (95.0%) compared with 70.0% and 25.0% for the LIA Mi-2α and Mi-2β, respectively. Positivity of the Mi-2β antibody was significantly associated with non-DM diagnosis. Conclusions Agreement for DM with assays for detecting Mi-2 is variable. Additional studies are required to validate Mi-2 immunoassays for routine patient evaluation.

Introduction: The most severe autoimmune subepidermal blistering condition of the skin and mucous membranes is bullous pemphigoid (BP). In Europe, it is estimated to affect 1 in every 4,000 people. Currently incidence range between 2-22/1,000,000 worldwide. It primarily affects the elderly and is diagnosed using clinical, histologic, and immunologic criteria. Clinically, it appears as diffuse eczematous, pruritic, urticaria-like lesions with the later emergence of tense bullae or blistering lesions filled with clear fluid. Case Presentation: Here, we report a case of a 50- years old female patient with a complaint of itchy lesions with wounds all over the body present with an 8-month of history. A subepidermal blister with eosinophils and neutrophils infiltration was discovered on histopathological evaluation. Salt-split indirect immunofluorescence revealed linear deposition of IgG at the dermo-epidermal junction. On further investigation, using diagnostic and Interventional aids a final diagnosis of Bullous pemphigoid.

Despite the expansion of available in vitro laboratory tests at a rate far exceeding that of dermatologic pharmaceuticals, the existing literature is dominated by discussion of the latter. With the advent of numerous new tests, it can be difficult for practicing dermatologists to stay up-to-date on the available options, methodologies, and recommendations for when to order one test over another. Understanding the inherent strengths and weaknesses of these options is necessary to inform appropriate ordering and proper interpretation of the results. The first article in this continuing medical education series summarizes information on methodology, test characteristics, and limitations of several in vitro laboratory tests used for the work up of undifferentiated patients suspected of having dermatologic autoimmune diseases and it provides a general guide to ordering these tests.

Background Antineutrophil cytoplasmic antibodies (ANCA) are important biomarkers in the diagnosis of ANCA-associated vasculitis, and indirect immunofluorescence (IIF) had been the method of choice for its detection from the very beginning. However, international consensus on ANCA testing (2017) advocates the use of high-quality immunoassays as the primary screening method. The purpose of this study was to evaluate the diagnostic performance of enzyme-linked immunosorbent assay (ELISA) compared to IIF in detecting ANCA. Methods One-hundred eighty-nine serum samples of suspected or known cases of systemic vasculitis were screened for ANCA by IIF and proteinase-3- and myeloperoxidase-ELISA. In IIF, positive results were further divided into cytoplasmic pattern of ANCA and perinuclear pattern of ANCA, depending upon the pattern of fluorescence. McNemar’s chi-squared test was applied to check the equality of proportions of positive results, and Kappa statistics was used to measure the agreement between the two methods. Diagnostic performance of ELISA was evaluated taking IIF as reference. Results IIF detected ANCA in 17.5% cases and ELISA detected it in 11.6% cases. A good agreement between the overall performance of ELISA and IIF was observed (K-value: 6.8, p-value: < 001). However, a significant difference in the proportion of positive results by the two methods was observed in McNemar’s test (two-sided p-value: 0.007). Taking IIF as standard, ELISA showed 60.6% sensitivity, 98.7% specificity, and predictive value of positive and negative results of 90.9 and 92.2%, respectively. Conclusion The new generation antigen-specific ELISAs had high specificity but the chances of missing cases in primary screening due to the low sensitivity and high false negativity (39.4%) need to be dealt with.

Screening for autoantibodies in HEp-2 cells by indirect immunofluorescence is currently accepted as the gold-standard test for the diagnosis of systemic autoimmune diseases. The main objective of the International Consensus on ANA Patterns is to achieve a consensus on the nomenclature and description of antinuclear antibody morphological patterns. This work aims to build on the International Consensus on ANA Patterns project to establish a nomenclature consensus in Portugal, thus contributing to harmonization in autoimmune diagnosis and promoting diagnostic quality in autoimmune systemic rheumatic diseases.Participating laboratories identified all the nuclear and cytoplasmic pattern designations in the International Consensus on ANA Patterns (including the anti-cell pattern code), and matched them with the corresponding Portuguese nomenclature in use. The results were aggregated and used as a foundation for nomenclature harmonization work. Consensus meetings followed an iterative process, until a final consensual proposal was drafted.Prior agreement between laboratories was over 75% for 23 of the total 29 anti-cell patterns. The degree to which each laboratory is aligned with the International Consensus on ANA Patterns international reference ranges from 22.1% to 100%. It was possible to write a consensual version of the International Consensus on ANA Patterns nomenclature for Portugal.There was a good consensus basis for the nomenclature in the International Consensus on ANA Patterns, despite relevant differences with some translations. The study highlights the need for collaboration among laboratories towards an unambiguous description of laboratory results.This study shows that there is good potential for collaboration between laboratories in order to produce the consensus needed to improve diagnosis and patient follow-up.Introdução: A pesquisa de autoanticorpos em células HEp-2 através de imunofluorescência indireta é o teste padrão atualmente aceite como a ferramenta central para o diagnóstico das doenças autoimunes sistémicas. O International Consensus on Antinuclear Antibody (ANA) Patterns tem como objetivo principal alcançar um consenso na nomenclatura e na descrição dos diferentes padrões morfológicos de anticorpos antinucleares. Este trabalho tem como objetivo ampliar o projeto do International Consensus on ANA Patterns de forma a estabelecer um consenso em Portugal para a sua nomenclatura, procurando contribuir para a harmonização no diagnóstico autoimune e promover a qualidade diagnóstica nas doenças reumáticas sistémicas autoimunes.Material e Métodos: Os laboratórios participantes identificaram cada designação de padrão citoplasmático e nuclear do International Consensus on ANA Patterns (incluindo o código padrão anti-célula), e fizeram corresponder a cada uma a respetiva nomenclatura portuguesa em uso. Os resultados foram agregados e serviram de base ao trabalho de harmonização da nomenclatura. Seguiram-se reuniões de consenso, num processo iterativo até à redação de uma proposta final consensualizada.Resultados: A concordância prévia entre laboratórios era superior a 75% para 23 do total de 29 padrões anti-célula. O grau em que cada laboratório está alinhado com a referência internacional do International Consensus on ANA Patterns varia entre 22,1% e 100%. Foi possível elaborar uma versão consensualizada da nomenclatura do International Consensus on ANA Patterns para Portugal.Discussão: Existia uma boa base de consenso para a nomenclatura do International Consensus on ANA Patterns, mas com diferenças importantes em algumas das traduções da terminologia. O estudo realça a necessidade de colaboração entre laboratórios para uma descrição inequívoca dos resultados laboratoriais.Conclusão: Este trabalho mostra o potencial positivo da colaboração entre laboratórios para gerar consensos que contribuam para a melhoria do diagnóstico e acompanhamento dos doentes.

Neosporose e toxoplasmose são doenças de importância na esfera reprodutiva e em saúde pública. Dados de soroprevalência destes protozoários em regiões de importante cadeia produtiva de bovinos no Brasil, como a região Sul, são necessários para compreender a extensão dos problemas causados por estas enfermidades, bem como melhorar o emprego de medidas profiláticas e de controle. Os resultados do levantamento sorológico realizado por este estudo, pela reação de Imunofluorescência Indireta (IFI) para detecção de anticorpos anti- T. gondii e anti- N. caninum, apontaram animais positivos para infecção por T. gondii (7,2%) e N. caninum (7,6%), abaixo das citadas na literatura no Estado do Rio Grande do Sul, que parece apresentar prevalências menores quando comparado com outras regiões do Brasil, caracterizando uma disseminação moderada destes agentes.

Background: Anti-nuclear antibodies (ANA) are known to be associated with autoimmune diseases and most commonly with connective tissue diseases (CTD). Screening for ANA is the first investigation of choice in the diagnosis of autoimmune diseases and indirect immunofluorescence (IIF) test using Hep-2 cells has been considered the gold standard method. The prevalence of autoimmune diseases and hence the ANA positivity and ANA IIF patterns differ in various geographical regions. Aims and Objectives: This study was aimed to estimate the ANA positivity rates among patients with suspected or confirmed auto immune diseases and to determine the common ANA IIF patterns in the positive samples. Materials and Methods: This is a retrospective study including 235 patients with either clinical suspicion of, or suffering from autoimmune diseases attending a tertiary health care hospital in Puducherry, South India, between November 2018 and April 2020. ANA IIF test (Hep 20-10/ primate liver, Euroimmun, Germany) was performed with 1:100 dilutions of patient serum. Results: The ANA positivity rate was 14.5% (34 out of 235 samples were positive). ANA was positive in 10 (13.3%) males and 24 (15%) females. The mean age of ANA positive patients were 43.4 years.The most common pattern was nucleus fine granular (n=8) followed by cytoplasm fine granular (n=7) and centromere (n=4). Conclusion: A significant ANA positivity rate was found in our study and a detailed clinical study has to be conducted to draw further conclusions. Keywords: Antinuclear antibodies, Indirect immunofluorescence test, ANA patterns.

Bullous pemphigoid (BP) is an autoimmune subepidermal blistering disease characterized by intensely pruritic urticarial plaques and bullae of the skin and, to a lesser extent, the mucosa.1,2 While many cases of BP are determined to be idiopathic, the incidence of BP with various medications resulting in drug-associated bullous pemphigoid (DABP) is on the rise.3 Glucagon-like peptide-1 (GLP-1) agonist-associated BP has been previously reported after administration of dulaglutide.4 Here we report a unique case of BP attributed to semaglutide, another medication in the GLP-1 agonist class.

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.

El megaesófago se presenta entre el 5 % y el 20 % de pacientes con acalasia, un trastorno motor esofágico primario reconocido hace más de 300 años, a considerarse en todo paciente con disfagia no explicada por un proceso obstructivo o inflamatorio luego de un estudio endoscópico detallado. Se presenta el caso de un paciente con disfagia progresiva, en quien se documentó megaesófago como complicación de una acalasia de largo tiempo de evolución, no tratada. Se descartó la enfermedad de Chagas mediante enzimoinmunoensayo (ELISA) e inmunofluorescencia indirecta (IFI), tal como recomiendan las guías actuales. Ante la baja frecuencia de esta entidad en nuestro medio y las implicaciones terapéuticas que tiene para los pacientes con acalasia, se realizó una revisión narrativa en la literatura sobre su diagnóstico y alternativas de manejo.

espanolAntecedentes Los anticuerpos anticitoplasma del neutrofilo (ANCA) se asocian con vasculitis. Existen diferentes metodos para determinar su presencia. Se ha descrito la interferencia de anticuerpos antinucleares (ANA) en la diferenciacion de los patrones P-ANCA y C-ANCA. Objetivo Determinar la frecuencia de ANCA en una poblacion con manifestaciones de enfermedad autoinmune; y evaluar la interferencia de los ANA en su interpretacion. Materiales y metodos Estudio de corte transversal retrospectivo, descriptivo no experimental incluyendo 3.330 datos con diagnostico presuntivo de enfermedad autoinmune y solicitud de ANCA. Las determinaciones de ANCA y de ANA se realizaron mediante inmunofluorescencia indirecta, L-ANCA® y CytoBead® ANCA. Antiproteinasa 3 y antimieloperoxidasa fueron determinados mediante ELISA y CytoBead® ANCA. Resultados Se encontraron ANCA positivos en el 10,21% y el 12,64% con ANCA positivos presentaban ANA positivos. La concordancia kappa para antiproteinasa 3 entre CytoBead® ANCA y ELISA fue del 100% (K = 1; p Conclusiones Los resultados indican una solicitud sobreestimada de este marcador como ayuda diagnostica en consulta de atencion primaria no direccionada. Para una adecuada evaluacion de ANCA se debe implementar la tecnica de inmunofluorescencia indirecta para tamizaje y confirmar con la determinacion de antigenos especificos para antiproteinasa 3 y antimieloperoxidasa por cualquiera de los ensayos confirmatorios. La alta concordancia mostrada por CytoBeads® ANCA hace que planteemos el empleo de dicha alternativa para la determinacion de ANCA y su confirmacion. Dada la interferencia de los ANA, se recomienda solicitar la prueba ANA por inmunofluorescencia indirecta ante la presencia de resultados P-ANCA positivos, con el fin de minimizar «falsos positivos». EnglishBackground 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 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”.

ANCA testing plays an established critical role in the diagnosis of ANCA Associated vasculitis (AAV). The spectrum of diseases associated with positive ANCA has recently broadened, thus calling into question the diagnostic implications of ANCA positivity in a hospital setting. We retrospectively studied all adult patients who had a positive ANCA test (by Indirect Immunofluorescence (IIF), ELISA or both) performed over the span of 19 years. Subjects were then divided into discordant (positive on one assay) and concordant ANCA (positive on both assays) groups based on their ANCA positivity status. The two groups were then compared with regards to their demographic, clinical and laboratory characteristics, the indication for ANCA testing in both groups and their final diagnoses. Of the 9189 ANCA tests ordered during the 19-year span of the study, 389 (4.2%) were positive. Two hundred and forty subjects met the exclusion criteria (patients aged less than 18 years or the lack of clinical and laboratory data in the medical file) thus resulting in a final cohort of 149 subjects. Of them, 122 subjects had discrepant ANCA results and 27 had matching ANCA results. Most cases in the discrepancy group were IIF positive and ELISA negative (86.8%). The diagnosis of AAV was highly unlikely in cases with discrepant IIF and ELISA serologies compared to cases with matching IIF and ELISA serologies (4.1% versus 44.4%, p value

Die Testung auf antinukleare Antikorper (ANA) an humanen Epithelzelllinien (HEp-2) unter Nutzung der indirekten Immunfluoreszenz (IIF) ist von zentraler Bedeutung, wenn Kollagenosen, andere Erkrankungen wie die primar biliare Cholangitis und Autoimmunhepatitis oder auch medikamenteninduzierte ANA ausgeschlossen bzw. diagnostiziert werden sollen. Die ausfuhrliche Beschreibung von 29 ANA-IIF-Mustern im International Consensus on ANA Patterns (ICAP) erleichtert die Vereinheitlichung der ANA-IIF-Diagnostik. Positive ANA-Tests sind bei gesunden Personen haufig und begrunden die Uberweisung an einen Rheumatologen. In diesen Fallen kann der Nachweis von Anti-DFS70-Antikorpern zum Ausschluss systemisch rheumatischer Autoimmunerkrankungen beitragen.

Background West Nile virus (WNV) is one of the widely distributed arboviruses in the world, and it is a pathogen of both humans and animals. The evidence that supports the prevalence of the WNV infection in Ethiopia is very scarce. Hence, this study aimed to assess the seropositivity of WNV among patients with acute febrile illness. Methods This health institution-based descriptive cross-sectional study was conducted on 532 acute febrile patients from May to August 2016 in Arba Minch Zuria district selected public health facilities, Southern Ethiopia. A pre-structured questionnaire was used to collect socio-demographic and clinical related information of the participants through convenient sampling techniques. In addition, trained nurses who were working in the health centers were responsible for interviewing acute febrile patients. About 5 mL of venous blood was collected aseptically from each of the study participants for the screening of the WNV immunoglobulin G (IgG) and immunoglobulin M (IgM) antibodies using indirect immunofluorescence technique (IIFT) as per manufacturer's protocol. Data analysis was done using statistical package for social sciences (SPSS) version 20 software and the results were presented by frequency and percentage using tables. Results A total number of 529 acute febrile patients (42.7% males and 57.3% females) were enrolled in the study with a response rate of 99.4%. The overall 7.4% of acute febrile patients were seropositive for WNV-specific IgG and the rate was higher in males (9.7%) when compared to females (5.6%). While the overall 4.5% were seropositive for WNV-specific IgM and the rate was 6.6% in males and 3.0% in females. Conclusion The finding of this study is an important alarm for clinicians/physicians to diagnose febrile patients in the divergent direction including with the diagnosis of flaviviruses. In addition, the finding will further contribute to understanding the epidemiology of WNV fever in Ethiopia and it will play a role in the delivery of public health measures to decrease the risk of WNV exposure in the areas.

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.

Anti-dense fine speckled 70 (DFS70) autoantibodies were reported to be more prevalent in healthy individuals than those with autoimmune diseases such as systemic lupus erythematosus (SLE). We determined anti-DFS70 autoantibody prevalence in a Latin American cohort of patients with SLE and healthy individuals.This study included 127 individuals with anti-nuclear antibodies (ANAs; 1:160) suggesting the presence of anti-DFS70, including 64 patients with SLE and 63 healthy controls. The anti-DFS70 autoantibodies were determined by immunoadsorption using NOVA LiteThe presence of anti-DFS70 autoantibodies was confirmed by indirect immunofluorescence in 21 (33.3%) healthy controls and 8 (12.5%) patients with SLE (p = 0.005). Among the anti-DFS70-positive patients with SLE, the most frequent compromise was renal involvement in six cases (75%), 4 patients (37.5%) were positive for anti-Sm, which was the most frequently associated antibody, and one patient (12.5%) was positive for anti-DNA.Anti-DFS70 autoantibodies might be considered a biomarker to differentiate patients with SLE from ANA-positive individuals without autoimmune diseases.• In a Latin American cohort, the anti-DFS70 was higher in individuals without autoimmune diseases compared with that in patients with SLE.• The anti-DFS70 might have utility as a biomarker of exclusion in patients with non-specific clinical signs of AARDs.

Objective: To determine the association between Anti-Nuclear Antibody (ANA) pattern in pediatric Systemic Lupus Erythematosus (SLE) patients and proteinuria as a sign of renal involvement in SLE. Methods: This was a cross-sectional study, using data from medical records involving 89 newly diagnosed with SLE (aged ≤ 18 years) in Department of Child Health Dr. Hasan Sadikin General Hospital, Bandung, from January 1st 2018 to June 30th 2019. Data of ANA pattern and proteinuria were collected from medical record. ANA pattern was examined by Immunofluorescence Assay (IFA) method. Chi-square was used to analyze the association between ANA pattern and proteinuria as a sign of renal involvement in pediatric SLE patients. Results: There were 89 patients, consisting of 7 male (7.9%) and 82 female (92.%) with median age of 14 (IQR=12-16). There were only 44 pattern of ANA as follow: homogenous 56.8%, speckled 36.4%, and nucleolar 6.8%. However, there is no significant association between homogenous pattern with proteinuria events (p=0.831). Conclusions: Homogenous pattern was the most frequent ANA pattern in children with SLE, and the pattern has no association with proteinuria events.

Scrub typhus is an acute febrile illness caused by Orientia tsutsugamushi. There are reports of resurgence of scrub typhus in different parts of India including Kerala. Many cases go undiagnosed due to the lack of knowledge regarding the diagnostic test to be used. This study was undertaken at Pushpagiri Institute of Medical Sciences and Research Centre, Tiruvalla in the Department of Microbiology from January 2014 to September 2015, to assess the validity of various tests in diagnosing scrub typhus. A total of 94 samples were subjected to IgM ELISA, Weil Felix (WF) test and Indirect Immunouorescence assay(IFA). The agreement between ELISA, IFA and Weil Felix test was assessed by κ (kappa) coefcient of Cohen using SPSS version 17. The sensitivity and specicity of ELISA with respect to IFA was 100% and 94.67% respectively, and that of IFA with respect to ELISA was 82.61% and 100% respectively. The sensitivity and specicity of WF test with respect to ELISA was 43.48 and 100% respectively and that of WF test with respect to IFA was 52.63% and 100% respectively. Although IgM ELISA is highly sensitive, it cannot be relied upon as the sole diagnostic test due to high false positivity. WF test is not a sensitive test, but when positive it is rather specic. IFA has high sensitivity and specicity, but is unsuitable for moderately equipped laboratories. Hence, we suggest that scrub typhus could be diagnosed using a combination of IgM ELISA and WF test and these tests should be included in the panel of serological tests ordered for patients presenting with undifferentiated febrile illnesses.

Objective: The aim of this study was to test the efficacy of autoantibodies to desmoglein 1 and desmoglein 3 detected by ELISA and indirect immunofluorescence in the diagnosis of oral pemphigus and to correlate the antibody titres with the severity of the disease. Materials and Methods: We report a retrospective cohort study of 22 patients with oral pemphigus and 64 controls from a single tertiary centre. Data about histopathological examination, direct immunofluorescence, indirect immunofluorescence and ELISA were analysed. Global validation of ELISA and IIF both alone and combined was established by calculating sensitivity, specificity, accuracy and both positive predictive value and negative predictive value. The relationship between Oral Disease Severity Score values and ELISA titres was analysed using Pearson's coefficient. Results: The best diagnostic performance was observed for anti-desmoglein 3 ELISA. The sensitivity was 75% and specificity 100% and positive predictive value and negative predictive value were 92.5% and accuracy 93.9%. The level of agreement with histopathology+direct immunofluorescence was substantial (k=.758). Anti-desmoglein 3 titres showed a significant correlation with Oral Disease Severity Score (p&nbsp

Clinicians may encounter a variety of skin conditions that present with vesiculobullous lesions in their everyday practice. Pemphigus vulgaris, pemphigus foliaceus, IgA pemphigus, and paraneoplastic pemphigus represent the spectrum of autoimmune bullous dermatoses of the pemphigus family. The pemphigus family of diseases is characterized by significant morbidity and mortality. Considering the risks associated with a delayed diagnosis or misdiagnosis and the potential for overlap in clinical features and treatment, evaluation for suspected pemphigus disease often requires thorough clinical assessment and laboratory testing. Diagnosis is focused on individual biopsies for histopathology and direct immunofluorescence. Additional laboratory methods used for diagnosis include indirect immunofluorescence and enzyme-linked immunosorbent assay. Recent advancements, including anti-CD20 therapy, have improved the efficacy and reduced the morbidity of pemphigus treatment. This contribution presents updates on the pathophysiology, clinical features, diagnostic work-up, and medical management of pemphigus. Improved strategies for diagnosis and clinical assessment are reviewed, and newer treatment options are discussed.

In this study, we aimed to assess the prevalence of uncommon staining patterns found during testing for the presence of antinuclear antibodies (ANA) and to determine their association with certain antibodies and clinical diagnoses. Presence of ANA and the staining pattern was determined in 10955 samples using indirect immunofluorescence (IIF) on HEp-2 cells. ANA-positive samples were assessed for presence of 14 specific antibody types using a microbead based system. Demographic data (age, sex) and clinical diagnoses were collected from the referral documentation. Particular staining patterns were then compared with a representative comparison group comprised of samples with common staining patterns using these criteria. There were 22 patterns present in less than 3% of samples each and these were jointly present in 42.43% of ANA-positive samples. Specific antibodies were found in proportions similar to the comparison group (46.06%) and varied significantly between patterns. Likewise, there were significant differences in antibody distribution in particular patterns. Some patterns were associated with presence of rheumatic diseases or inflammatory arthropathies, while in others there was a concurrent diagnosis of liver disease, or a neoplastic process. Many of the uncommon IIF patterns have distinctive characteristics that warrant further investigation in order to determine their role in diagnosing various diseases, not limited only to the illnesses of the rheumatic spectrum. IIF on HEp-2 cells remains an irreplaceable method because of the diversity of ANA, only a number of which can be detected using other standardised methods.

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., There is still room for improvement in ANA screening testing. Currently there are different methods available for systemic autoimmune rheumathic disease sift, but the optimization of screening protocols does not only rely on the selection of the best method–there is no perfect test‐ but in the detection of the optimal combination for the specific laboratory routine. In our study we compared the performance of two protocols in our routine samples: solid phase test+IIF and ELISA+IIF. IIF together with EliA CTD Screen (solid phase test) resulted in the highest sensitivity, while EliA CTD Screen showed the best sensitivity/specificity balance.

We have assessed the risk factors for the occurrence of hospital-acquired (HA) and community-acquired (CA) viral acute respiratory tract infections (ARTIs) in children. Children (1–60 months) who were having ARTI on admission (CA) and develops ARTI following 48 h after admission or 3 days of discharge (HA) were included. Indirect immunofluorescence assay (IFA) was performed and multivariable analyses were done to determine the risk factors for the development of viral CA and HA-ARTI. Total of 818 with ARTIs, 226 (27.6%) RSV cases were detected. Out of 226, 86 (38.0%) HA-RSV cases were detected. CA-viral-ARTI was significantly high (p

This study was carried out to evaluate the accuracy of various antibody tests for scrub typhus, namely, the indirect immunofluorescence assay (IFA) from the Korea Centers for Disease Control and Prevention (KCDC) and four commercial kits (companies A to D). The test accuracy was based on the diagnosis of scrub typhus, as defined by a positive PCR or culture. In total, serum samples from 97 patients with scrub typhus and 200 non-scrub typhus patients were tested. The respective sensitivity and specificity of each test were as follows. For the KCDC IFA, sensitivity and specificity were 55.7% (95% confidence interval [CI], 45.2 to 65.8%) and 94.8% (95% CI, 90.4 to 97.3%) for IgM and 42.3% (95% CI, 32.3 to 52.7%) and 96.3% (95% CI, 92.6 to 98.5%) for IgG, with diagnostic cutoffs of ≥1:16 for IgM and ≥1:256 for IgG. For kit A, the sensitivity and specificity were 70.1% (95% CI, 59.8 to 78.8%) and 74.6% (95% CI, 67.6 to 80.6%) for total immunoglobulins, with a cutoff of ≥1:40. For kit B, the sensitivity and specificity were 64.3% (95% CI, 51.9 to 75.1%) and 94.9% (95% CI, 81.4 to 99.1%) for IgM and 67.1% (95% CI, 54.8 to 77.6%) and 74.4% (95% CI, 57.6 to 86.4%) for IgG. For kit C, the sensitivity and specificity were 53.6% (95% CI, 43.2 to 63.7%) and 99.5% (95% CI, 96.8 to 100%) for IgM and 36.1% (95% CI, 26.8 to 46.5%) and 100% (95% CI, 97.6 to 100%) for IgG. For kit D, the sensitivity and specificity were 73.2% (95% CI, 63.1 to 81.4%) and 89.5% (95% CI, 84.2 to 93.2%) for total immunoglobulins. These results are all unsatisfactory, highlighting an urgent need for the development of more highly sensitive and specific tests.

ObjectiveTo compare acute treatment responses and long-term outcome in leucine-rich glioma-inactivated 1 (LGI1) antibody encephalitis.MethodsRetrospective case series of 118 patients with LGI1 antibody encephalitis evaluated at Mayo Clinic across all US sites from 1 May 2008 to 31 March 2019. Patient clinical data were identified and analysed through the neuroimmunology laboratory and electronic medical record. LGI1 antibody detection was by cell-based indirect immunofluorescence assay of serum, cerebrospinal fluid or both. Clinical outcomes were faciobrachial dystonic seizure (FBDS) resolution, modified Rankin Scale (mRS) score, Kokmen Short Test of Mental Status (STMS) score (0–38 point scale) and neuropsychometric testing results.ResultsCompared with intravenous immunoglobulin (IVIg) (n=21), patients treated with single-agent acute corticosteroids (intravenous, oral or both) (n=49) were more likely to experience resolution of FBDS (61% vs 7%, p=0.002) and improvements in mRS score (ΔmRS score 2 vs 0, p=0.008) and median Kokmen STMS scores (ΔKokmen STMS score 5 points vs 0 points, p=0.01). In 54 patients with long-term follow-up (≥2 years), the median mRS score was 1 (range 0–6) and the median Kokmen STMS score was 36 (range 24–38) after all combinations of immunotherapy. Neuropsychometric testing in 32 patients with long-term follow-up (≥2 years) demonstrated short-term memory impairments in 37%.ConclusionsCorticosteroids appeared more effective acutely than IVIg in improving LGI1 antibody encephalitis in this retrospective comparison of immunotherapies. While improvement with immunotherapy is typical and long-term outcome is favourable, short-term memory deficits are noted in approximately a third of the patients.

Longevity, high prevalence, and multifocal distribution of this disease pose unprecedented public health challenges., Epidemic levels of Rocky Mountain spotted fe­­­ver (RMSF) have persisted in Mexicali, Mexico, since the initial outbreak was first reported in December 2008. We compared clinical and epidemiologic data of cases in Mexicali during 2009–2019 between patients with an IgG titer reactive with Rickettsia rickettsii bacteria by indirect immunofluorescence antibody (IFA) assay and those who demonstrated DNA of R. rickettsii in a whole blood sample when tested by PCR. We identified 4,290 patients with clinical and epidemiologic features compatible with RMSF; of these, 9.74% tested positive by IFA and 8.41% by PCR. Overall, 140 patients died (11-year case-fatality rate 17.97%). Substantial differences in the frequency of commonly recognized clinical characteristics of RMSF were identified between PCR-positive and IFA-positive cases. The Mexicali epidemic is unique in its size and urban centralization. Cases confirmed by PCR most accurately reflect the clinical profile of RMSF.

Objective We studied the clinical phenotypes and prevalent genotypes of Orientia tsutsugamushi in our area using indirect immunofluorescence assay (IFA). Methods We prospectively screened all febrile children presenting to our hospital over three years. From among children who were scrub typhus positive by ELISA we selected a sample of convenience for IFA testing to determine the genotypes of O. tsutsugamushi using four strains namely Boryong, Gilliam, Karp and Kato. Results Of all scrub positive patients (n=77), we tested 14 samples using IFA and all 14 samples were IFA positive. Karp genotype (n=7) was most prevalent followed by Kato (3), Boryong (1) and Gilliam (1) genotypes; 2 patients were positive for mixed genotype. There was high prevalence of organ dysfunction among IFA positive children. Three most common organ dysfunctions included hematological derangement in all, liver involvement in 10 (71%), and encephalopathy and shock in 4 each. Conclusions Karp was the most prevalent genotype of O. tsutsugamushi in our area.

Author(s): Abel, Mary Kathryn; Suresh, Raagini; Raffi, Jodie; McCalmont, Timothy H; Murase, Jenny E

BACKGROUND Bullous pemphigoid (BP) is an autoimmune blistering disease associated with autoantibodies against BP180 and/or BP230 antigens. The immunoassays available for serological diagnostics include indirect immunofluorescence (IIF) on monkey esophagus (ME), salt-split skin (SSS), and enzyme-linked immunosorbent assay (ELISA) for BP180-NC16a and BP230. Only a few studies validated innovative BIOCHIP mosaic, but none compared agreement between BIOCHIP substrates with conventional methods separately. METHODS We evaluated the agreement between BIOCHIP and conventional methods and assessed sensitivity and specificity in BP diagnosis. The study comprised 51 BP patients and 39 controls. RESULTS Analysis showed very good agreement between BIOCHIP-SSS vs classic IIF-SSS (0.933, P

Objectives: This study aims to evaluate the interpretation of the antinuclear antibody (ANA)-indirect immunofluorescence (IIF) test results based on the interpreter-related subjectivity and to examine the inter-center agreement rates with the performance of each laboratory. Patients and methods: The ANA-IIF testing was carried out in a total of 600 sera and evaluated by four laboratories. The inter-center agreement rates were detected. The same results given by the four centers were accepted as gold standard and the predictive values of each center were calculated. Results: The inter-center agreement was reported for ANA-IIF test results from 392 of 600 (65.3%) sera, while 154 of 392 results were positive. Four study centers reported 213 (35.5%), 222 (37.0%), 266 (44.3%), and 361 (60.2%) positive test results, respectively. In terms of the patterns, the highest and lowest positive predictive values were 72.3% and 42.7%, respectively, while the highest and lowest negative predictive values were 99.6% and 61.5%, respectively. The agreement for semi-quantitative evaluation at three levels of fluorescence intensity stated by four centers was detected in 100 sera at 87% 3(+), while the other two levels were 6% and 7%. The highest predictive value for the highest fluorescence intensity of 3(+) was found to be 71.9%. Conclusion: Significant differences may be observed among laboratories in terms of qualitative results, patterns, and semi-quantitative determination of the fluorescence intensity in the ANA-IIF testing, particularly at low fluorescence intensity levels and in those with speckled patterns. In case of any discrepancy between ANA-IIF test and clinical prediagnosis, the test should be repeated in another laboratory, if necessary. © 2019 Turkish League Against Rheumatism.

Objective Anti-golgi antibodies (AGAs) are rarely encountered and often considered in relation to autoimmune diseases in clinical practice. This research was performed for studying the prevalence and clinical significance of AGAs in Chinese population. Methods We retrospectively reviewed 22,619 laboratory reports of AGAs detected by indirect immunofluorescence (IIF) were consecutively collected from the First People's Hospital of Wenling between June 2012 and June 2017. Eight patients with AGAs were followed up for relevant clinical and laboratory characteristics. Result A total of 22,619 laboratory reports were collected. Of 19 patients with AGAs, 7 cases (all females) had autoimmune diseases (AID) and 12 cases (6 females and 6 males) had non-AID. High titer AGAs ranging from 1:1000 to 1:3200 were persistently present in AID patients, while low-titer AGAs ranging from 1:100 to 1:320 were transient in non-AID patients. Conclusion This is the first study to assess the AGA positive rate and relevant clinical manifestations in a hospitalized Chinese population. AGAs were rare and occurred in a variety of diseases. They were persistently strongly positive in AID, whereas low-titered and transient in non-AID.

The concomitant occurrence of psoriasis vulgaris (PV) and bullous pemphigoid in a patient is rare. We report a 55-year-old male, with history of PV since 4 years, on irregular topical medication, who developed multiple fluid-filled lesions all over the body. A combination treatment with prednisolone, cyclosporine, and dapsone followed by methotrexate was proved suitable and effective.

Background: Nonbullous pemphigoid is an under-recognized phenotype of the autoimmune bullous disease pemphigoid, characterized by the absence of blisters. Several disease aspects have not been studied previously.Objective: To describe the characteristics of nonbullous pemphigoid.Methods: A retrospective review study of medical records. The diagnosis of pemphigoid was based on meeting 2 of the following 3 criteria: (1) pruritus, (2) positive direct immunofluorescence microscopy, or (3) positive indirect immunofluorescence microscopy on salt-split skin.Results: The review included 69 patients. The mean delay in diagnosis was 29 months. Skin examination most often showed pruritic papules/nodules (37%) or pruritus without primary skin lesions (22%). Histopathologic findings were mainly nonspecific. Results of direct and indirect immunofluorescence microscopy were positive in 60% and 69%, respectively. During follow-up, blisters formed in 17%, which was associated with a positive indirect immunofluorescence microscopy (P = .014) and a positive BP180 immunoblot result (P = .032). The Kaplan-Meier estimates of mortality at 1, 2, and 3 years were 14%, 34%, and 46%, respectively, with an 8.6-fold increased all-cause mortality risk.Limitations: The retrospective study design.Conclusions: Nonbullous pemphigoid presented with heterogeneous pruritic skin lesions, resulting in delayed diagnosis. Direct and indirect immunofluorescence microscopy are essential to diagnose nonbullous pemphigoid, in contrast to histopathology, mainly showing nonspecific findings. An increased all-cause mortality risk was observed during follow-up.

Background When the dense fine speckled (DFS) pattern-antinuclear antibodies (ANA) are detected in the indirect immunofluorescence (IIF) assay, the presence of anti-dense fine speckles 70 (DFS70) antibodies has been suggested to facilitate the exclusion of ANA-associated rheumatic diseases (AARD). We evaluated the potential use of anti-DFS70 antibodies for verifying AARD in patients with a positive ANA result of the DFS pattern. Methods A total of 5509 patients who were requested ANA testing were included. The DFS pattern was confirmed using two IIF assays. Semiquantitative DFS70 ELISA (Euroimmun, Germany) was examined in samples with the DFS pattern. Results Among 639 ANA-positive patients, 19.6% displayed the DFS pattern. And 17.6% of patients with the DFS pattern were diagnosed with AARD. The low titer of 1:80 was more prevalent in the non-AARD group than in the AARD group (64.1% vs. 4.5%, p Conclusions The DFS pattern is present in both AARD and non-AARD cases. In the DFS pattern, a low titer of 1:80 and isolated anti-DFS70 antibodies without AARD-associated antibodies represent a low likelihood of AARD. The presence of anti-DFS70 antibodies cannot exclude AARD and should be analyzed in combination with AARD-associated antibodies in the diagnostic algorithm.

Resumen Introducción. La leptospirosis representa un problema de salud pública y es una causa importante de morbimortalidad en la región de Urabá, cuya notificación se ve afectada por las deficiencias en el diagnóstico. Objetivo. Establecer la incidencia de la leptospirosis en los municipios del llamado ‘eje bananero’ de la región de Urabá, documentar la magnitud del subregistro y proponer orientaciones para el diagnóstico por laboratorio por parte de la red de salud pública. Materiales y métodos. Se compararon dos fuentes de información sobre la leptospirosis: el sistema oficial nacional de vigilancia y un estudio transversal de 479 pacientes febriles, llevado a cabo entre abril de 2010 y mayo de 2012. El diagnóstico se hizo con base en tres pruebas: inmunofluorescencia indirecta, microaglutinación y hemocultivo. La exhaustividad de cada fuente de información se estimó mediante el método de captura y recaptura. Resultados. El 58 % (278/479) de los pacientes fueron positivos para leptospirosis, por lo menos, en una de las pruebas y, el 10,43 % (29/278), en las tres. La inclusión de una cepa nativa en el panel de la prueba de microaglutinación aumentó el porcentaje de positividad en 15 %. La tasa acumulada de incidencia fue de 66,5 por 100.000 habitantes y la proporción de letalidad fue de 2,15 %. El subregistro de la morbilidad por leptospirosis en la región de Urabá, fue de 27,8 % y, el de la mortalidad, de 66,6 %. Conclusión. El subregistro de leptospirosis en la región reitera la necesidad de usar más de una prueba diagnóstica para identificar Leptospira spp. en pacientes de zonas endémicas. Este subregistro podría ser una situación común en todo el país. Abstract Introduction: Leptospirosis represents a public health problem and is a significant cause of morbidity and mortality in the region of Urabá. However, its notification reveals diagnostic limitations. Objective: To establish the incidence of leptospirosis in the municipalities of the so-called eje bananero in the Urabá region, to describe the magnitude of underreporting, and to propose guidelines for laboratory diagnosis by the public health network. Materials and methods: Two leptospirosis information sources were used: The national official surveillance system and a cross-sectional study of 479 acute-phase patients from April, 2010, to May, 2012. The diagnosis was made using three different tests: Indirect immunofluorescence, microagglutination test, and blood cultures. The exhaustiveness percentage of each information source was calculatedusing thecapture and recapture test. Results: From the total number of cases, 58% (278/479) were positive for leptospirosis at least by a test and 10.43% (29/278) of cases were positive by all three methods. The inclusion of a native strain in the microagglutination test panel increased the percentage of positivity by 15%. The cumulative incidence rate was 66.5/100,000 inhabitants and the case fatality ratio was 2.15%. The underreporting rates of leptospirosis in the Urabá region were 27.8% in morbidity and 66.6% in mortality. Conclusion: Under-registration of leptospirosis in the region highlights the necessity to use more than one diagnostic test to identify Leptospira in patients from endemic areas. Under-registration could be a common situation throughout the country.

Objective: The aim of this study was to investigate the predictive value of positive enzyme immunoassay (EIA) for detection of specific autoantibodies in sera negative for antinuclear antibody (ANA) by indirect immunofluorescence (IIF) assay, but positive for ANA by EIA. Methods: Eighty sera that tested negative for ANA by IIF, but positive for ANA by EIA were included. All sera were tested for specific autoantibodies by line immunoassay (LIA). The positive predictive value (PPV) of EIA was calculated using LIA result as a reference standard. Medical records of patients were reviewed. Clinical findings at the time of blood sampling for ANA testing and at 5 years after sampling were obtained. Results: Twenty-eight sera (35%) were found to be positive by LIA. The PPV of EIA for detection of specific autoantibodies at the manufacturer’s recommended cut-off was 35.0% (95% CI: 24.5-45.5%). The most prevalent antibodies were anti-SSA/Ro60 (64.3%, 95% CI: 46.5-82.0%), anti-Ro52 (25.0%, 95% CI: 9.0-41.0%), and anti-SSB/La (10.7%, 95% CI: 0-22.2%). A diagnosis of systemic autoimmune rheumatic disease was established in 7 patients (25%) at the time of blood sampling, and 4 patients (14.3%) were diagnosed with non-rheumatic autoimmune disease. Conclusion: EIA testing in IIF-ANA negative sera yielded a chance to detect antinuclear antibodies. However, the poor PPV of EIA may have low benefit in real-life clinical practice. Anti-SSA/Ro60 was the most prevalent antibody detected. A high proportion of LIA-ANA positive patients were not diagnosed as autoimmune disease at the time of antibody detection.

Aim: to present a clinical case of linear IgA bullous dermatosis.Materials and methods. A 44 years old patient complaining of skin rashes on the trunk and extremities, accompanied by severe itching was examined. We carried out a morphological investigation of biopsy samples derived from the lesion and apparently unaffected skin areas using the method of indirect immunofluorescence.Results. The patient having rashes in the form of multiple vesicles and small bubbles with a tight cover, which had been grouped into figures resembling pearl necklaces, demonstrated the presence of focal subepidermal cracks (subepidermal bubble in one location), as well as a linear deposition of IgA along the epidermal basement membrane. According to the clinical picture and following the histological and immunofluorescent investigation of skin biopsies, the patient was diagnosed with linear IgA bullous dermatosis. Lesion regression was achieved as a result of systemic therapy with prednisolone at a dose of 50 mg per day.Conclusion. The diagnosis of linear IgA bullous dermatosis should be made on the basis of skin biopsy investigation by the method of indirect immunofluorescence. Systemic glucocorticosteroids are seen as an effective approach to the treatment of such patients.

Importance: A substantial number of patients with bullous pemphigoid do not develop skin blisters and may not have received the correct diagnosis. Diagnostic criteria and an optimal diagnostic strategy are needed for early recognition and trials.Objectives: To assess the minimal requirements for diagnosis of bullous and nonbullous forms of pemphigoid and to evaluate the optimal diagnostic strategy.Design, Setting, and Participants: This paired, multivariable, diagnostic accuracy study analyzed data from 1125 consecutive patients with suspected pemphigoid who were referred to the Groningen Center for Blistering Diseases from secondary and tertiary care hospitals throughout the Netherlands. Eligible participants were patients with paired data on at least (1) a skin biopsy specimen for the direct immunofluorescence (DIF) microscopy test; (2) indirect immunofluorescence on a human salt-split skin substrate (IIF SSS) test; and (3) 1 or more routine immunoserologic tests administered between January 1, 2002, and May 1, 2015. Samples were taken from patients at the time of first diagnosis, before introduction of immunosuppressive therapy, and within an inclusion window of a maximum of 4 weeks. Data analysis was conducted from October 1, 2015, to December 1, 2017.Main Outcomes and Measures: Pairwise DIF, IIF SSS, IIF on monkey esophagus, BP180 and BP230 enzyme-linked immunosorbent assays, and immunoblot for BP180 and BP230 tests were performed. The results were reported in accordance with 2015 version of the Standards for Reporting Diagnostic Accuracy.Results: Of the 1125 patients analyzed, 653 (58.0%) were women and 472 (42.0%) were men, with a mean (SD) age of 63.2 (19.9) years. In total, 343 participants received a pemphigoid diagnosis, with 782 controls. Of the 343 patients, 74 (21.6%, or 1 in 5) presented with nonbullous pemphigoid. The DIF microscopy was the most sensitive diagnostic test (88.3% [n = 303]; 95% CI, 84.5%-91.3%), whereas IIF SSS was less sensitive (77.0% [n = 263]; 95% CI, 72.2%-81.1%) but was highly specific (99.9%; 95% CI, 99.3%-100%) and complemented most cases with negative DIF findings. Results of the BP180 NC16A enzyme-linked immunosorbent assay did not add diagnostic value for initial diagnosis in multivariable logistic regression analysis of combined tests. These findings lead to the proposed minimal criteria for diagnosing pemphigoid: (1) pruritus and/or predominant cutaneous blisters, (2) linear IgG and/or C3c deposits (in an n-serrated pattern) by DIF on a skin biopsy specimen, and (3) positive epidermal side staining of IgG by IIF SSS on a serum sample; this proposal extends bullous pemphigoid with the unrecognized nonbullous form.Conclusions and Relevance: Both DIF and IIF SSS tests should be performed for diagnosis of the bullous and nonbullous variants of pemphigoid, and the BP180 NC16A enzyme-linked immunosorbent assay is recommended as an add-on test for disease activity monitoring.

Background The introduction of automated anti-nuclear antibody (ANA) indirect immunofluorescence (IIF) analysis may allow for more harmonized ANA IIF reporting, provided that a thorough quality assurance program controls this process. The aim of this study was to evaluate various quality indicators used for ANA IIF analysis with the final goal of optimizing the iQC program. Methods In an experimental setup, we introduced artificial errors, mimicking plausible problems during routine practice on a QUANTA-Lyser-NOVA View® system (Inova Diagnostics, San Diego, CA, USA). Predetermined quality indicators were evaluated against predefined acceptance criteria. In addition, we retrospectively investigated the applicability of the selected quality indicators in the daily routine practice during three pre-defined periods. Results Both the experimental as the retrospective study revealed that pre-analytical, analytical and post-analytical errors were not highlighted by company internal quality control (iQC) materials. The use of patient derived iQC samples, median fluorescence intensity results per run and the percentage of positive ANA IIF results as additional quality indicators ensured a more adequate ANA IIF quality assurance. Furthermore, negative and moderate positive sample iQC materials merit clinical validation, as titer changes of >1 correspond to clinically important shifts. Traditional Westgard rules, including a clinically defined stop limit, revealed to be useful in monitoring of the supplemental quality indicators. Conclusions A thorough ANA IIF quality assurance for daily routine practice necessitates the addition of supplemental quality indicators in combination with well-defined acceptance criteria.

Pemphigus comprises a heterogeneous group of autoimmune blistering diseases, which can affect both skin and mucous membranes, especially oral mucosa. This group of diseases shows usually a chronic-relapsing course. Since pemphigus is a rare disease, the diagnosis is often delayed, because it is based upon the recognition of consistent clinical, histologic, and direct immunofluorescence findings, as well as indirect immunofluorescence, and/or enzyme-linked immunosorbent assay. Usually the patients are treated for multiple other conditions before starting the correct therapy, leading to a critical reduction of the patients' quality of life. This review is a succinct compilation of pearls gathered from clinical experience in pemphigus and the myths that may have influenced everyday practice but have been proven false. This review provided a selection of such dilemmas and controversies, focusing on myths and pearls that can help young dermatologist in the clinic, while also dispelling them.

To date, only a few studies have analyzed the clinical characteristics and genetic features of human granulocytic anaplasmosis (HGA) in South Korea. Thus, in this study, we investigated the clinical characteristics of HGA and methods used for clinical diagnosis. The clinical characteristics of patients with HGA were studied retrospectively. We reviewed the medical charts of 21 confirmed patients with HGA admitted to the Chosun University Hospital, located in Gwangju, South Korea. Twenty-one HGA patients visited the hospital 2–30 days (median 7 days) after the onset of symptoms. Fourteen patients (66.7%) had fever, which was alleviated 2 h (range 0–12.75 h) after starting treatment with doxycycline. Of the 18 patients who underwent peripheral blood (PB) smear test, only one (5.6%) had morulae. Additionally, only 4/17 patients (23.5%) had morulae in the PB smear reconducted after the confirmation of anaplasmosis. All 21 patients recovered without significant complications. As per results of the blood tests conducted at the time of admission, 7/21 (33.3%) and 5/21 (23.8%) patients showed at least 1:16 and 1:80 of IgM and IgG titers, respectively. Most HGA patients in Korea recovered without significant complications. The indirect immunofluorescence antibody diagnosis or morulae identification for HGA in this study had low sensitivity in the early stage of the disease.

Background and Aims Antibodies to M-type phospholipase A2 receptor (PLA2R-Ab) are considered to be a promising biomarker for laboratory diagnosis of primary membranous nephropathy (PMN) and may be useful in the evaluation of the response to therapy and CKD prognosis. The aim of the study was to compare two immunoassay methods – indirect immunofluorescence (IIF) and enzyme immunoassay (ELISA) for the determination of circulating PLA2R-Ab in patients with PMN. Method The study included 54 patients aged 55 (40-63) yrs. (M: F [33:21]) with PMN before treatment (n=16) and treated with immunosuppressive therapy (IST) (n=38), and apparently healthy individuals of the corresponding gender and age (n=10). Proteinuria and estimated glomerular filtration rate (eGFR) were determined in all participants. The levels of PLA2R-Ab were determined by IIF and quantitative/ semi-quantitative ELISA (EURUIMMUN AG test, Germany). In 16 PMN patients without treatment and 28 PMN patients treated with IST the level of PLA2R-Ab was measured one time and in 10 PMN patients treated IST – in dynamic, from 2 to 5 times. Statistical comparisons among groups were performed using Mann–Whitney U-test and Kruskal-Wallis H tests. The association between variables was estimated using Spearman’s coefficient. Sensitivity and specificity of the methods were calculated. Results The correlation coefficient between IIF and ELISA was 0.82 (p Conclusion Both quantitative and semi-quantitative ELISA were considered to be more preferable methods since the obtained results correlate with renal dysfunction and allow to assess the concentration of PLA2R-Ab in the course of disease more accurately, that may contribute to timely correction of treatment and improvement of outcome.

Background Although tick-borne pathogens have been reported as an important cause of imported fever, the incidence of Anaplasma phagocytophilum, the causative agent of human granulocytic anaplasmosis (HGA), in travellers is unknown. Methods We conducted a prospective cohort study to investigate the aetiologies of fever in returning travellers (November 2017—July 2019). Polymerase chain reaction for msp2 gene amplification and indirect immunofluorescence assay for A. phagocitophilum were performed in all returning travellers with undifferentiated non-malarial fever. Results Among 141 travellers included, 8 patients were diagnosed with probable or confirmed HGA. The overall incidence rate of HGA was 19.9 cases/1000 person—week of travel. The main destination of travel was Asia, accounting for 62.5% patients with HGA. Co-infections were found in 37.5% of patients with HGA. Conclusions Diagnosis of HGA and empirical treatment with doxycycline should be considered in travellers with fever.

Congenital Chagas disease (CCD) has become a global health problem. Historically, the diagnosis of CCD has been carried out using parasitological methods and traditional serological techniques, however, new serological techniques such as chemiluminescent microparticle immunoassays (CMIA) have been developed in the last few years with many advantages compared with traditional serological tests. A total of 75 children born to 72 Latin American Chagas-infected mothers were consecutively enrolled and studied by CMIA and indirect immunofluorescence (IIF) at 0–2, 6, 9, and 12 months of age. At the end of the follow-up, 74 out of 75 children were considered uninfected and one child was diagnosed with CCD. Our study emphasizes the need to carry out serological follow-up on every newborn from a mother with Chagas disease and shows that CMIA assay is a great diagnostic tool as a single serological test at 9 months of age to rule out CCD or to identify possible transmission.

Objective To evaluate the perception of rheumatologists regarding the recommendations of the Brazilian Consensus for detection of Autoantibodies (BCA) on HEp-2 Cells by Indirect Immunofluorescence assay (IFA) and how BCA recommendations help in clinical practice. Methodology A structured questionnaire regarding the BCA recommendations for detection and interpretations of autoantibodies in HEp-2 cells was applied to randomly selected rheumatologists. The results were tabulated using the Microsoft® Excel program, expressed as a simple percentage and the dichotomous data were analyzed using the Chi-square test and the Epi Info® program. Results Four hundred fuorteen rheumatologists participated in the study: 70% of them considered their knowledge of the HEp-2 IFA test satisfactory or excellent, and 43% said they knew the BCA recommendations in general, without distinguishing the edition of the BCA to which they refer. The Revista Brasileira de Rheumatologia/Advances in Rheumatology was the means of dissemination most consulted by specialists (50%). According to the rheumatologists’ opinion, the most relevant pattern was the homogeneous nuclear (78%) and 65% stated they were satisfied with the BCA recommendations at a level of satisfaction greater than or equal to 80%. There was no significant difference in the perception of rheumatologists from the several Brazilian geographic regions. Conclusion Brazilian rheumatologists are aware of the BCA guidelines and most are satisfied with the content published, considering that the BCA recommendations assist positively in the clinical practice. Most rheumatologists recognize the patterns associated with rheumatic autoimmune diseases and have used BCA recommendations to interpret the results of the HEp-2 IFA test.