Positive Indirect Immunofluorescence Research Articles

Clinical features of anti-SAE1 antibody-positive myositis and interstitial lung disease: a multicenter, retrospective study in Taiwan.

The clinical characteristics of patients positive for anti-small ubiquitin-like modifier 1-activating enzyme subunit 1 (SAE1) antibodies and diagnosed with idiopathic inflammatory myopathies (IIMs) vary across different cohorts and ethnicities, particularly concerning interstitial lung disease (ILD). We aimed to assess the clinical utility of the line immunoblot assay (LIA) in detecting anti-SAE1 autoantibodies and evaluate the clinical relevance and chronology of ILD development in relation to SAE1 autoantibody positivity among Taiwanese patients. We retrospectively conducted a population-based cohort analysis involving 6,496 patients who visited Chang Gung Memorial Health System across Taiwan from May 2018 to December 2021. Patients were assayed for myositis-specific autoantibodies (MSAs) and myositis-associated autoantibodies (MAAs) using the LIA method, and the antinuclear antibody (ANA) indirect immunofluorescence (IIF) method was used to evaluate ANA patterns. Of these, 70 SAE1-positive patients (1.08%) were included and followed up until December 2023. Associations with clinical characteristics and final diagnosis, particularly ILD, were assessed. Among the 70 SAE1-positive patients, 10 (14.3%) were strongly positive and 60 (85.7%) were weakly positive. In the strong positive group, 70% (7/10) were diagnosed with IIM, with most (5/7) showing a concordant ANA IIF pattern (speckled type). Six patients presented ILD either before (1/6) or after (5/6) IIM diagnosis; the majority (4/6) were classified as organizing pneumonia. The remaining 30.0% (3/10) had connective tissue disease (CTD) other than IIM without detectable ILD during follow-up, and none demonstrated a concordant ANA IIF pattern. In the weakly positive group, only 5.0% (3/60) had IIM and 3.3% (2/60) had ILD. The positive predictive value for strong positive SAE1 autoantibodies in diagnosing IIM was significantly higher than for weak positives (70.0% vs. 5.0%; p < 0.001). The study suggests that strong positive SAE1 autoantibodies detected via LIA are more closely associated with IIM compared to weak positive results. A high prevalence of ILD was observed among strong positive Taiwanese patients, indicating the need for prompt screening. Patients with weak positive or discordant ANA IIF results may represent false positives with a lower ILD risk.

Discordant indirect immunofluorescence compared to multiplex bead immunoassay in ANCA associated vasculitides, a retrospective analysis

Abstract Based on the revised 2017 international consensus on testing for anti-neutrophil cytoplasmic antibodies (ANCA) in granulomatosis with polyangiitis (GPA) and microscopic polyangiitis (MPA), we conducted a retrospective analysis to assess whether testing algorithm modification would be appropriate in our laboratory. Currently, at our institution, the primary screening method for ANCA associated vasculitides (AAV) remains indirect immunofluorescence (IIF), which is reflexed to immunoassay (IA) for confirmation. IA is performed by multiplex bead technology to detect anti-myeloperoxidase (MPO) and anti-proteinase 3 (PR3) antibodies. We reviewed cases from January 1, 2021, to November 30, 2022 using Softlab (laboratory information system) with the aim to find discordant results between IIF and IA (IIF-positive and IA-negative, and IA-positive and IIF-negative). We limited our analysis to in-house patients with clinical information available (EPIC electronic medical record). 204 cases met the criterion for positive IIF and negative IA (positive IA defined as index value &amp;gt;0.9), of which 5% were close to the cut off index value (0.7-0.9). 64% of these 204 patients were never concordant in results available for review (IIF-positive, IA-negative). Potential confounding factors for these never concordant cases include 44% historically or concurrently ANA-positive (10% with clinical diagnosis of lupus), 11% with inflammatory bowel disease, and 10% with an underlying ophthalmologic diagnosis, both overlapping with positive ANA. Notably 36% of the patients who were IIF negative over this period had historically positive MPO or PR3 testing, and the current testing window may reflect higher sensitivity of IIF methods, which in this population includes index values near cutoff but definitionally negative. In contrast, of 247 cases with IA-positive results, 51 cases were discordant with IIF results (IA-positive, IIF-negative). 57% of the discordant cases had a historical diagnosis of AAV who were receiving treatment or in remission. 34% of these AAV discordant cases were near the cut off index value (1.0-1.2). Furthermore, we found that all patients with a clinical diagnosis of AAV were monitored by serial laboratory testing at varying frequency, which included IIF with titers and IA (at least once). As there are no explicit guidelines for laboratory testing to monitor AAV patients, it is unclear whether ordering both assays is of clinical utility in management. With advantages of efficiency and faster turnaround time, IAs may be considered as the primary screening method. In our patient population, we did not identify patients who were IIF-positive only with clinically meaningful disease. Although there may be clinical scenarios that warrant to perform both assays simultaneously, for those with GPA and MPA we conclude that it may be appropriate to transition to use IAs as the primary screening method and use IIF as a confirmatory assay, which may not need to be repeated if positive once.

Negative ANA-IIF in SLE patients: what is beyond?

The antinuclear antibody (ANA) test has high sensitivity in diagnosing and classifying systemic lupus erythematosus (SLE).ObjectivesTo describe the immunological pattern of SLE patients through investigating specific antinuclear autoantibodies by enzyme dot immunoassay and studying their frequency in both positive and negative ANA indirect immunofluorescence assay (IIF) cases.MethodsIn a cross-sectional study, blood samples from 393 newly diagnosed SLE patients were analyzed using (IIF) on HEp-2 cells and ANA dot immunoassay by automated enzyme immunoassay (EIA) to detect 19 antibodies.ResultsNinety-one percent of the patients are females; their mean age was 37 ± 12.28. Antinuclear antibody (ANA) was detected by IIF in 82.4% of cases, with 181 (46.1%) speckled and 167 (42.4%) homogeneous ANA patterns. The majority of patients (96%) demonstrated autoantibodies via EIA. Among the ANA-IIF-negative patients, 97.2% demonstrated autoantibodies. There was a significant difference in the frequency of certain autoantibodies between SLE patients with negative and positive ANA-IIF (1.44 0.73, 3.12 2.09, p = 0.00) respectively.ConclusionThe results of analyzing 19 autoantibodies with the ANA staining pattern increased the significance of analyzing the immune profile even if IIF is negative when clinical symptoms strongly suggest SLE diagnosis. Certain autoantibodies may evade staining by the IFA approach while they are present in the patient’s serum, and they may not be detected by the ANA EIA profile if it does not contain that antigenic substrate.Key Points• Indirect immunofluorescence on Hep-2 is the conventional method for ANA detection and is regarded as the “gold standard” for testing in clinical practice for SLE.• In our study, ANA profile dot enzyme immunoassay (EIA)-based test was performed to evaluate 19 autoantibodies in SLE patients either positive or negative for ANA-IIF.• The presence of anti-dsDNA with ANA-IIF-negative serum in 32.4% of SLE patients provides evidence that not all anti-dsDNA antibodies are identified on standard HEp-2 substrates.• certain autoantibodies can evade staining by the ANA-IIF method despite being present in the SLE patient’s blood; this supports the ANA profile enzyme dot immunoassay as a more sensitive test.

Anti-Ki/anti-PA28γ autoantibodies contribute to the HEp-2 indirect immunofluorescence nuclear speckled pattern.

Antinuclear antibodies (ANAs) are associated with several autoimmune diseases. Indirect immunofluorescence (IIF) on human epithelial type 2 (HEp-2) cells is the golden standard for ANA detection in the clinic. In case of a positive HEp-2 IIF test result, follow-up tests are done to determine autoantibody specificity. For a fraction of the HEp-2 IIF-positive samples, the nature of the autoantigens remains uncharacterized. Our objective was to characterize autoantigens in such samples. To characterize autoantigens in an unbiased way, we combined protein immunoprecipitation with liquid chromatography (LC) tandem mass spectrometry (MS/MS) sequencing. Using such approach we detected the Ki antigen, also referred to as PA28γ, in the immunoprecipitate of serum samples of three individuals with an autoimmune disease. The HEp-2 nuclear speckled IIF fluorescent signal of all three serum samples was abolished after pre-absorption of the serum with recombinant Ki antigen, confirming that autoantibodies against Ki underlie the HEp-2 IIF signal. Our data suggest that anti-Ki autoantibodies can underlie a nuclear speckled HEp-2 IIF pattern.

Occurrence of autoantibodies against skin proteins in patients with hereditary epidermolysis bullosa predisposes to development of autoimmune blistering disease.

Skin blistering disorders are associated with inherited defects in proteins involved in the dermal-epidermal adhesion or autoantibodies targeting those proteins. Although blistering in hereditary epidermolysis bullosa (EB) is pathogenetically linked to genetic deficiency of distinct proteins of the epidermis or the dermal-epidermal junction, circulating autoantibodies against these proteins have also been identified in EB patients. So far, autoantibodies have been considered bystanders in EB and active pathogenicity of them in EB has not been disclosed. In sera of a cohort of 258 EB patients, we found by ELISA in 22% of the patients autoantibodies against the bullous pemphigoid antigen BP180. The titers correlated negatively with collagen VII skin expression and positively with disease severity. Among those patients, we identified six (2.33%) with clinical features of an autoimmune bullous disorder (AIBD) and positive indirect immunofluorescence (IIF) staining. In literature, we found four more cases of EB patients developing disease-aggravating AIBD. Co-existence of these two rare skin disorders suggests that EB patients have a predisposition for the development of AIBD. Our work highlights that EB patients with increased itch or blister formation should be evaluated for additional AIBD and repeated screening for changes in autoantibody titers and skin-binding specificities is advised.

Prognostic factors for mortality in bullous pemphigoid: A systematic review and meta-analysis.

ObjectiveTo systematically evaluate the prognostic factors for mortality in bullous pemphigoid.MethodsPubMed, Embase, Cochrane Library, China National Knowledge Infrastructure, China Biology Medicine disc and Wanfang Database were searched to collect literature on the prognostic factors for mortality in bullous pemphigoid. The quality of studies was assessed by Newcastle–Ottawa Quality Assessment Scale. Two researchers extracted relevant data and scored study quality independently. The hazard ratio (HR) was calculated using the random effects model. Study heterogeneity was assessed using both Cochran’s Q test and I2 statistics. The causes of heterogeneity were assessed by subgroup analysis and/ or sensitivity analysis when heterogeneity was significant. When ten or more studies were included as outcome indicators, publication bias was evaluated by funnel plot and Egger’s test.ResultsOut of a total of 1,546 articles retrieved, 15 studies involving 2,435 patients were included. The meta-analysis showed that the mortality of patients with bullous pemphigoid increased with positive bullous pemphigoid 180 antibody (HR = 1.85, 95%CI: 1.25~2.75, P = 0.002); concomitant dementia (HR = 2.26, 95%CI: 1.43~3.59, P<0.001); stroke (HR = 2.09, 95% CI: 1.23–3.55, P = 0.007); heart disease (HR = 1.96, 95% CI: 1.41–2.73, P<0.001) and diabetes mellitus (HR = 2.39, 95% CI: 1.55–3.69, P<0.001). Sex, positive indirect immunofluorescence and hypertension were not associated with prognosis.ConclusionPositive bullous pemphigoid 180 antibody, dementia, stroke, heart disease and diabetes mellitus were the prognostic factors for mortality in bullous pemphigoid.

Relationship between the age and sex of the patient with the results of the indirect immunofluorescence test in patients with bullous dermatoses

Background/Aim: Autoimmune bullous diseases are characterised by the production of autoantibodies to epidermal or subepidermal adhesive proteins. The aim of this study was to determine the relationship between age and sex of patients with the results of indirect immunofluorescence test in patients with newly diagnosed bullous dermatoses. Methods: The investigation presents a retrospective study of newly diagnosed patients with autoimmune bullous diseases at the Clinic for Skin and Venereal Diseases of the University Clinical Centre in Banja Luka in the period 2016-2021. In addition to demographic data, the results of an indirect immunofluorescence test in two titres (≥ 1:10 and ≥ 1:100) were analysed. Results: In this study, almost the same number of patients with pemphigus (45.2 %) and pemphigoid (54.8 %) was found. There were more women than men in the total sample (p = 0.049). The average age of subjects with pemphigoid was higher than that of patients with pemphigus (p = 0.001). 48.2 % of patients with pemphigus and 51.8 % of patients with pemphigoid had a positive indirect immunofluorescence test. A positive test for epidermal intercellular substance in both sexes at a titre ≥ 1:100 is higher than a titre ≥ 1:10 (p = 0.029). Patients with autoantibody titres ≥ 1:100 to desmoglein-1 were statistically significantly older than patients with titres ≥ 1:10 (p = 0.047). Conclusion: Number of patients with pemphigus and pemphigoid were similar, with no difference in sex distribution between the two groups of patients, but patients with pemphigoid were older than patients with pemphigus. The difference between high and low autoantibody titres in both sexes was found only in the group of pemphigus on epidermal intercellular substance and desmoglein-1.

Clinical Manifestations of Bartonella henselae Infection Among Children

Objectives: The aim of this study was to analyze clinical manifestations, epidemiology and laboratory parameters of B. henselae infection among children treated at the University Hospital for Infectious Diseases “Dr. Fran Mihaljević”, Zagreb from January 2014 until June 2019. Materials and methods: We retrospectively analyzed the epidemiology, clinical and laboratory characteristics among children with positive indirect immunofluorescence assay for B. henselae IgM and IgG or positive B. henselae polymerase chain reaction from lymph node aspirate. Results: A total of 104 patients, 47 (45,1%) female and 57 (54,8%) male were enrolled. The median age was 9,7 (range, 1,1 to 17,3 years). A history of cat contact was present in 101 (97,1%) children. Acute infection was serologically confirmed in 87 (83,6%), in 5 (4,8%) with PCR while both methods were positive in 12 (11,5%) patients. The presentation on B. henselae infection were regional lymphadenopathy , disseminated disease, encephalopathy and fever of unknown origin. Suppurative inflammation was the most common complication in patients with lymphadenopathy 12/92 (13%). Full recovery was the most frequent outcome (96,1%). Conclusion lesion: B. henselae infection among children is usually a mild disease presented as regional lymphadenopathy. Serology and polymerase chain reaction are useful tests for diagnosis. Treatment duration and choice of therapy depend on clinical manifestation and developed complications.

P40: Positive indirect immunofluorescence pemphigoid antibodies: a clinicopathological analysis

P40: Positive indirect immunofluorescence pemphigoid antibodies: a clinicopathological analysis

Internal audit of two antineuronal immunoblots and the role of indirect immunofluorescence in sample screening

Anti-neuronal antibodies are associated with a range of clinical phenotypes that are often preceded by malignancy. Most of these antibodies can be identified by indirect immunofluorescence (IIF) using commercial primate neural tissue slides. The EUROImmun 12 and upgraded RAVO 14 antigen line immunoblots were directly compared to replace a 9 antigen RAVO immunoblot. Initial assessment was with samples with known positive specificities by IIF. The second phase involved parallel testing of routine pooled negative IIF samples as per institutional standard antineuronal testing algorithm. IIF results and clinical correlation were used to arbitrate on discordant immunoblot results. Correlation in known positives was 80%. The EUROImmun immunoblot did not identify disease causing SOX1 and Recoverin antibodies. Routine pooled analysis did not identify discrepancy between the platforms. Assessment of 318 patient samples did not reveal any cases where a diagnosis was correctly identified by line immunoblot and not IIF. A single sample with positive Yo antibodies was identified by both platforms without clinical or IIF correlation, making this a likely false positive. Subsequent to this audit, Westmead ICPMR has migrated to the 14 antigen RAVO line blot for confirmation of positive IIF or for patients with equivocal results only.

Clinical implications of ANCA positivity in a hospital setting: a tertiary center experience.

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 19years. 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 18years 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 < 0.001). The diagnosis of AAV in unlikely in subjects with discrepancies between IIF and ELISA, particularly with only positive IIF.

Recognising the spectrum of scleromyositis: HEp-2 ANA patterns allow identification of a novel clinical subset with anti-SMN autoantibodies

ObjectiveTo describe systemic sclerosis (SSc) with myopathy in patients without classic SSc-specific and SSc-overlap autoantibodies (aAbs), referred to as seronegative scleromyositis.MethodsTwenty patients with seronegative scleromyositis diagnosed by expert opinion were...

Pemphigus and Bullous Pemphigoid

Pemphigus disorders are characterized by acantholysis, whereas pemphigoid disorders are characterized by a dermal-epidermal split. Diagnosis of pemphigus or pemphigoid relies on a combination of positive anti-desmoglein or anti-collagen XVII serology, confirmatory direct immunofluorescence, and clinical features. Treatment for immunobullous disease revolves around various immunosuppressants, most often some combination of rituximab, prednisone, and IVIg. Paraneoplastic pemphigus is characterized by hemorrhagic crusting of the lips with positive indirect immunofluorescence on rat bladder epithelium, which should prompt a search for malignancy. Hailey-Hailey disease is a genetically mediated pemphigus that typically occurs in skin folds and responds to a number of agents including botulinum toxin, topical steroids, and other anecdotal therapies. This review contains 17 figures, 2 tables, and 109 references. Keywords: blister, pemphigus, bullous, rituximab, bullae, prednisone

Pemphigus and Bullous Pemphigoid

Pemphigus disorders are characterized by acantholysis, whereas pemphigoid disorders are characterized by a dermal-epidermal split. Diagnosis of pemphigus or pemphigoid relies on a combination of positive anti-desmoglein or anti-collagen XVII serology, confirmatory direct immunofluorescence, and clinical features. Treatment for immunobullous disease revolves around various immunosuppressants, most often some combination of rituximab, prednisone, and IVIg. Paraneoplastic pemphigus is characterized by hemorrhagic crusting of the lips with positive indirect immunofluorescence on rat bladder epithelium, which should prompt a search for malignancy. Hailey-Hailey disease is a genetically mediated pemphigus that typically occurs in skin folds and responds to a number of agents including botulinum toxin, topical steroids, and other anecdotal therapies. This review contains 17 figures, 2 tables, and 109 references. Keywords: blister, pemphigus, bullous, rituximab, bullae, prednisone

P181 Detection of anti-neutrophil cytoplasmic antibodies: a single-centre evaluation of the value of indirect immunofluorescence versus antigen-specific immunoassays

Abstract Background Anti-neutrophil cytoplasmic antibodies (ANCAs) are valuable laboratory markers used for the diagnosis of medium and small-vessel vasculitis, including granulomatosis with polyangiitis (GPA), eosinophilic granulomatosis with polyangiitis (EGPA) and microscopic polyangiitis (MPA). According to the 1999 international consensus, indirect immunofluorescence (IIF) should be used to screen for ANCAs; samples containing ANCAs should then be tested by immunoassays for proteinase 3 (PR3)-ANCAs and myeloperoxidase (MPO)-ANCAs. As dependable immunoassays for PR3ANCAs and MPO-ANCAs have become broadly available, there is increasing agreement that high-quality immunoassays are the preferred screening method for the diagnosis of ANCA-associated vasculitis (AAV). This single centre study was performed to evaluate the accuracy of MPO and PR3 immunoassays in comparison to IIF in the diagnosis of AAV. Methods We retrospectively reviewed the clinical information of patients that underwent ANCA testing between 1/6/19-30/6/19, via our electronic web portal. Data collected included patients’ demographics, ANCA /MPO/PR3 result, and diagnosis as established by their treating physician. A total of 403 ANCA requests were submitted to the Immunology Department at the Royal Wolverhampton NHS Trust. We excluded 55 patients with insufficient information. Due to multiple ANCA tests in a single patient we included only the first test request. Therefore, the study population includes 345 patients. IIF at our centre is performed for all ANCA requests using a commercially available ASP1200 automated dilutor to process the IIF slides with Werfen (Inova) ANCA slides. For the detection of antibodies against MPO and PR3 a commercially available immuno-assay Phadia-250 ImmunoCAP analyser is used in patients with positive IIF. Diagnostic performance was assessed using sensitivity and specificity. Results The proportion of ANCA negative patients by IIF (177 patients,51.3%) was almost identical to that of the ANCA positive group (168 patients,48.7%). Only 13 patients (3.8%) received a diagnosis of AAV. Of the ANCA positive patients 12 (7.1%) had an AAV. 6 of the 13 patients with vasculitis had a supportive tissue biopsy. One ANCA negative patient and one ANCA positive MPO/PR3 negative patient had a diagnosis of vasculitis, previously diagnosed and now on steroid treatment; both were ANCA and MPO positive at diagnosis. 15 patients were MPO or PR3 positive, 11/15 had a diagnosis of vasculitis (73%). Our data found IIF has a sensitivity of 91.67%, specificity 53.01% for detection of AAV. In comparison immunoassay was found to have a sensitivity of 91.67% and specificity of 97.44%. Conclusion ANCAs are helpful in the diagnosis of AAV, but their use as a diagnostic biomarker should be undertaken in the appropriate clinical context. Immunoassay has a superior diagnostic performance at our centre. Our data suggests screening for ANCA with IIF is not of added value when using high-quality antigen-specific immunoassays. Disclosures N. Cleaton None. T. Adizie None. N. Barkham None.

Comparison of C3d immunohistochemical staining to enzyme-linked immunosorbent assay and immunofluorescence for diagnosis of bullous pemphigoid

Bullous pemphigoid (BP), the most common autoimmune blistering disease, may be diagnostically challenging. Direct immunofluorescence (DIF), indirect immunofluorescence (IIF), enzyme-linked immunosorbent assay (ELISA), and recently, C3d immunohistochemistry (IHC), are used as adjuncts to diagnosis. To compare C3d IHC to DIF, IIF, and ELISA testing in BP diagnosis. C3d IHC was performed on skin biopsyspecimens from 51 patients (27 with BP and 24 with other blistering diseases) and compared to DIF and IIF, with anti-BP180 or anti-BP230 ELISA results used as the gold standard. We found C3d IHC, DIF, and IIF had similar sensitivity (74.1%, 63.1%, and 70.4%), specificity (95.8%, 100%, and 100%), positive predictive value (95.2%, 100%, and 100%), and negative predictive value (76.7%, 70.6%, and 75%) for BP. Cases with positive C3d IHC, DIF, and IIF had significantly higher anti-BP180 and anti-BP230 by ELISA than cases with negative testing (P<.0001). False-negative IIF results were associated with lower BP230 compared with true-positive results (P=.03). This was a single-center, retrospective study. Our study compared C3d IHC to DIF and IIF in BP diagnosis, demonstrating C3d IHC on fixed tissue provides similar diagnostic utility to immunofluorescence and ELISA.

Plucked Hair as a Substrate for Indirect Immunofluorescence in Cases of Pemphigus Vulgaris.

Introduction:Indirect immunofluorescence (IIF) is used to determine the circulating autoantibodies in cases of Pemphigus. For IIF in Pemphigus several substrates had been used. This study was done to determine whether plucked hair can be utilized as a substrate for IIF in cases of Pemphigus. Aim & Objectives: To determine the efficacy of utilizing the plucked hair as a substrate for indirect immunofluorescence among patients with Pemphigus vulgaris.Methodology:Thirty two consecutive patients with active and fresh diagnosed cases of pemphigus vulgaris (PV) who did not undergo any treatment and those patients who had positive DIF findings of characteristic fish net or chicken wire pattern of intercellular IgG deposits in the epidermis of perilesional skin were included in the study. A total of 32 control subjects without any auto immune disorders were selected and the blood samples were taken from these patients for IIF analysis. Anagen hair were collected from the healthy volunteers without pemphigus in the same way as for trichogram. Telogen hair were obtained by combing the hair and collecting the loose strands of hair on the comb and further confirmed under microscope. Three hairs each of anagen and telogen stage were collected from each subject to be utilized as a substrate for IIF.Results:Out of 32 samples 20 samples showed positive results for Ig G alone and 5 samples showed positivity for Ig G and C3. One sample showed positivity for C3alone. All these 26 samples were considered to have positive IIF test based on the intercellular pattern of deposit. Positive IIF results were observed in anagen hair samples and were negative in all the telogen hair samples. Six anagen hair samples did not show any positive findings in IIF for the study group. All the 32 control samples showed negative reports in IIF.Conclusion:In conclusion, IIF microscopy on plucked hair as a substrate is a more sensitive immunoassay for the detection of circulating intercellular autoantibodies in PV and the lower negative predictive value of this substrate is a limitation. Further large scale studies might provide better information regarding the practical utility.

Nonbullous pemphigoid: Insights in clinical and diagnostic findings, treatment responses, and prognosis

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. To describe the characteristics of nonbullous pemphigoid. 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. The review included 69 patients. The mean delay in diagnosis was 29months. 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 3years were 14%, 34%, and 46%, respectively, with an 8.6-fold increased all-cause mortality risk. The retrospective study design. 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.

Harmonizing by reducing inter-run variability: performance evaluation of a quality assurance program for antinuclear antibody detection by indirect immunofluorescence.

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.

Paraneoplastic pemphigus: Retrospective study of a case series

Paraneoplastic pemphigus (PNP) is a rare condition associated with poor prognosis. It associates polymorphic mucocutaneous manifestations with neoplasia. Diagnosis is difficult because of the various clinical and histological features involved and the lack of specificity of immunological examinations. We retrospectively analyzed the records of patients presenting with PNP in the Poitou-Charentes region between 2000 and 2015. Seven patients were included. They presented 9 neoplasias (1 lymphoma, 1 melanoma, and 7 carcinomas) diagnosed from 4 months before to 25 months after the occurrence of cutaneous (6/7) and/or mucosal (6/7) polymorphic lesions. Histological examination revealed epidermal acantholysis (7/7), keratinocytic necrosis (4/7), and interface lichenoid dermatitis (5/7). Intercellular deposits of IgG and C3 or along the dermo-epidermal junction were detected with direct immunofluorescence (IF) (7/7). Four of 6 patients tested had positive indirect IF on rat bladder epithelium. Follow-up ranged from 1-132 months with a one-year survival of 85.7%. The clinical and histopathological presentations observed in our patients were polymorphic, with overlap between the clinical and histological features of PNP and classical pemphigus. Prognosis and survival appear better in our series than in the literature. It is possible that in some cases, the association of pemphigus with neoplasia was fortuitous, which might account for the better prognosis. A new consensus on the diagnostic criteria for PNP is needed to help practitioners to consensually diagnose it for prognostic or therapeutic trials.