The porcine epidemic diarrhea virus (PEDV) infection inflicted substantial economic losses upon the global pig-breeding industry. This pathogen can infect all pigs and poses a particularly high fatality risk for suckling piglets. The S1 subunit of spike protein is a crucial target protein for inducing the particularly neutralizing antibodies that can intercept the virus-host interaction and neutralize virus infectivity. In the present study, the HEK293F eukaryotic expression system was successfully utilized to express and produce recombinant S1 protein. Through quantitative analysis, five monoclonal antibodies (mAbs) specifically targeting the recombinant S1 protein of PEDV were developed and subsequently evaluated using enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence assay (IFA), and flow cytometry assay (FCA). The results indicate that all five mAbs belong to the IgG1 isotype, and their half-maximal effective concentration (EC50) values measured at 84.77, 7.42, 0.89, 14.64, and 7.86 pM. All these five mAbs can be utilized in ELISA, FCA, and IFA for the detection of PEDV infection. MAb 5-F9 exhibits the highest sensitivity to detect as low as 0.3125 ng/mL of recombinant PEDV-S1 protein in ELISA, while only 0.096 ng/mL of mAb 5-F9 is required to detect PEDV in FCA. The results from antigen epitope analysis indicated that mAb 8-G2 is the sole antibody capable of recognizing linear epitopes. In conclusion, this study has yielded a highly immunogenic S1 protein and five high-affinity mAbs specifically targeting the S1 protein. These findings have significant implications for early detection of PEDV infection and provide a solid foundation for further investigation into studying virus-host interactions.

UNASSIGNED: To determine the clinical significance of Ro52 protein/tripartite motif-containing 21 antibody and specific antinuclear antibody patterns using indirect immunofluorescence technique.
METHODS: The retrospective study was conducted at the clinical laboratory of the First Affiliated Hospital of Chongqing Medical University, China, and comprised data from January 2017 to December 2021 of patients who underwent antinuclear antibody and anti-extractable nuclear antigen antibody detection. Inpatients with Ro52 antibody-positive status were taken as the cases, while anti-Ro52 negative patients with clear clinical diagnosis were taken as the controls. Data was analysed using SPSS 19.
RESULTS: There were 1802 cases and 1211 controls. Positive Ro52 showed significantly greater frequency in patients with primary Sjogren\'s syndrome, systemic lupus erythematosus, inflammatory myositis, dry eyes and interstitial lung disease (p<0.05). Ro52 antibody showed high positive predictive value for primary Sjogren\'s syndrome 25(96.15%), systemic lupus erythematosus 259(91.20%), connective tissue disease-associated interstitial lung disease 45(86.67%) and inflammatory myositis 60(86.67%). Antinuclear antibody indirect immunofluorescence patterns most frequently detected were nuclear speckled 128(40.89%) and cytoplasmic speckled 126(40.26%) (p<0.05). Interstitial lung disease was associated with the presence of cytoplasmic speckled antinuclear antibody indirect immunofluorescence pattern 24(19.2%), while tumours 47(36.5%) and hepatitis B 26(20.3%) seemed to be more frequent with nuclear speckled pattern (p<0.05). The simultaneous reactivity extractable nuclear antigen antibodies most frequently detected were antinuclear antibody+Ro52+anti-Sjogren\'s syndrome A+ 558(33.96%).
CONCLUSIONS: Ro52 antibody positivity was found to be associated with Sjogren\'s syndrome, systemic lupus erythematosus, inflammatory myositis, dry eye and interstitial lung disease. The antinuclear antibody immunofluorescence pattern of Ro52 positive was single and primarily granular cytoplasm type. Antinuclear antibody negative and Ro52 positive in the serum of patients also had certain significance in auxiliary disease diagnosis.

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

OBJECTIVE: This study aims to estimate the prevalence of anti-mitochondrial antibody subtype M2 (AMA-M2) and assess its consistency with AMA in a general population.
METHODS: A total of 8954 volunteers were included to screen AMA-M2 using enzyme-linked immunosorbent assay. Sera with AMA-M2 >50 RU/mL were further tested for AMA using an indirect immunofluorescence assay.
RESULTS: The population frequency of AMA-M2 positivity was 9.67%, of which 48.04% were males and 51.96% were females. The AMA-M2 positivity in males had a peak and valley value of 7.81% and 16.88% in those aged 40 to 49 and ≥70 years, respectively, whereas it showed a balanced age distribution in females. Transferrin and immunoglobulin M were the risk factors for AMA-M2 positivity and exercise was the only protective factor. Of 155 cases with AMA-M2 >50 RU/mL, 25 cases were AMA-positive, with a female-to-male ratio of 5.25:1. Only 2 people, with very high AMA-M2 of 760 and >800 RU/mL, met the diagnostic criteria of primary biliary cholangitis (PBC), making the prevalence of PBC 223.36 per million in southern China.
CONCLUSIONS: We found that AMA-M2 has a low coincidence rate with AMA in the general population. A new decision-making point for AMA-M2 is needed to improve consistency with AMA and diagnostic accuracy.

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

To explore the clinical diagnostic efficacy of antineutrophil cytoplasmic antibody associated vasculitis (AAV) by comparing the consistency and coincidence rate of serum anti-myeloperoxidase (MPO) antibody and anti-protease 3 (PR3) antibody detected by digital liquid chip method (DLCM) and enzyme-linked immunosorbent assay (ELISA). To provide reference for the selection of detection methods of anti-MPO antibody and anti-PR3 antibody in clinical laboratory. This study is a cross-sectional study, a total of 307 cases of antineutrophil cytoplasmic antibodies were detected in the Department of Clinical Immunology, West China Hospital of Sichuan University from January to March 2021. The serum samples and related clinical information were collected. At the same time, the levels of anti-MPO antibody and anti-PR3 antibody in serum samples were detected by ELISA and DLCM, indirect immunofluorescence (IIF) was used to re-test the differential samples between the two methods. SPSS 26.0 was used to analyze the test results, Cohen\'s kappa coefficient analysis was used to compare the consistency of the two methods, and paired chi-square test was used to compare the sensitivity and specificity of the two methods to AAV. The results showed that the positive cases of anti-MPO antibody detected by ELISA and DLCM were 63 and 44, and the negative cases were 244 and 263; the positive cases of anti-PR3 antibody detected by ELISA and DLCM were 34 and 28, and the negative cases were 273 and 279. The results of anti-MPO antibody and anti-PR3 antibody detected by the two methods had good consistency and coincidence rate, in which the total coincidence rate of anti-MPO antibody was 92.51%, the positive coincidence rate was 66.67%, and the negative coincidence rate was 99.18%. The results of consistency analysis showed that kappa=0.741 had well consistency. The total coincidence rate of anti-PR3 antibody is 96.74%, the positive coincidence rate is 76.47%, and the negative coincidence rate is 99.27%. The consistency analysis results show that kappa=0.821 had strong consistency. The results of IIF re-test of differential samples showed that the coincidence rate between DLCM and IIF was higher. The results of comparative analysis of anti-MPO antibody and anti-PR3 antibody showed that the specificity of DLCM was better than that of ELISA, and its sensitivity was lower than that of ELISA. In conclusion, the results of anti-MPO antibody and anti-PR3 antibody detected by DLCM were consistent with those of ELISA. In the combined detection of anti-MPO antibody and anti-PR3 antibody, the specificity of DLCM is better than that of ELISA.
通过比较数码液相芯片法（digital liquid chip method，DLCM）检测血清抗髓过氧化物酶（myeloperoxidase，MPO）抗体和抗蛋白酶3（proteinase 3，PR3）抗体与酶联免疫吸附法（enzyme-linked immunosorbent assay，ELISA）检测结果的一致性与符合率，探讨其对抗中性粒细胞胞浆抗体相关血管炎（antineutrophil cytoplasmic antibodies associated vasculitis，AAV）的临床诊断效能，为临床实验室抗MPO抗体和抗PR3抗体检测方法的选择提供参考。本研究为横断面研究，选取2021年1—3月在四川大学华西医院实验医学科临床免疫室检测抗中性粒细胞胞浆抗体的样本307例，收集其血清样本及相关临床信息，同时采用ELISA和DLCM平行检测血清样本中抗MPO抗体和抗PR3抗体水平，使用间接免疫荧光法（indirect immunofluorescent，IIF）就两种方法学差异样本进行复测验证。采用SPSS 26.0对检测结果进行统计分析，运用Cohen′s kappa系数分析比较两种方法检测结果的一致性，运用配对χ²检验比较两种方法检测结果对AAV的敏感度和特异度。结果显示，采用ELISA和DLCM两种方法平行检测抗MPO抗体阳性例数分别为ELISA 63例和DLCM 44例，阴性例数分别为ELISA 244例和DLCM 263例；检测抗PR3抗体阳性例数分别为ELISA 34例和DLCM 28例，阴性例数分别为ELISA 273例和DLCM 279例。两种方法检测抗MPO抗体和抗PR3抗体的检测结果均有良好的一致性与符合率，其中抗MPO抗体总符合率为92.51%，阳性符合率为66.67%，阴性符合率为99.18%，一致性分析结果显示 kappa=0.741，具有较强一致性；抗PR3抗体总符合率为96.74%，阳性符合率为76.47%，阴性符合率为99.27%，一致性分析结果显示 kappa=0.821，具有很强一致性。IIF复测差异样本结果均显示DLCM结果与IIF结果符合率更高。抗MPO抗体和抗PR3抗体联合检测性能比较分析结果显示，DLCM的特异度优于ELISA，其敏感度较ELISA偏低。综上，DLCM检测抗MPO抗体和抗PR3抗体检测结果与ELISA结果比较均具有良好的一致性与符合率。在联合检测抗MPO抗体和抗PR3抗体时，DLCM特异度优于ELISA。.

To investigate the distribution and clinical significance of the rods and rings (RR) pattern in various diseases.
A total of 169,891 patients in Peking Union Medical College Hospital (PUMCH) and 29,458 patients in Inner Mongolia People\'s Hospital (IMPH) from January 2018 to December 2020 were included, and the results of ANA (antinuclear antibodies) and special antibodies were analyzed retrospectively.
The positive rates of ANA and RR patterns were 34.84%, 0.16% in PUMCH, and 44.73%, 0.23% in IMPH. Anti-RR antibodies mainly appear in adults (≥ 41 years), mostly of low or medium fluorescence titers. Isolated RR patterns were mostly presented (60.30% and 69.12%, respectively), and the RR pattern mixed with the speckled pattern was most commonly observed among patients having two or more patterns. The RR pattern existed in a variety of diseases including hepatitis C, AIDs, pulmonary diseases, nephropathy diseases, and even healthy people. The highest prevalence of the RR pattern was observed in hepatic diseases, such as hepatic dysfunction (0.79%), hepatic cirrhosis (1.05%), PBC (0.85%), and AIH (0.65%), etc. The positive rate of specific antibodies in RR pattern cases was 31.25%, and anti-Ro52 (27, 20.61%) was the most common target antibody.
The RR pattern had a low prevalence in ANAs test samples and varied in different nationalities and regions. Except for hepatitis C, it could be observed in AIDs, pulmonary diseases, nephropathy, other hepatic diseases, and even healthy people, but the positive rate was slightly higher in hepatic diseases. Its mechanism of action and clinical relevance still need clarification.

BACKGROUND: Classical swine fever (CSF) is a severe infectious disease of pigs that causes significant economic losses to the swine industry.
OBJECTIVE: This study developed a solid-phase blocking enzyme-linked immunosorbent assay (spbELISA) method for the specific detection of antibodies against the CSF virus (CSFV) in porcine serum samples.
METHODS: A spbELISA method was developed based on the recombinant E2 expressed in Escherichia coli. The specificity of this established spbELISA method was evaluated using reference serum samples positive for antibodies against other common infectious diseases. The stability and sensitivity were evaluated using an accelerated thermostability test.
RESULTS: The spbELISA successfully detected the antibody levels in swine vaccinated with the C-strain of CSFV. In addition, the detection ability of spbELISA for CSFV antibodies was compared with that of other commercial ELISA kits and validated using an indirect immunofluorescence assay. The results suggested that the spbELISA provides an alternative, stable, and rapid serological detection method suitable for the large-scale screening of CSFV serum antibodies.
CONCLUSIONS: The spbELISA has practical applications in assessing the vaccination status of large pig herds.

To investigate the distribution and clinical significance of nuclear dense fine speckled (DFS) pattern in various diseases. A total of 95 289 patients who received DFS tests at Peking Union Medical College Hospital from January 2019 to December 2020 were included in this study. The results of indirect immunofluorescence assay (IIF) for detection of antinuclear antibody (ANA) were evaluated. The positive rates of ANA and DFS were 39.60% (37 733/95 289) and 1.19% (1 139/95 289) respectively. The positive rate of DFS in ANA-positive patients was 3.02% (1 139/37 733). DFS and ANA positivity were significantly different among different age groups rather than gender. The positivity rate of DFS reached the peak (55.57%, 633/1 139) in young patients between 21-40 years, while positive ANA with negative DFS was mainly observed in patients between 41-60 years (37.26%, 13 636/36 594). Additionally, single ANA-positivity were mainly detected in rheumatology department (59.23%, 18 402/31 066), whereas positive DFS was more common in obstetrics and gynecology department (3.08%, 49/1 593). There were 82.88% (944/1 139) patients with positive DFS diagnosed with non-autoimmune disease (non-AID), and 19.49%(222/1 139) with dermatosis. Positive DFS with higher titer (≥1∶320) was detected more frequently in autoimmune disease (AID) patients (5.13%, 10/195) than in non-AID patients (1.69%, 16/944) (P<0.05). The DFS pattern is rare in ANA positive patients, which is mainly observed in women between 21-49 years. High titer of DFS is prevalent in AID patients, but positive DFS is detected more in non-AID patients, especially those with dermatosis.
探讨抗核抗体（ANA）致密斑点型（DFS）在不同疾病中的分布及临床意义。回顾分析2019年1月至2020年12月北京协和医院95 289例连续送检标本间接免疫荧光法（IIF）检测ANA的结果。结果显示，ANA阳性率39.60%（37 733/95 289），DFS阳性率1.19%（1 139/95 289）；在ANA阳性患者中，DFS阳性率3.02%（1 139/37 733）。DFS阳性者在21~40岁比例为55.57%（633/1 139），而ANA阳性（除外DFS阳性）者在41~60岁比例为37.26%（13 636/36 594）。ANA阳性（除外DFS阳性）者主要在风湿免疫科（59.23%，18 402/31 066），而DFS阳性者主要在妇产科（3.08%，49/1 593）。1 139例DFS阳性者中，自身免疫病（AID）者195例（17.12%，195/1 139）；非AID者944例（82.88%，944/1 139），其中以皮肤病占比最高（19.49%，222/1 139）。AID患者中出现高滴度（≥1∶320）DFS的比例（5.13%，10/195）高于非AID患者（1.69%，16/944；P<0.05）。DFS是ANA阳性者中少见的荧光核型，主要分布在21~49岁女性患者；DFS阳性者主要出现在非AID患者中，以皮肤病最常见；DFS阳性滴度≥1∶320时常见于AID患者。.

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