Ocular cicatricial pemphigoid (OCP) is a chronic bilateral, blinding, cicatrizing form of conjunctivitis with relapsing and remitting periods. It has strong evidence for an immune type II hypersensitivity that leads to subconjunctival fibrosis and extensive systemic bullae formation. To the best knowledge of the authors, this is the first reported case of direct immunofluorescence (DIF) assay-proven OCP in an elderly Filipino man.

A 68-year-old male presented with bilateral corneal conjunctivalization, symblepharon, ectropion, conjunctival hyperemia testing positive with conjunctival biopsy for basement membrane antibodies with DIF for the left eye, while turning out negative for the right eye. He was managed as a case of OCP, both eyes, and was given topical steroids and antibiotics. Oral Dapsone was started by Dermatology and Rheumatology Services.

OCP is a rare autoimmune and blinding disease. Early diagnosis and prompt treatment are vital as ocular complications permanently affect the quality of life of patients as seen in our patient. DIF assay remains the gold-standard for diagnosis. Systemic immunosuppression is the mainstay of treatment. Adjunctive supportive topical medication may be given to alleviate ocular discomfort. A multidisciplinary approach is essential to provide holistic care to each patient.

Objective To prepare monoclonal antibodies against bovine CD4 and identify their basic biological characteristics. Methods Recombinant bovine CD4 (rHis-BoCD4 and rGST-BoCD4) was successfully expressed and purified by constructing a prokaryotic plasmid of bovine CD4 gene. The bovine CD4 monoclonal antibody was produced using hybridoma technology. The subtype and potency of the monoclonal antibody were identified and analyzed by ELISA, while specificity was analyzed through indirect immunofluorescence assay (IFA) and Western-blot. Results Four hybridoma cell lines, namely, 1H4, 6A10, 3F9 and 4G10, stably secreting monoclonal antibodies against BoCD4 were successfully obtained. The subclasses of the monoclonal antibodies subclass 6A10 was IgG2b and the rest of the monoclonal antibodies were of IgM type. Western-blot results showed that the four anti-bovine CD4 mAb strains were able to specifically bind to the bovine CD4 protein expressed in vitro. Indirect immunofluorescence assay showed that four monoclonal antibodies were able to specifically recognize the natural bovine CD4 protein. Flow cytometry assay showed that 3F9 was best to recognize bovine natural CD4 molecules. Conclusion Four monoclonal antibody strains with high specificity to natural bovine CD4 protein were successfully prepared, which lays the foundation for the subsequent studies on the function of bovine CD4 and diagnosis and treatment of bovine T-lymphocyte diseases.
Animals ; Antibodies, Monoclonal/isolation & purification* ; Cattle ; CD4 Antigens/genetics* ; Hybridomas/immunology* ; Antibody Specificity/immunology* ; Mice ; Mice, Inbred BALB C ; Enzyme-Linked Immunosorbent Assay ; Fluorescent Antibody Technique, Indirect

Ocular cicatricial pemphigoid (OCP) is a chronic bilateral, blinding, cicatrizing form of conjunctivitis with relapsing and remitting periods. It has strong evidence for an immune type II hypersensitivity that leads to subconjunctival fibrosis and extensive systemic bullae formation. To the best knowledge of the authors, this is the first reported case of direct immunofluorescence (DIF) assay-proven OCP in an elderly Filipino man.

A 68-year-old male presented with bilateral corneal conjunctivalization, symblepharon, ectropion, conjunctival hyperemia testing positive with conjunctival biopsy for basement membrane antibodies with DIF for the left eye, while turning out negative for the right eye. He was managed as a case of OCP, both eyes, and was given topical steroids and antibiotics. Oral Dapsone was started by Dermatology and Rheumatology Services.

OCP is a rare autoimmune and blinding disease. Early diagnosis and prompt treatment are vital as ocular complications permanently affect the quality of life of patients as seen in our patient. DIF assay remains the gold-standard for diagnosis. Systemic immunosuppression is the mainstay of treatment. Adjunctive supportive topical medication may be given to alleviate ocular discomfort. A multidisciplinary approach is essential to provide holistic care to each patient.

Objective:This study aims to evaluate the clinical utility of total IgE （tIgE） and specific IgE （sIgE） levels in nasal secretions for diagnosing allergic rhinitis. The investigation is enhanced through an improved method of nasal secretion collection and advanced quantum dot immunofluorescence detection technology. Methods:A total of 88 subjects were enrolled in this study, and demographic data and clinical characteristics were collected through standardized questionnaires. Anterior rhinoscope was used to check the local condition of the nasal cavity. Each participant underwent skin prick test（SPT）. The total IgE（tIgE） and sIgE in nasal secretions were quantitatively analyzed by improved nasal secretion collection strategy and quantum dot immunofluorescence method, and the correlation between them and clinical symptoms and signs was discussed. The receiver operating characteristic curve（ROC） was used to calculate the optimum threshold and detection efficiency of total IgE and sIgE in nasal secretions. Results:The improved method successfully collected nasal secretions from all subjects. Based on SPT results, participants were categorized into three groups: normal control （20 cases）, non-allergic rhinitis （22 cases）, and allergic rhinitis （46 cases）. Analysis showed that both tIgE and sIgE levels in nasal secretions correlated with nasal symptoms and signs. A tIgE level of ≥9.42 IU/mL was identified as a cut-off for allergic rhinitis diagnosis, demonstrating an 85.37% agreement with SPT results. Furthermore, cut-off values for house dust mite sIgE （≥0.34 IU/mL） and dermatophagoides Farinae sIgE （≥0.41 IU/mL） yielded a diagnostic agreement of 97.56% with SPT. Notably, two patients in the non-allergic rhinitis group tested negative for SPT but positive for dust mite sIgE in nasal secretions and exhibited positive results in the nasal provocation test, indicating potential local allergic rhinitis. Conclusion:The assessment of tIgE and mite-specific IgE levels in nasal secretions presents a rapid, reliable, and non-invasive approach for diagnosing allergic rhinitis, particularly in cases of local allergic rhinitis.
Humans ; Immunoglobulin E/analysis* ; Quantum Dots ; Male ; Female ; Adult ; Young Adult ; Middle Aged ; Rhinitis, Allergic/immunology* ; Adolescent ; Fluorescent Antibody Technique/methods* ; Case-Control Studies ; Nasal Mucosa/immunology*

Abnormal accumulation of collagen fibrils is a hallmark feature of oral submucous fibrosis (OSF). However, the precise characteristics and underlying mechanisms remain unclear, impeding the advancement of potential therapeutic approaches. Here, we observed that collagen I, the main component of the extracellular matrix, first accumulated in the lamina propria and subsequently in the submucosa of OSF specimens as the disease progressed. Using RNA-seq and Immunofluorescence in OSF specimens, we screened the cartilage oligomeric matrix protein (COMP) responsible for the abnormal collagen accumulation. Genetic COMP deficiency reduced arecoline-stimulated collagen I deposition significantly in vivo. In comparison, both COMP and collagen I were upregulated under arecoline stimulation in wild-type mice. Human oral buccal mucosal fibroblasts (hBMFs) also exhibited increased secretion of COMP and collagen I after stimulation in vitro. COMP knockdown in hBMFs downregulates arecoline-stimulated collagen I secretion. We further demonstrated that hBMFs present heterogeneous responses to arecoline stimulation, of which COMP-positive fibroblasts secrete more collagen I. Since COMP is a molecular bridge with Fibril-associated collagens with Interrupted Triple helices (FACIT) in the collagen network, we further screened and identified collagen XIV, a FACIT member, co-localizing with both COMP and collagen I. Collagen XIV expression increased under arecoline stimulation in wild-type mice, whereas it was hardly expressed in the Comp^-/- mice, even with under stimulation. In summary, we found that COMP may mediates abnormal collagen I deposition by functions with collagen XIV during the progression of OSF, suggesting its potential to be targeted in treating OSF.
Oral Submucous Fibrosis/pathology* ; Cartilage Oligomeric Matrix Protein/genetics* ; Animals ; Mice ; Humans ; Fibroblasts/metabolism* ; Collagen Type I/metabolism* ; Arecoline/pharmacology* ; Mouth Mucosa/metabolism* ; Cells, Cultured ; Fluorescent Antibody Technique

Pemphigus foliaceous is a rare autoimmune blistering disease, while psoriasis is a common immune‑mediated inflammatory skin disease. The coexistence of psoriasis and pemphigus foliaceous has rarely been reported. We report a case of a 55‑year‑old Filipino female with an 8‑year history of chronic plaque‑type psoriasis biopsy‑proven. After 5 years, she developed generalized flaccid bullae and crusted erosions over the face, trunk, and extremities, with no mucous membrane involvement. Skin punch biopsy, direct immunofluorescence, and enzyme‑linked immunosorbent assay were consistent with pemphigus foliaceous. The combination of topical corticosteroids and oral methotrexate was selected as the therapeutic approach, leading to a notable improvement in the patient’s condition. This case report underscores the significance of identifying the simultaneous presence of psoriasis alongside autoimmune blistering diseases like pemphigus foliaceous. Examining predisposing and triggering factors, performing re‑biopsy, and further work‑up as the disease evolves may yield more profound insights. Nonetheless, effectively managing this condition poses a significant challenge.
Fluorescent Antibody Technique, Direct ; Methotrexate ; Psoriasis

Molting is an important physiological phenomenon of many metamorphosis insects, during which the old and new epidermis are separated by enzymes present in the molting fluid. Various proteomic studies have discovered the presence of Bombyx mori carboxypeptidase A (Bm-CPA) in the molting fluid of silkworm, but its function remains unclear. In order to better understand the role of Bm-CPA in the molting process of silkworm, Bm-CPA was analyzed by bioinformatics analysis, real-time fluorescence quantitative PCR, antibody preparation, immunofluorescence staining, and expression in Pichia pastoris. The results showed that Bm-CPA had a conserved M14 zinc carboxypeptidase domain and glycosylation site. Its expression was regulated by ecdysone 20E, and large expression was observed in the epidermis of the upper cluster stage. Immunofluorescence staining showed that Bm-CPA was enriched in the epidermis during the molting stage, and the inhibitor of Bm-CPA led to the larval death due to the inability to molt. We also successfully obtained a large number of recombinant Bm-CPA proteins by Pichia pastoris expression in vitro. These results may facilitate further understanding the molting development process of silkworm.
Animals ; Molting/genetics* ; Bombyx/genetics* ; Carboxypeptidases A/metabolism* ; Proteomics ; Larva/metabolism* ; Fluorescent Antibody Technique ; Insect Proteins/metabolism*

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.
Humans ; Antibodies, Antineutrophil Cytoplasmic/analysis* ; Myeloblastin ; Cross-Sectional Studies ; Sensitivity and Specificity ; Fluorescent Antibody Technique, Indirect ; Enzyme-Linked Immunosorbent Assay/methods*

Different fragments of SARS-CoV-2 nucleocapsid (N) protein were expressed and purified, and a fluorescence immunochromatography method for detection of SARS-CoV-2 total antibody was established. The effect of different protein fragments on the performance of the method was evaluated. The N protein sequence was analyzed by bioinformatics technology, expressed in prokaryotic cell and purified by metal ion affinity chromatography column. Different N protein fragments were prepared for comparison. EDC reaction was used to label fluorescence microsphere on the synthesized antigen to construct sandwich fluorescence chromatography antibody detection assay, and the performance was systemically evaluated. Among the 4 prepared N protein fragments, the full-length N protein (N419) was selected as the optimized coating antigen, N412 with 0.5 mol/L NaCl was used as the optimal combination; deleting 91-120 amino acids from the N-terminal of N412 reduced non-specific signal by 87.5%. the linear range of detection was 0.312-80 U/L, the limit of detection was 0.165 U/L, and the accuracy was more than 95%. A fluorescence immunochromatographic detection method for analysis of SARS-CoV-2 total antibody was established by pairing N protein fragments. The detection result achieved 98% concordance with the commercially available Guangzhou Wanfu test strip, which is expected to be used as a supplementary approach for detection of SARS-CoV-2. The assay could also provide experimental reference for improving the performance of COVID-19 antibody detection reagents.
Antibodies, Viral ; COVID-19 ; Chromatography, Affinity ; Fluorescent Antibody Technique ; Humans ; Microspheres ; SARS-CoV-2 ; Sensitivity and Specificity

Hailey-Hailey disease (HHD) is an uncommon acantholytic disorder of the skin. This is a case of a 64-year-old Filipino female with a chronic history of painful and malodorous intertriginous plaques. Histopathologic evaluation showed overlapping features of pemphigus vulgaris and Hailey-Hailey disease. A negative direct immunofluorescence test clinched the diagnosis of Hailey-Hailey disease. The patient was advised regarding preventive measures and treated with topical antibiotics and corticosteroids with improvement of her lesions.
Pemphigus, Benign Familial ; Fluorescent Antibody Technique, Direct