No abstract available.
Chlamydia* ; Fluorescent Antibody Technique, Indirect*

No abstract available.
Diagnosis* ; Fluorescent Antibody Technique, Indirect* ; Herpes Simplex* ; Simplexvirus*

The pathophysiology of glandular dysfunction in Sjögren's syndrome (SS) has not been fully elucidated. Previously, we reported the presence of autoantibodies to AQP-5 in patients with SS, which was associated with a low resting salivary flow. The purpose of this study was to investigate the presence of anti-AQP1 autoantibodies. To detect anti-AQP1 autoantibodies, cell-based indirect immunofluorescence assay was developed using MDCK cells that overexpressed human AQP1. By screening 112 SS and 52 control sera, anti-AQP1 autoantibodies were detected in 27.7% of the SS but in none of the control sera. Interestingly, the sera that were positive for anti-AQP1 autoantibodies also contained anti-AQP5 autoantibodies in the previous study. Different from anti-AQP5 autoantibodies, the presence of anti-AQP1 autoantibodies was not associated with the salivary flow rate. Although anti-AQP1 autoantibodies are not useful as a diagnostic marker, the presence of autoantibodies to AQP1 may be an obstacle to AQP1 gene therapy for SS.
Aquaporin 1 ; Autoantibodies* ; Fluorescent Antibody Technique ; Fluorescent Antibody Technique, Indirect ; Genetic Therapy ; Humans ; Madin Darby Canine Kidney Cells ; Mass Screening

BACKGROUND: Neutrophils fixed on a slide are used in indirect immunofluorescence assay (IIF) for anti-neutrophil cytoplasmic antibody (ANCA). A simple method to prepare ANCA slides is described in this report. METHODS: This method uses an overlay method to separate WBC from EDTA whole blood and Octospottrade mark kits to cytocentrifuge WBC onto 8-well slides. This in-house ANCA IIF was compared with commercial ANCA IIF using 40 sera, and with enzyme linked immunosorbent assay for anti-myeloperoxidase (MPO) antibody (MPO ELISA) using 117 sera. RESULTS: In ANCA IIF parallel tests using 10 positive and 30 negative sera, in-house slides were in good qualitative agreement, 97.5% (39/40) with commercial kits (kappa=0.93). Fluorescence pattern and intensity of positive reaction was also highly concordant between two methods. In MPO ELISA positive cases, the positivity rate of in-house ANCA IIF was 97.8% (45/46), which accounts for its assay sensitivity for the presence of anti-MPO antibody. CONCLUSIONS: In-house ANCA slides was convenient and cost-efficient to be prepared. As these slides were found to have acceptable agreement and sensitivity as compared with commercial kits, in-house ANCA IIF is considered to be useful to screen ANCA in combination with MPO ELISA in routine clinical laboratory practice.
Antibodies, Antineutrophil Cytoplasmic ; Edetic Acid ; Enzyme-Linked Immunosorbent Assay ; Fluorescence ; Fluorescent Antibody Technique, Indirect ; Neutrophils

Group A rotaviruses are the most common causes of gastroenteritis among infants and young children. The outer capsid layer of the virus is composed of two structural proteins, VP4 and VP7, and they play important roles in protection by eliciting neutralization antibodies. Group A rotaviruses are subdivided into distinct G and P serotypes according to the antigenic differences of the VP7 and VP4, respectively. Rotavirus G9 serotype was thought to be the fifth most common serotype circulating among the population worldwide. In this study, G9 human rotaviruses (HRV) were isolated from fecal samples using MA104 cells and characterized. Characteristic cytopathic effects of rotavirus were observed and rotaviral antigens were confirmed by indirect immunofluorescence antibody test in MA104 cells inoculated with isolated HRV strains. The nucleotide sequences of the VP7 gene of Korean G9 HRV isolated in this study were determined and compared with those of other recent and prototype G9 rotavirus strains from other parts of the world. Also, the nucleotide sequences of VP4 and NSP4 gene of Korean G9 HRV were determined and compared with those of other rotavirus strains from other countries. The results showed that the Korean HRV isolates belong to a G9, P[8] and NSP4 B genotype. The Korean G9 HRV isolates and their nucleotide sequence data would be usefully applied for the vaccine development of HRV in the near future.
Antibodies ; Base Sequence ; Capsid ; Child ; Fluorescent Antibody Technique, Indirect ; Gastroenteritis ; Genotype ; Humans* ; Infant ; Rotavirus*

BACKGROUND: Immunolcgical assays are required for the accurate diagnosis of autoimmune bullous dermatoses including pemphigus vulgaris(PV) and pemphigus foliaceus(PF). In the detection of circulating autoantibodies to pemphigus antigens(desmosomal components), the priority remains controversial between indirect immunofluorescence(IF) and immunoblot(IB) assay. OBJECTIVE: In the present study we compared the sensitivity of indirect IF and that of IB using amplified alkaline phosphatase staining system in the detection of pemphigus autoantibodies. PATIENTS: We selected eight patients with serum endpoint titer of 1:80 in preliminary study. Among these patients three were PV and five were PF. METHODS/RESULTS: The titers of IgG autoantibodies found on indirect IF were confirmed as 1: 80 in all patients, whereas the titers examined by IB assay were much higher, 1: 640 to 1: 2560. In the 3 sera of PV patients, the titers of two cases were 1: 1280 and the third case was 1: 2560. In 5 cases of PF, one was 1:640, two were 1: 1280, and two were 1:2560. CONCLUSION: This result suggests that the immunoblot examination using amplified alkaline phosphatase staining system demonstrates higher sensitivity compared with indirect IF(p=0.0003 by Mann-Whitney U test) in the detection of pemphigus autoantibodies.
Alkaline Phosphatase ; Autoantibodies* ; Diagnosis ; Fluorescent Antibody Technique, Indirect* ; Humans ; Immunoglobulin G ; Pemphigus* ; Skin Diseases, Vesiculobullous

BACKGROUND: Immunolcgical assays are required for the accurate diagnosis of autoimmune bullous dermatoses including pemphigus vulgaris(PV) and pemphigus foliaceus(PF). In the detection of circulating autoantibodies to pemphigus antigens(desmosomal components), the priority remains controversial between indirect immunofluorescence(IF) and immunoblot(IB) assay. OBJECTIVE: In the present study we compared the sensitivity of indirect IF and that of IB using amplified alkaline phosphatase staining system in the detection of pemphigus autoantibodies. PATIENTS: We selected eight patients with serum endpoint titer of 1:80 in preliminary study. Among these patients three were PV and five were PF. METHODS/RESULTS: The titers of IgG autoantibodies found on indirect IF were confirmed as 1: 80 in all patients, whereas the titers examined by IB assay were much higher, 1: 640 to 1: 2560. In the 3 sera of PV patients, the titers of two cases were 1: 1280 and the third case was 1: 2560. In 5 cases of PF, one was 1:640, two were 1: 1280, and two were 1:2560. CONCLUSION: This result suggests that the immunoblot examination using amplified alkaline phosphatase staining system demonstrates higher sensitivity compared with indirect IF(p=0.0003 by Mann-Whitney U test) in the detection of pemphigus autoantibodies.
Alkaline Phosphatase ; Autoantibodies* ; Diagnosis ; Fluorescent Antibody Technique, Indirect* ; Humans ; Immunoglobulin G ; Pemphigus* ; Skin Diseases, Vesiculobullous

OBJECTIVETo compare indirect immunofluorescence assay (IIFA) and enzyme-linked immunosorbent assay (ELISA) for detecting antinuclear antibodies (ANA) and anti-double-stranded DNA antibodies (anti-dsDNA).

METHODSA total of 125 serum samples were obtained from patients with established or suspected autoimmune disease, and 82 samples were used for ANA detection and 57 for anti-dsDNA detection using both IIFA and ELISA. Fourteen samples were examined for both ANA and anti-dsDNA. In cases where discrepancy occurred in the results by the two methods, extractable nuclear antigens were detected using immunoblotting.

RESULTSThe positivity rate of ANA detected by IIFA and ELISA was significantly different (87.8% and 73.17%, respectively, P<0.01), but the positivity rate of anti-dsDNA was similar between IIFA and ELISA (77.19% and 71.93%, respectively, P>0.05). The percent agreement between the two testing methods with different cutoff values of ANA and anti-dsDNA showed significant differences (P<0.01), and for some uncommon patterns, the percent agreement of the two methods was lowered in ANA detection but remained unchanged for anti-dsDNA with different ANA patterns. High percent agreements of the two methods were obtained with the cutoff ANA titer of 1:100 and the cutoff anti-dsDNA value of weak positivity, but they demonstrated a significant difference in testing low-titer ANA and anti-dsDNA.

CONCLUSIONIIFA is more sensitive than ELISA in detecting the total ANA and anti-dsDNA. ELISA prescreening combined with IIFA can obtain the information of the nuclear pattern and allow the observation of the titer alterations. The combination of two or more testing methods can greatly enhance the accuracy of the results.

Type VII collagen is one of the major structural components of the corneal epithelial adhesion complex. Using the immunogold technique combined with indirect immunofluorescence analysis, the fine structural distribution of type VII collagen was studied in the corneas obtained from 5 enucleated hyman eyes (age range, 1-77 years) including one pathologic cornea from graft rejection. The findings on normal cornea corroborated the results from previous studies. In pathologic cornea from graft rejection, type VII collagen antibodies generated linear and irregular patchy fluorescence staining along the epithelial-stromal interface and immunogold binding to type VII collagen mainly occurred within the undulating lamina densa, more densealy distributed anchoring plaques and anchoring fibrils. The distribution of type VII collagen in pathologic human cornea from graft rejection is similar to normal human cornea. But, in pathologic cornea, type VII collagen is more densely distributed in superficial stroma and forms more extended anchoring network, which may be derived from the increased secretion of the type VII collagen due to the activated basal epithelial cell during healing process.
Antibodies ; Collagen Type VII* ; Cornea* ; Epithelial Cells ; Fluorescence ; Fluorescent Antibody Technique, Indirect ; Graft Rejection ; Humans ; Immunohistochemistry

Granulocyte-specific antigens, such as NA1, NA2, NBI, NB2, NC1, NDI, NE1, are a group of antigens specifically expresesed only on the granulocytes. Antibodies against these are involved in some clinical disorders such as alloimmune neonatal neutropenia(ANN), autoimmune neutropenia(AIN), and transfusion-related acute lung injury(TRALI). We investigated the frequencies of NA1, NA2, NB1, and Mart antigens among Koreans by the granulocyte indirect immunofluorescence test employing flow cytometry. The subjects were 105 Koreans(male 65, female 40), whose mean age was 31.7+/-8.2 years (range 16~57). The antigen and gene frequencies were as follows, NA1, 0.78, 0.53, NA2, 0.75, 0.50, NB1 0.86, 0.62, and Mart, 1.00, 1.00, respectively. The proportions of NB 1 -positive granulocytes among NB 1-positive individuals were variable(range, 27~100%). Through this study, the authors procured granunlocyte-specifiic antigen papnel, which is essential in the identification of causative antibody(-ies) in immune neutropenias.
Antibodies ; Female ; Flow Cytometry ; Fluorescent Antibody Technique, Indirect ; Gene Frequency ; Granulocytes ; Humans ; Lung ; Neutropenia