• Title/Summary/Keyword: Fluorescent antibody technique

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Comparative Diagnostic Studies on Serologic and Molecular Biological Tests Against Haemorrhagic Fever with Renal Syndrome (신증후출혈열 환자의 혈청학적 및 분자생물학적 진단 검사법 비교)

  • 우영대;문희주;배형준
    • Biomedical Science Letters
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    • v.6 no.2
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    • pp.141-149
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    • 2000
  • The etiologic agents of haemorrhagic fever with renal syndrome (HFRS) in Korea are Hantaan and Seoul virus in the genus Hantavirus, family Bunyaviridae. Antibody titers of sera from HFRS patients against Hantaan virus were measured by immunofluorescent antibody technique (IFAT), enzyme-linked immunosorbent assay (ELISA), high density composite particle agglutination (HDPA) and plaque reduction neutralization test (PRNI). PRNT and nested reverse transcriptase polymerase chain reaction (nested RT-PCR) was used for serotypic differentiation of Hantaviruses against Hantaan and Seoul virus. Eight doubtful HFRS patients showed higher fluorescent, IgG ELISA, agglutination and neutralizing antibody titer by IFAT, ELISA IgG, HDPA and PRNT, respectively Five out of them showed high IgM antibody titer by IgM capture ELISA against Hantaan virus, remarkably. Fifteen HFRS patients showed higher fluorescent antibody titer by IFAT. In PRNT, 12 out of them showed high neutralizing antibody titer against HTNV, 2 against SEOV and 1 against both viruses. In nested RT-PCR using serotype specific-primer, 3 out of them showed positive against HTNV and 1 against SEOV.

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Detection of IgG and IsM Antibodies with Immunofluoreseent Antibody Technique in Buman Trichomoniasis (질트리코모나스증에서 간접형광항체법을 이용한 혈청내 항질트리코모나스 IgG 및 IgM 항체의 측정)

  • 윤경찬;김경민;안명희;민득영;차동수
    • Parasites, Hosts and Diseases
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    • v.25 no.1
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    • pp.7-12
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    • 1987
  • The indirect fluorescent antibody(IFA) test was used to detect serum IgG and IgM antibodies to Trichomonas vaginalis in 31 vaginal trichomoniasis, 7 candidiasis and in 20 non-infected healthy women with antigen prepared from axonic culture of Trichomenas vaginalis isolated from vulvovaginitis patient. The results were as follows: 1. In 31 vaginal trichomoniasis the positive reactions of IgG antibody were 27 in the 1/8 dilution or higher and :l in the 1/4 dilution whereas in healthy women the reaction showed significantly low as in the 1/4 dilution or below. 2. The sensitivity and specificity of IFA test for IgG antibody to trichomonad antigen in this study were 87.1% and 100%, respectively. 3. No significant difference of IgM antibody levels between vaginal trichomoniasis and healthy women was observed. 4. No relation between the levels of IgG and IsM antibodies to trichomonad antigen by IFA test was observed. 5. No relation between the time lapse and the level of serum IgG antibodies in IFA test of vaginal trichomoniasis was regarded. In conclusion the present study suggests that IFA test in trichomoniasis could be a useful tool for detection of anti-trichomonad IgG antibodies and applicable as an immunodiagnostic method.

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Diagnosis of Viral Infection of Pensaeide (보리새우류의 바이러스 감염증의 진단)

  • 허문수;정초록;장경립
    • Journal of Life Science
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    • v.9 no.4
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    • pp.453-459
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    • 1999
  • Baculovirus(WSBV) was isolated from infected Penaeide was collected from shrimp farm at southern sea of Korea from 1993 to 1995. The Infectious virus was purified and used for diagnosis of infected shrimp. Anti-viral serum were used for immunological detection as enzyme linked immunoabsorbent assay(ELISA) and indirect fluorescent antibody technique(IFAT). In IFAT, stomach, lymphoid organ and antenae gland of infected shrimp showed fluorescent reaction. In ELISA, tissues of spontaneously infected shrimp appeared higher O.D. values than in artificial infected shrimp. Primer set was constructed from sequence of 420bp of cloned Baculovirus(WSBV) genome. Specific band for infected shrimp was detected in Polymerase chain reaction(PCR)

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Staphylococcusaureus protein A as a means of assessing sperm penetrability in cervical mucus in vitro

  • Al-Daghistani, Hala I.
    • Clinical and Experimental Reproductive Medicine
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    • v.47 no.3
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    • pp.186-193
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    • 2020
  • Objective: The effectiveness of Staphylococcus protein A (SPA) in improving the penetration ability of sperm and reducing antisperm antibody (ASA) titers in immunologically infertile males was evaluated. Methods: Seminal fluid samples were obtained from 15 infertile men, and ASA titers were assessed with the latex agglutination test. Identification of immunoglobulin (Ig) classes and characterization of the antigens involved in the immune response were performed using indirect immunofluorescence. Local ASAs typically present as a mixture of IgG and IgA classes. The capillary tube penetration method was used to assess the capability of spermatozoa to penetrate the cervical mucus (CM). Results: ASAs associated with the neck region of sperm showed a significantly lower migration distance in the CM of infertile females than ASAs associated with the head or tail segments. ASA-positive seminal fluid exhibited significant increases in the mean migration distance (2.6 ± 1.4 cm vs. 1.54 ± 1.1 cm, respectively; p< 0.001) and sperm concentration (174 ± 121.0 × 103/mL vs. 101 ± 93.7 × 103/mL, respectively; p= 0.033) after treatment with SPA compared to pre-treated samples. A significant reduction (p< 0.01) in the recorded ASA titer was detected. Conclusion: These results indicate that SPA can be used as a sorting regimen for insemination programs. However, further studies are warranted to assess its influence on pregnancy rate.

Live Cell Detection of Monoclonal Antibody Light and Heavy Chain mRNAs using Molecular Beacons (분자 비컨을 이용한 살아 있는 세포에서 단일클론항체 경쇄와 중쇄 mRNA 검출에 의한 세포주 선별방법)

  • Jeong, Seunga;Rhee, Won Jong
    • KSBB Journal
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    • v.31 no.1
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    • pp.33-39
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    • 2016
  • Developing the method for the selection of animal cell line producing therapeutic monoclonal antibody (mAb) is invaluable as its market is rapidly growing. Although the quality of produced mAb is as important as quantity, however there is no method developed for the selective screening of cell lines on the basis of both quantity and quality. From recent reports, the ratio of light and heavy chain mRNAs of mAb in the cell is a key parameter for the indication of product quality. Therefore, it is obvious that developing the novel method that can detect both light and heavy chain mRNAs in single live cell will provide unprecedented opportunities in bio-industry. Here, we have constructed oligonucleotide probes, molecular beacons for the detection of light or heavy chain mRNAs, respectively, in the live cells producing mAbs. Both beacons showed increased fluorescent intensity after transient transfection of plasmid expressing mAbs analyzed by fluorometer. Flow cytometric analysis clearly demonstrated that both molecular beacons can simultaneously detect the expression of light and heavy chain mRNAs of mAb in the same cell. The technique described in the thesis provides the new direction and concept for developing the method for the smart selection of cell lines producing recombinant proteins including therapeutic mAbs.

Rapid diagnosis of Pseudotuberclosis in yellowtail (Seriola quinqueradiata) by immunofluorescent antibody technique (면역형광항체법(免疫螢光抗體法)에 의(依)한 방어의 유결절증(類結節症) 신속(迅速) 진단(診斷))

  • Bang, Jong-Deuk;Jung, Sung-Hee;Chun, Seh-Kyu
    • Journal of fish pathology
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    • v.3 no.1
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    • pp.11-19
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    • 1990
  • Yellow tail (Seriola quinqueradiata) infected by Pasteurella piscicida have been occurred to mass mortality without showing apparent surface lesions in cage culture farms. In this case, it is necessary to consider countermeasure by rapid diagnosis of infected fish. The purpose of the present study was to investigate usefulness of the direct fluorescent antibody technique(FAT) for rapid diagnosis of pseudotuberclosis of cultured yellowtail caused by P. piscicida. Antibody produced by inoculating rabbit with formalin killed pseudotuberclosis bacteria antigen(strain KNP-2). Immunoglobulin-G(IgG) was purified from antisera by DEAE-cellulose column chromatography and conjugate with fluorescein isothiocyanate. Fluorescein-labeled antisera was purified by sephadex G-25 gel column chromatography. The fluorescein/protein molar ratio of labeled antisera was determined as 8.8-9.5. Diagnosis of cultured yellowtail was examined in cage culture farms which located in Tongyung, kyungnam from July to October 1990. The causative bacteria of pseudotuberclosis could be detected within two hours after the specimens were transferred to the laboratory for FAT, and it showed that FAT could be adapted as a rapid and accurate diagnostic method of pseudotuberclosis in yellowtail.

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IgG Western Blot for Confirmatory Diagnosis of Equivocal Cases of Toxoplasmosis by EIA-IgG and Fluorescent Antibody Test

  • Khammari, Imen;Saghrouni, Fatma;Yaacoub, Alia;Meksi, Sondoss Gaied;Ach, Hinda;Garma, Lamia;Fathallah, Akila;Said, Moncef Ben
    • Parasites, Hosts and Diseases
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    • v.51 no.4
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    • pp.485-488
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    • 2013
  • The performance values of available techniques used in serodiagnosis of toxoplasmosis are satisfactory but they raise problems of equivocal and discordant results for very low IgG titers. Recently marketed, LDBio-Toxo II IgG Western blot (IB) showed an excellent correlation with the dye test. We estimated the proportion of equivocal and discordant results between the enzyme immunoassay Platelia Toxo IgG (EIA-IgG) and fluorescent antibody test (FAT) and assessed the usefulness of the IB as a confirmatory test. Out of 2,136 sera collected from pregnant women, 1,644 (77.0%) tested unequivocally positive and 407 (19.0%) were negative in both EIA-IgG and FAT. The remaining 85 (4%) sera showed equivocal or discordant results. Among them, 73 (85.9%) were positive and 12 (14.1%) were negative in IB. Forty-one (89.1%) equivocal sera in EIA-IgG and 46 (86.8%) equivocal sera in FAT were positive in IB. Reducing the cut-off values of both screening techniques improved significantly their sensitivity in detecting very low IgG titers at the expense of their specificity. In conclusion, equivocal results in routine-used techniques and their discordance in determination of the immune status in pregnancy women were not uncommon. IB test appeard to be highly useful in these situations as a confirmatory technique.

The Characterization of Anti-HER-2/neu Monoclonal Antibody using Different in vivo Imaging Techniques

  • Moon, Cheol;Kim, Eun Jung;Choi, Dan Bee;Kim, Byoung Soo;Kim, Sa Hyun;Choi, Tae Hyun
    • Biomedical Science Letters
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    • v.21 no.1
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    • pp.23-31
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    • 2015
  • Recently, specific antibodies have been used extensively to diagnose and treat various diseases. It is essential to assess the efficacy and specificity of antibodies, especially the in vivo environment. Anti-HER-2/neu mAb was evaluated as a possible transporting agent for radioimmunotherapy. The monoclonal antibody was successfully radio-labeled with $^{131}I$. In vitro binding assays were performed to confirm its targeting ability using another radio-iodine, $^{125}I$. Binding percentage of $^{125}I$ labeled anti-HER-2/neu mAb in HER-2/neu expressing CT-26 cells was found to be 4.5%, whereas the binding percentage of $^{125}I$ labeled anti-HER-2/neu mAb in wild-type CT-26 was only 0.45%. In vivo images were obtained and analyzed through $\gamma$-camera and an optical fluorescent modality, IVIS-200. $\gamma$-camera images showed that $^{131}I$ labeled anti-HER-2/neu mAb accumulated in HER-2/neu CT-26 tumors. Optical imaging based on near infrared fluorescence labeled anti-HER-2/neu mAb showed higher fluorescence intensities in HER-2/neu CT-26 tumors than in wild-type CT-26 tumors. Anti-HER-2/neu mAb was found to specifically bind to its receptor expressing tumor. Our study demonstrates that in vivo imaging technique is a useful method for the evaluation of an antibody's therapeutic and diagnostic potentials.

Propagation of lymphocystis disease virus (LCDV) in the FIN cell line originated from olive flounder Paralichthys olivaceus fin

  • Hossain, Mosharrof;Kim, Wi-Sik;Jung, Sung-Ju;Oh, Myung-Joo
    • Journal of fish pathology
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    • v.24 no.2
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    • pp.47-51
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    • 2011
  • The present study demonstrated lymphocystis disease virus (LCDV) propagation through cytopathic effects (CPE) formation and LCDV detection in olive flounder fin (FFN) cells by polymerase chain reaction (PCR) and fluorescent antibody technique (FAT) methods. Tissue filtrates from the cluster cells produced CPE in FFN cells, which initially cells became enlarged and gradually underwent fusion en masse. Infectivity of culture grown LCDV using the FFN cells reached $10^{2.3}$ $TCID_{50}$/ml at 4 days post infection and the highest titer was measured $10^{6.5}$ $TCID_{50}$/ml at 12 days. The viral DNA was detected in the cell culture supernatants showing CPE and the CPE cells by PCR. Antigen specific strong fluorescence reacting with monoclonal antibody against the virus revealed the presence of viral antigen in the cytoplasm of infected FFN cells. These results suggest that the FFN cell line originated from the olive flounder has a susceptibility of the LCDV.

Direct Immunofluorescence in Clinically Diagnosed Oral Lichen Planus

  • Lee, Kyung-Eun;Suh, Bong-Jik
    • Journal of Oral Medicine and Pain
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    • v.41 no.1
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    • pp.16-20
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    • 2016
  • Purpose: Oral lichen planus (OLP) is relatively common mucosal disease in clinical dentistry. OLP is intractable and regarded having malignant potential. Until now, there is some debate on how far OLP can be malignant, and which characteristics can be a risk factor for malignant transformation. Clinician need to know some differences between OLP and lesions similar to OLP to manage properly and suppose prognosis correctly. Therefore, the aim of this study was to divide clinical OLP into two groups and to compare the results of direct immunofluorescence (DIF) between two groups. Methods: This study was conducted on outpatients who visited at the department of Oral Medicine in Chonbuk National University Hospital from January 2007 to November 2015. Patients with DIF result were retrospectively reviewed. The selected patients were classified 'clinical typical of OLP' (CTO) or 'clinical compatible with OLP' (CCW) by modified World Health Organization diagnostic criteria of OLP and oral lichenoid lesion. Results: DIF were classified by deposition intensity or pattern of anti-human antibody and fibrinogen. The classification of fluorescence pattern in each specimen was graded as positive, possibly positive or negative. Conclusions: Both CTO and CCW had positive and possibly positive pattern. Prevalence of positive pattern was 68.8% in CTO and 52.6% in CCW and that of possibly positive pattern was 9.4% in CTO and 5.3% in CCW. Prevalence of negative was 21.8% in CTO and 42.1% in CCW.