• Title/Summary/Keyword: neutralizing antibody test

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Generation of a recombinant rabies virus expressing green fluorescent protein for a virus neutralization antibody assay

  • Yang, Dong-Kun;Kim, Ha-Hyun;Park, Yu-Ri;Yoo, Jae Young;Park, Yeseul;Park, Jungwon;Hyun, Bang-Hun
    • Journal of Veterinary Science
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    • v.22 no.4
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    • pp.56.1-56.10
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    • 2021
  • Background: Fluorescent antibody virus neutralization (FAVN) test is a standard assay for quantifying rabies virus-neutralizing antibody (VNA) in serum. However, a safer rabies virus (RABV) should be used in the FAVN assay. There is a need for a new method that is economical and time-saving by eliminating the immunostaining step. Objectives: We aimed to improve the traditional FAVN method by rescuing and characterizing a new recombinant RABV expressing green fluorescent protein (GFP). Methods: A new recombinant RABV expressing GFP designated as ERAGS-GFP was rescued using a reverse genetic system. Immuno-fluorescence assay, peroxidase-linked assay, electron microscopy and reverse transcription polymerase chain reaction were performed to confirm the recombinant ERAGS-GFP virus as a RABV expressing the GFP gene. The safety of ERAGS-GFP was evaluated in 4-week-old mice. The rabies VNA titers were measured and compared with conventional FAVN and FAVN-GFP tests using VERO cells. Results: The virus propagated in VERO cells was confirmed as RABV expressing GFP. The ERAGS-GFP showed the highest titer (108.0 TCID50/mL) in VERO cells at 5 days post-inoculation, and GFP expression persisted until passage 30. The body weight of 4-week-old mice inoculated intracranially with ERAGS-GFP continued to increase and the survival rate was 100%. In 62 dog sera, the FAVN-GFP result was significantly correlated with that of conventional FAVN (r = 0.95). Conclusions: We constructed ERAGS-GFP, which could replace the challenge virus standard-11 strain used in FAVN test.

Survey of antibody to chicken anemia agent by virus neutralization test (Virus 중화시험법에 의한 닭 빈혈성인자의 항체조사)

  • Ryoo, Gwang-seon;Koh, Hong-bum
    • Korean Journal of Veterinary Research
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    • v.33 no.2
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    • pp.227-234
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    • 1993
  • A serological survey for antibody to chicken anemia agent(CAA) was carried out by virus neutralization test. Antibody to CAA was detected in broilers and layers at different age groups. The results obtained were summarized as follows ; 1. Of a total of 1,035 chicken sera from 1.16 flocks 617 samples of sera were detected as positive(59.6%) and 95 flocks of a total flocks as positive(81.9%). 2. Proportion of positive sera by age were 92.3 %(88.9~100%) at 1 to 2 weeks of age, 56.4%(16.7~77.8%) at 3 to 9 weeks of age, 85.0%(50.0~100%) at 10 to 14 weeks of age and all tested sera were positive at over the 15 weeks age. 3. In each broiler and layer chicken 63.6% and 68.4% chicks possessed positive sera respectively. 4. Neutralizing antibody titer in age group was various from 1:10 to 1:6,400 and mean titer was 1:400 to 1:800.

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Production and characterization of monoclonal antibodies against rabies virus (광견병바이러스에 대한 단크론항체 생산 및 특성)

  • Lee, Seung-Chul;Yoon, Young-Sim;Song, Yun-Kyung;Woo, Gye-Hyeong;Jean, Young-Hwa;Kang, Shien-Young
    • Korean Journal of Veterinary Service
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    • v.33 no.2
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    • pp.105-111
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    • 2010
  • Rabies virus which belongs to the genus Lyssavirus of the family Rhabdoviridae is known as a highly neurotropic virus and causes fatal encephalitis accompanied by severe neurological symptoms in almost all mammals, including humans. In this study, monoclonal antibodies (MAbs) against rabies virus were produced, characterized and applications of MAbs as diagnostic reagents were assessed Spleen and inguinal lymph node cells from Balb/c mouse immunized with purified rabies virus were fused with SP2/O myeloma cells using polyethylene glycol (PEG) and hybridoma cells producing rabies virus-specific MAbs were screened by an indirect fluorescent antibody test. A total of ten MAbs were produced against rabies virus. The protein specificity and neutralizing activity of MAbs were determined by Western blot analysis and fluorescent antibody virus neutralization test, respectively. As a result, two MAbs, 5G3 and 6H4 had specificity for nucleoprotein (N protein) and two other MAbs, 5B1 and 5C1 had neutralizing activity for rabies virus. Some MAbs recognized the rabies virus-infected bovine brain stem cells by immunohistochemistry (IHC) assay. In conclusion, it was confirmed that MAbs produced in this study were rabies virusspecific and could be used as reliable diagnostic reagents for the detection of rabies virus.

Development of Rapid Detection Technique for Canine Distemper Virus (개 디스템퍼 바이러스의 신속검출법 개발)

  • Kim, Doo;Ahn, So-Jeo;Kwon, Hyuk-Moo
    • Journal of Veterinary Clinics
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    • v.17 no.1
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    • pp.13-20
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    • 2000
  • Recently cases of canine distemper have occurred in Korea despite vaccination was carried out nationwidely. This study was performed to establish rapid diagnosis of canine distemper by RT-PCR, nested PCR, and serological test. A total of 30 dogs, which were suspected canine distemper clinically, was examined. RT-PCR and nested PCR were specific for the amplification of CDV H gene and sensitive to detect 7 TCID50 of Onderstepoort strain. By RT-PCR, H gene was detected in 6(20%) of 30 peripheral bloods from dogs. And H gene was detected in 10(33.3%) of 30 samples by nested PCR. H gene was detected from 1 brain of 6 years-old Beagle dog and 1 lung of 2 months-old Shihtzu dog, in which peripheral blood H gene was not detected. Serum neutralizing antibody titer against Onderstepoort strain ranged from 4 to 1,024 in 30 patients. No correlation was observed between the results of nested PCR and titiers of neutralizing antibody.

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Production and characterization of monoclonal antibodies against porcine transmissible gastroenteritis virus (돼지 전염성 위장염 바이러스에 대한 단크론항체 생산 및 특성)

  • Jang, Young-en;Cho, Sun-hee;Kim, Byung-han;Ahn, Jae-moon;Kang, Shien-young
    • Korean Journal of Veterinary Research
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    • v.38 no.2
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    • pp.336-344
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    • 1998
  • Eight monoclonal antibodies(MAbs) against the transmissible gastroenteritis virus (TGEV) were produced and characterized. Four of the MAbs were produced against a reference TGEV, Purdue strain(P115) and the others were produced against the Korean vaccine virus, Pyungtaek strain. Only one MAb(5C8) produced against P115 had neutralizing activity and was found to be E2 protein-specific. The other seven MAbs(4E2, 4G8, 5H6, 1F8, 2C6, 5H5, and 3A6) had specificity of nucleocapsid protein and no neutralizing activity. All MAbs reacted with different strains of TGEV, but none of the MAbs was reactive with porcine enteropathogenic viruses such as rotavirus, epidemic diarrhea virus and enterovirus by fluorescence antibody(FA) test.

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END SN Antibody Titers of Sows and Piglets Vaccinated with Living HC Vaccine During '83 Hog Cholera Outbreaks in Korea ('83 돈(豚)콜레라 유행시(流行時)의 면역모돈(免疫母豚)과 자돈(仔豚)의 END혈청중화항체가(血淸中和抗體價) 조사(調査))

  • Jeon, Yun-seong;Yeh, Jae-gil;Seo, Ik-soo
    • Korean Journal of Veterinary Research
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    • v.25 no.1
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    • pp.69-75
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    • 1985
  • Hog cholera serum neutralizing antibody of piglets and sows were titrated by means of END SN method. The piglets of a variety of ages, precolostrally immunized with LOM living HC vaccine were subjected to the test. The sows were vaccinated with lapinized living HC vaccine after 25 days from the parturition. Throughout the studies the following results were obtained and summarised. 1. Hog cholera antibody titers of inbred sows immunized with lapinized living HC vaccine after 25 days from parturition were high except Hampshire group(Table 2). 2. Sows, different stage of the pregnancy or the day of parturition, and of 3 way crossed, that were immunized with lapinized living HC vaccine have shown no significant difference on HC antibody titer(Table 2, 3). 3. HC antibody titers of piglets, immunized with a single dose of LOM HC vaccine before feeding colostrum, were high in case of the younger group(1 week) compare to the older(7 week) (Table 4). 4. The piglets that were booster immunized with LOM HC vaccine at the age of 7 weeks have shown an inconsistent antibody titers(Table 5).

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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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A Study on Serologic Diagnosis for Dengue Virus Infection

  • Sang-Wook Park;Je-Hoon Yang;Hyung-Joon Bae;Hi-Joo Moon;Young-Dae Woo
    • Biomedical Science Letters
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    • v.8 no.4
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    • pp.269-273
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    • 2002
  • Dengue fever (DF) is an acute febrile illness caused by dengue viruses in the family Flaviviridae, genus Flavivirus. DF has so far posed any problem in Korea, however it has been recently believed to be associated with oversea's traveler infected with dengue virus. Antibody titers of sera from DF patients against dengue virus were measured by indirect immunofluorescence assay (IFA) and plaque reduction neutralization test (PRNT), including the haematologic test. Three of patients with DF showed highly fluorescent and neutralizing antibody titers by IFA and PRNT assay. Two of them showed higher, remarkably. Meanwhile, one of them was tested and resulted in severe tirombocytopenia, elevated serum levels of aspartate aminotransferase (AST), alanine aminotransferase (ALT) activities as well as mild leucopenia, increased monocytes and basophils and depressed lymphocytes in haematological differential count.

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Infection Patterns of Porcine Epidemic Diarrhea Virus (PEDV) by Sera-epidemiological Analysis in Korean Pig Farms (혈청역학적 분석을 통한 한국의 돼지 유행성 설사병 바이러스 장염양상)

  • Park, Choi-Kyu;Pak, Son-Il
    • Journal of Life Science
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    • v.19 no.9
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    • pp.1304-1308
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    • 2009
  • To investigate the infection patterns of porcine epidemic diarrhea virus (PEDV) in Korean pig farms, a total of 4,768 swine sera samples from 159 pig farms were taken twice, in June (n=82) and October (n=77) in 2007. In each farm selected for the survey, 10 samples from breeding pigs and 4 from each of the 5 age groups (30, 60, 90, 120, and 150 days) were taken, and all serum samples were tested for PEDV by the serum neutralization test. The overall seroprevalence was 62.6% (2,983/4,768), with the highest prevalence in breeding pigs (93.5%, 1,485/1,589). The prevalence showed an increasing trend with increasing age (30.8, 27.2, 44.7, 61.6, and 71.2% respectively in the 30, 60, 90, 120, and 150 days age groups) (p<0.0001 for $x^2$ trend test). The association between age and PEDV prevalence was similar in both surveys, indicating that the infection of PEDV seemed to be occurring repeatedly in the farms surveyed. This inference could also be explained by the fact that prevalence in sows was very high despite low vaccination coverage, as they are continuously exposed to PEDV in potentially infected farms for a longer period. Based on the neutralizing antibody levels in sows and growing pigs, the majority of farms (91.8%, n=146 farms) were endemically infected with PEDV, and most of pigs seemed to be intensively infected with PEDV at around early growth (41.8%) and weaning (31.5%). On the other hand, serum neutralizing antibodies were not detected in pigs older than 30 days of age in farms classified as having no PEDV infection (n=13 farms), indicating the level of maternal antibody against PEDV is decreased on a non-detectable level before the piglet is 60 days old in the field situation. The results indicated that most farms surveyed in 2007 were affected with endemic PEDV infection. Therefore, a national monitoring and control program for the endemic type PEDV infection needs further attention.

Analysis of the Three Dimensional Structure of Envelope Protein of the Japanes encephalitis virus Isolated in Korea (국내에서 분리된 일본뇌염 바이러스의 Envelope Protein의 3차구조 분석)

  • Nam, Jae-Hwan;Chae, Soo-Lim;Kim, Eung-Jung;Yoon, Kyung-Sik;Lee, Ho-Dong;Koh, Hyun-Chul;Cho, Hae-Wol
    • The Journal of Korean Society of Virology
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    • v.27 no.2
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    • pp.209-216
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    • 1997
  • Three dimensional structures of envelope protein from Korean isolates and Nakayama-NIH strain of Japanese encephalitis virus (JEV) were deduced by a computer program (HyperChem 4.0 Chemplus 1.0) based on the data of the three dimentional structure of Tick-borne encephalitis virus. In the three dimensional structure of envelope protein, neutralizing epitope and T-helper cell recognition site of C-terminal region of Korean isolates were structually similar to those of Nakayama-NIH but the N-terminal region was not. Korean JE isolates were compared with Nakayama-NIH strain by using cross-neutralization antibody test. Neutralizing activities of Korean isolates derived from guinea pigs were higher than those of Nakayama-NIH strain against Korean isolates, although the polyclonal antibody titers of Nakayama-NlH showed 1:160 to 1:640 against Korean isolates. According to the results from three dimentional structures and cross-neutralization analyses, the antigenic difference between Korean JE isolates and Nakayama-NIH strain may be dependent on structural difference of envelope protein.

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