• Parasites, Hosts and Diseases
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• 제35권3호
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• pp.197-202
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• 1997

고양이 4마리의 폐흡충 피낭유충을 50마리씩 감염시키고, 1주부터 13주까지 감염단계별 혈청을 채취하고 폐흡충 충란, 피낭유충, 4주, 7주 및 16주 충체 추출물 항원에 대한 항체 반응을 관찰하였다. 초기 항채 반응은 감염 3주 후부터 관찰되었다. 항원별로는 4주 충체 항원에 대한 항체가가 가장 먼저, 높게 증가하였고, 7주, 16주 충체 항원에 대한 반응이 그 다음으로 높았다. 충란 항원에 대한 항체 반응은 10주 이후 증가하였다. 피낭유충 충체 항원에 대한 반응은 관찰한 초기 감염 전체 기간에 걸쳐 비교적 낮았다. Immunoblot을 실시한 결과, 충란 항원 특이 28, 46, 94 kDa 항원대에 대한 반응은 10주 이후부터 관찰할 수 있었다. 4주 충체 항원은 10-25, 29, 31 kDa 등이 감염 초기부터 항원성을 나타내고 있었고, 그 중에는 감염 13주 혈청에 대하여 이미 반응이 약하게 된 항원대도 있었고 다른 기생충 질환 혈청과 교차반응을 나타내는 것이 알려진 항원도 있었다. 16주 성충 충체 항원의 32, 35 kDa 항원대는 감연 4주 후부터 특이한 반응을 나타내고 있었다. 이상의 결과, 감염 경과에 따라 폐흡충 항원단백질의 항원 결정기도 성충의 것으로 바뀌는 것을 알 수 있었고 성충 충체항원의 32 및 35 kDa 단백질은 4주 이후 초기 폐흡충증도 진단할 수 있는 항원 성분으로 생각하였다.

• Asian Pacific Journal of Cancer Prevention
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• 제14권3호
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• pp.1635-1642
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• 2013

Helicobacter pylori antigen was prepared from an isolate from a patient with a duodenal ulcer. Serum samples were obtained from culture-positive H. pylori infected patients with duodenal ulcers, gastric ulcers and gastritis (n=30). As controls, three kinds of sera without detectable H. pylori IgG antibodies were used: 30 from healthy individuals without history of gastric disorders, 30 from patients who were seen in the endoscopy clinic but were H. pylori culture negative and 30 from people with other diseases. OFF-GEL electrophoresis, SDS-PAGE and Western blots of individual serum samples were used to identify protein bands with good sensitivity and specificity when probed with the above sera and HRP-conjugated anti-human IgG. Four H. pylori protein bands showed good (${\geq}$ 70%) sensitivity and high specificity (98-100%) towards anti-Helicobacter IgG antibody in culture-positive patients sera and control sera, respectively. The identities of the antigenic proteins were elucidated by mass spectrometry. The relative molecular weights and the identities of the proteins, based on MALDI TOF/TOF, were as follows: CagI (25 kDa), urease G accessory protein (25 kDa), UreB (63 kDa) and proline/pyrroline-5-carboxylate dehydrogenase (118 KDa). These identified proteins, singly and/or in combinations, may be useful for diagnosis of H. pylori infection in patients.

• Journal of Microbiology and Biotechnology
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• 제16권7호
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• pp.1026-1031
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• 2006

The optimization of the production of a fusion protein containing the antigenic domain 1 (AD-1) is of a great importance, considering its use in diagnostic tests. The fusion protein is produced by the fermentation of a recombinant strain of Escherichia coli containing the plasmid Mbg58, which expresses the AD-1 (aa 484-650) of human cytomegalovirus glycoprotein B as a fusion protein together with aa 1-375 of ${\beta}-galactosidase$. An important characteristic of promoters (lac and derivatives) used in recombinant protein production in E. coli is their inducibility. Induction by IPTG is widely used for basic research; however, its use in large-scale production is undesirable because of its high cost and toxicity. In this work, studies using different inducers and carbon sources for the production of a fusion protein containing the AD-l were performed. The results showed that lactose could be used as an inducer in the fermentation process for the production of this protein, and that expression levels could exceed those achieved with IPTG. The use of lactose for protein expression in E. coli should be extremely useful for the inexpensive, large-scale production of heterologous proteins in E. coli. Addition of sucrose to the fermentation medium improved the yield of recombinant protein, whereas addition of fructose or trehalose decreased the yield.

• Parasites, Hosts and Diseases
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• 제34권2호
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• pp.135-142
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• 1996

톡소포자충(Toxoplosma gondii) 주요막단백질의 하나인 30 kDa 단백질(p30)의 항원부위를 결정하고자 p30의 아미노산 분석에 따른 친수성 부위 및 혐수성 부위에 맞게 유전자를 증폭하고 발현시켜 항원성을 검토하였다 p30의 절편으로는 p30 전체 p30의 N-말단 Signal Sequence와 C- 탈단의 혐수성 부위를 제거한 S28. S28의 N-말단 2/3부위인 Al9. S28의 C-말단 2/3부위인 Pl9. 528의 N-탈난 1/3부위인 X9 중앙 1/3부위인 Y10 및 C-말단 1/5부위인 Z9로 구성하였다. 각절편에 대한 primer에는 EcoR I의 clampsequence를 포함시켜 중합효소반응으로 증폭시켰으며 G57를 발현하는 pGEX-4T-1 vector에 삽입시킨 후 Eschericha coli(.JM105 strain)에 형질변형시키고 IgG로 각 절편이 GST와 융합단백질로 발현되도록 하였다 SDS-PAGE상에서 p30은 63 kDa. S28는 54 kDa Al9과 Pl9은 각각 45 kDa. X9은 35 kDa. Y10은 36 kDa 및 29은 35 kDa 단백질로 발현되었다. 각각의 단백질은 westemblot상에서 GSTdetectionkit와 잘 반응하여 융합단백질임을 확인하였다. 톡소포자충증 환자 혈청과 westem blot에서 p30. S28 및 Al9은 반응하여 항원성이 인정되었으나 Pl9 . X9, Y10 및 Z9는 반응하지 않았다 따라서. p30의 중간 1/3 부위의 존재하에 N-말단 1/3부위가 항원성을 나타내는 구조적 항원이거나. 첫 1/3부위와 중 간 1/3부위의 경계에 위치한 polypeptide가 항원성을 발현하는 것으로 추정되었다.

• Journal of Veterinary Science
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• 제21권1호
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• pp.5.1-5.12
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• 2020

The major glycoproteins of bovine gammaherpesvirus 4 (BoHV-4) are gB, gH, gM, gL, and gp180 with gB, gH, and gp180 being the most glycosylated. These glycoproteins participate in cell binding while some act as neutralization targets. Glycosylation of these envelope proteins may be involved in virion protection against neutralization by antibodies. In infected cattle, BoHV-4 induces an immune response characterized by low neutralizing antibody levels or an absence of such antibodies. Therefore, virus seroneutralization in vitro cannot always be easily demonstrated. The aim of this study was to evaluate the neutralizing capacity of 2 Argentine BoHV-4 strains and to associate those findings with the gene expression profiles of the major envelope glycoproteins. Expression of genes coding for the envelope glycoproteins occurred earlier in cells infected with isolate 10/154 than in cells infected with strain 07/435, demonstrating a distinct difference between the strains. Differences in serological response can be attributed to differences in the expression of antigenic proteins or to post-translational modifications that mask neutralizing epitopes. Strain 07/435 induced significantly high titers of neutralizing antibodies in several animal species in addition to bovines. The most relevant serological differences were observed in adult animals. This is the first comprehensive analysis of the expression kinetics of genes coding for BoHV-4 glycoproteins in 2 Argentine strains (genotypes 1 and 2). The results further elucidate the BoHV-4 life cycle and may also help determine the genetic variability of the strains circulating in Argentina.

• 약학회지
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• 제50권4호
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• pp.263-267
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• 2006

CD1d is an unique antigen presenting molecule which provides antigenic repertoires to NKT cells. To examine molecules required for CD1d antigen presentation, we determined an interaction between CD1d and several endoplasmic reticulum (ER) resident molecular chaperones by co-immunoprecipitation. Results indicated that calnexin and calreticulin seem to be bound to mouse CD1d, but TAP and tapasin do not bind. Further, we screened an yeat two hybrid system to identify proteins that help mouse CD1d transportation in the cytosol. We found that two proteins, heat shock protein a sub-unit $(Hsp90{\alpha})$ and protein kinase C and casein kinase substrate in neurons 3 (PACSIN-3), interact with CD1d. Future study will be focus on the role of these molecules during the CD1d antigen presentation.

• 한국미생물·생명공학회지
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• 제25권2호
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• pp.129-136
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• 1997

Helicobacter pylori is a spiral-shaped, microaerophilic human gastric pathogen causing chronic-active gastritis in association with duodenal ulcer and gastric cancer. To investigate the possibility of H. pylori outer membrane proteins (OMPS) as the oral vaccine antigens, sarcosine-insoluble outer membrane fraction has been prepared from H. pylori NCTC 11637. The major OMPs having apparent molecular masses of 62 kDa, 54 kDa and 33 kDa were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), which were identified as urease B subunit (UreB), heat shock protein (Hsp54 kDa) and urease A subunit (UreA), respectively. Minor protein bands of 57 kDa, 52 kDa, 40 kDa, 36 kDa and 31 kDa were also observed. The antigenicity of H. pylori OMPs and antigenic cross-reactivity among the strains were determined by immunoblot analysis using anti-H. pylori OMPs antisera or intestinal lavage solutions. The results showed that UreB, Hsp54 kDa, UreA and 40 kDa proteins vigorously stimulated mucosal immune response rather than systemic immunity. From this results, these proteins seemed to be useful as the antigen candidates for the oral vaccine. The immunoblotting results with surface proteins from eight isolated H. pylori strains were similar to that of H. pylori NCTC 11637. The IgA which had been arised from oral administration of H. pylori OMPs, was able to bind H. pylori whole-cells.

• Parasites, Hosts and Diseases
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• 제26권3호
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• pp.163-168
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• 1988

스파르가눔증을 혈청학적으로 진단하기 위하여 효소면역측정법을 실시할 경우, 그 민감도와 특이도가 높다고 이미 보고되었다. 실제로 조직기생충 감염이 의심되는 환자에 대하여 특이 IgG항체가를 일상적으로 검사하면 중추신경계 환자중에서 뇌스파르가눔증은 드물지 않게 발견할 수 있다. 그러나 혈청학적 진단에 이용되는 스파르가눔 조항원(조항원)은 조충증(조충증), 유구낭미충증이나 포충증 환자의 혈청과 비특이적인 교차반응을 일으키는 경우가 있다. 이러한 교차반응의 문제를 해결하려면 우선 스파르가눔 항원의 구성 단백질이 환차혈청과 반응하는 양상을 알아야 할 필요가 있다. 이 연구에서는 외과적으로 스파르가눔증을 확인한 환자 15명에서 얻은 혈청이 스파가눔의 생리식염수 추출액의 항원단백질중 어느 분획과 반응하는지 검토하였다. 그리고 뇌유구낭미충증 환자 24명(그중 8명은 효소면역측정법으로 교차반응이 있었음)의 혈청과 교차반응을 일으키는 단백질 분획을 관찰하였다. 스파르가눔충체의 생리식염수 추출액을 10∼15% linear gradient gel에서 SDS-폴리아크릴아마이드 전기영동을 실시하여 단백질대 30개를 구별할 수 있었다. 분리한 단백질(대)를 nitrocellulose 종이에 옮긴후 스파르가눔증 환자 혈청 및 conjugate와 차례로 반응시키고 발색반응을 일으킨 결과 (효소면역 전기영동이적법, immunoblot), 스파르가눔 항원단백질중 29kDa 및 36kDa가 가장 많이 또 가장 진하게 반응한 것이어서 민감하고 항원성이 높은 단백질 분획이라고 판단하였다. 그리고 위의 단백질 이외에도 158kDa, 130kDa, 107kDa, 78kDa, 72kDa, 52kDa, 23kDa, 21kDa, 15kDa 및 6kDa 등에도 반응이 있었다. 유구낭충증 환자 24명의 혈청중 스파르가눔 항원에 혈청학적 교차반응이 있었던 혈청은 36kDa 및 29kDa 단백질에 반응하고 있어 이 단백질이 스파르가눔의 종 특이(종특이)단백질 항원은 아닌 것으로 판단되나 그 성질에 대해서는 앞으로 더 추구할 가치가 있다고 생각한다.

• BMB Reports
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• 제39권1호
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• pp.16-21
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• 2006

The coat protein (CP) of Papaya ringspot virus (PRSV) was analyzed for presentation of the antigenic peptide of animal virus, Canine parvovirus (CPV), in Escherichia coli (E. coli). The 45 nucleotides fragment coding for the 15-aa peptide epitope of the CPV-VP2 protein was either inserted into the PRSV-cp gene at the 5', 3' ends, both 5' and 3' ends or substituted into the 3' end of the PRSV cp gene. Each of the chimeric PRSV cp genes was cloned into the pRSET B vector under the control of the T7 promoter and transformed into E. coli. The recombinant coat proteins expressed from different chimeric PRSV-cp genes were purified and intraperitoneally injected into mice. All of the recombinant coat proteins showed strong immunogenicity and stimulate mice immune response. The recombinant coat proteins containing the CPV epitope insertion at the C terminus and at both N and C termini elicited ten times higher specific antisera in immunized mice compared with the other two recombinant coat proteins which contain the CPV epitope insertion at the N terminus and substitution at the C terminus.

• Clinical and Experimental Reproductive Medicine
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• 제20권2호
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• pp.137-147
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• 1993

The purpose of this study is to identify the presence of the circulating antibodies directed toward ovarian proteins{antiovarian antibodies, AOA) and the nature of antigenic ovarian structure by comparing the binding activities to 4 types of ovarian proteins, particulated and solubilized forms of pig ovarian and granulosa cell membranes in sera of patients with premature ovarian failure(POF) and to evaluate the usefulness of circulating AOA as a follow up tool after treatment. Measurements of AOA were performed by enzyme linked immunosorbent assay(ELISA) in sera of 58 patients with POF, 51 had normal chromosomes and 7 had X chromosome abnormalities. Sera of 21 natural menopausal women and 17 castrated women were also tested and sera of 32 healthy premenopausal women were served as controls. ELISA reactivities against particulated porcine granulosa cell membrane proteins was the greatest among 4 different ovarian proteins. Fifteen(29%) of 51 POF patients with normal chromosome and 1(14.3%) of 7 POF patients with X chromosome abnormalities had AOA while none of 32 controls and 21 natural menopausal women and 17 castrated women had AOA. One POF patient with 47, XXX was identified AOA positive. The ELISA reactivities were followed up monthly up to 5 months in 4 AOA positive POF patients after estrogen-progestin{E-P) therapy. There was a decreasing tendency of the ELISA reactivities in all these patients after E-P therapy and two of them converted to AOA negative. These data suggest that antigenic structure may be components of granulosa cell membrane and the determination of circulating AOA may be useful in the follow up after treatment in patients with autoimmune POF.