• Journal of Microbiology
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• v.45 no.6
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• pp.572-577
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• 2007

To enhance therapeutic potential of murine monoclonal antibody, humanization by CDR grafting is usually used to reduce immunogenic mouse residues. Most humanized antibodies still have mouse residues critical for antigen binding, but the mouse residues may evoke immune responses in humans. Previously, we constructed a new humanized version (AKA) of mouse CC49 antibody specific for tumor-associated glycoprotein, TAG-72. In this study, to select a completely human antibody light chain against TAG-72, guided selection strategy using phage display was used. The heavy chain variable region (VH) of AKA was used to guide the selection of a human TAG-72-specific light chain variable region (VL) from a human VL repertoire constructed from human PBL. Most of the selected VLs were identified to be originated from the members of the human germline VK1 family, whereas the VL of AKA is more homologous to the VK4 family. Competition binding assay of the selected Fabs with mouse CC49 suggested that the epitopes of the Fabs overlap with that of CC49. In addition, they showed better antigen-binding affinity compared to parental AKA. The selected human VLs may be used to guide the selection of human VHs to get completely human anti-TAG72 antibody.

• Parasites, Hosts and Diseases
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• v.54 no.2
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• pp.239-241
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• 2016

Chikungunya virus (CHIKV), a tropical pathogen, has re-emerged and has massive outbreaks abruptly all over the world. Containing many dominant epitopes, the envelope E2 protein of CHIKV has been explored for the vaccination or diagnosis. In the present study, the antigenicity of a recombinant expressed intrinsically disorder domain (IUD) of E2 was tested for the detection of the antibody against CHIKV through western blot method. The gene of the IUD of E2 was inserted into 2 different vectors and expressed as recombinant GST-E2 and recombinant MBP-E2 fusion protein, respectively. Two kinds of fusion proteins were tested with 30 CHIKV patient sera and 30 normal sera, respectively. Both proteins were detected by 25 patients sera (83.3%) and 1 normal serum (3.3%). This test showed a relatively high sensitivity and very high specificity of the recombinant E2 proteins to be used as diagnostic antigens against CHIKV infection.

• Journal of Periodontal and Implant Science
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• v.33 no.3
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• pp.331-340
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• 2003

본 연구의 목적은 인간의 동맥경화증에서 Porphyromonas gingivalis (P. gingivalis)와 인체 열충격단백 60의 T-세포 및 교차성 B-세포 epitope를 규명하고 수립된 T-세포주의 T-세포 주요조직적합체 양상을 파악하려는 데 있다. P. gingivalis 열충격단백-반응성 T 세포주와 환자의 혈청을 이용하여 P. gingivalis 열충격단백60 분자를 구성하는 104개의 중복성 합성 펩타이드의 T-세포 epitope과 B-세포 epitope을 규명하였다. 인체 열충격단백60에 대한 B-세포 epitope도 같은 방법으로 파악하였다. P. gingivalis, P. gingivalis 열충격단백60 또는 인체 열충격단백60에 대한 IgG 항체는 모든 동맥경화증 환자에서 상승하였다. P. gingivalis 열충격단백60의 3, 15, 24, 33, 45, 53, 64, 84, 88, 99번 펩타이드가 주요한 T-세포 epitope였고 이것들은 T-세포 및 B-세포 공동 epitope이기도 했다. 또한 인체 열충격단백60 교차반응 B-세포 epitope은 15, 29, 53, 56, 69, 74번 펩타이드로 판명되었다. 대부분 환자의 주요조직적합체는 $HLA-DRB1^{\ast}1504$와 $HLA-DZB1^{\ast}0603$으로 나타났다. 결론적으로 P. gingivalis 열충격단백60은 제 2급 주요조직적합제-제한적으로 분해되고 전달되었으며 이 단백질이 공통적인 T-세포 및 B-세포 epitope를 가지면서 동시에 인체 열충격단백60과 교차성 B-세포 epitope을 가지면서 동맥경화증의 면역조절기능에 관여한다고 볼 수 있다.

• Korean Journal of Veterinary Research
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• v.41 no.3
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• pp.343-350
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• 2001

The neutralization epitopes of the outer capsid protein VP4 of a porcine rotavirus, Gottfried strain, were studied using neutralizing monocolonal antibodies(N-MAbs). Eight N-MAbs which are specific for the VP4 of Gottfried strain were used for analyzing the antigenic sites of VP4. Three different approaches were used for this analysis; i)testing the serological reactivity of each N-MAb against different G and P types of human and animal rotavirusese ii) analyzing N-MAb-resistant viral escape mutants and iii) performing nucleotide sequence analysis of the VP4 gene of each N-MAb-resistant viral escape mutant. From experimental results, at least four antigenic sites(I, II, III, and IV) were identified. Antigenic site I recognized by N-MAbs 24B9, 23G10, and 26A2 was separated from antigenic site II recognized by N-MAbs 30H5, 32B3, and 29B3. However, these antigenic sites were overlapped with antigenic site III recognized by N-MAb 21A1. The other antigenic site IV recognized by N-MAb 16D2 was separated from antigenic sites I, II, and III.

• IMMUNE NETWORK
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• v.9 no.5
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• pp.203-207
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• 2009

Background: The FimA of Porphyromonas gingivalis is a crucial pathogenic component of the bacteria and has been implicated as a target for vaccine development against the periodontal diseases. Methods: In this study, the purified fimbriae (FimA subunit polymers) protein was used for immunization in their native form and B hybridoma clones producing antibodies specific to FimA were established. Results: The monoclonal antibodies prepared from selected two clones, designated #123 (IgG2b/ kappa) and #265 (IgG1/kappa), displayed different patterns of binding activity against the cognate antigen. Both antibodies reacted with conformational epitopes expressed by partially dissociated oligomers, but not with monomer as elucidated by Western blot analysis. Ascites fluid containing the monoclonal antibodies showed the inhibitory activity against P. gingivalis to saliva-coated hydroxyapatite beads, an in vitro model for the pellicle-coated tooth surface. Conclusion: These results suggest that the monoclonal antibodies could be used as vaccine material against the periodontal diseases through passive immunization.

• Journal of Periodontal and Implant Science
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• v.41 no.2
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• pp.54-59
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• 2011

Purpose: The aim of this study was to define the immunoreactive specificity of Porphyromonas gingivalis (P. gingivalis) heat shock protein (HSP) 60 in periodontitis and atherosclerosis. Methods: In an attempt to define the cross-reactive bacterial heat-shock protein with human self-antigen at molecular level, we have introduced a novel strategy for cloning hybridoma producing anti-P. gingivalis HSP 60 which is polyreactive to bacterial HSPs or to the human homolog. Results: Five cross-reactive clones were obtained which recognized the #19 peptide (TLVVNRLRGSLKICAVKAPG) among 37 synthetic peptides (20-mer, 5 amino acids overlapping) spanning the whole molecule of P. gingivalis HSP 60. We have also established three anti-P. gingivalis HSP 60 monoclonal antibodies demonstrating mono-specificity. These clones recognized the #29 peptide (TVPGGGTTYIRAIAALEGLK). Conclusions: Peptide #19 and #29 of P. gingivalis HSP 60 might be important immunoreactive epitopes in the immuno-pathogenic mechanism of bacterial antigen-triggered autoimmune diseases.

• BMB Reports
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• v.30 no.6
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• pp.415-420
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• 1997

Although $CD4^+$ T cell responses to protein-derived antigen have well been understood, the epitopes recognized by hapten-specific $CD4^+$ T cells have not been fully defined. In this study, we characterized the response of a T cell hybridoma (5Di0.1B8) which is specific for a hapten. N-hydroxysuccinimidyl-4-azidobenzoate (HSAB) restricted by MHC class II $I-A^d$. Using three different antigen presenting cells (APCs) expressing $I-A^d$, the role of class II MHC proteins in haptenic antigen presentation and subsequent activation of 5D10.1B8 has been examined. Activation of 5D10.1B8 T cells by HSAB analogs was also performed. Our results show that each APC activated T cells differentially and that interleukin-2 (IL-2) augmented antigen-presenting ability of all the APCs, suggesting that increased expression of class II MHC protein by IL-2 played an important role in HSAB presentation and T cell activation. Finally, early T cell receptor-dependent signals induced by HSAB or its analogs were examined by phosphotyrosine immunoblot analysis, and showed that tyrosine phosphorylation level of a 18-20 kD protein increased upon stimulation.

• BMB Reports
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• v.44 no.9
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• pp.607-612
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• 2011

Several investigators have shown that CpG-DNA has outstanding effects as a Th1-responsive adjuvant and that its potent adjuvant effects are enhanced by encapsulation with a liposome of proper composition. In this study, we showed that encapsulation with phosphatidyl-${\beta}$-oleoyl-${\gamma}$-palmitoyl ethanolamine (DOPE): cholesterol hemisuccinate (CHEMS) complex enhances the immunostimulatory activity of CpG DNA and the binding of CpG-DNA to TLR9. We also examined involvement of myeloid differentiation protein (MyD88) and NF-${\kappa}B$ activation in liposome-encapsulated CpG-DNA-induced IL-8 promoter activation. In this manuscript, the natural phosphodiester bond CpG-DNA encapsulated by DOPE : CHEMS complex is designated as Lipoplex(O). Importantly, we successfully screened B cell epitopes of envelope protein (E protein) of hepatitis C virus (HCV-E) and attachment glycoprotein G of human respiratory syncytial virus (HRSV-G) by immunization with complexes of several peptides and Lipoplex(O) without carriers. Therefore, Lipoplex(O) is potentially applicable as a universal adjuvant for peptide-based epitope screening and antibody production.

• Journal of Periodontal and Implant Science
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• v.33 no.2
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• pp.179-191
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• 2003

Since periodontal infections are suggested as risk factors for the development of cardiovascular diseases, the present study was performed to evaluate the T cell immune responses specific to Pophylomonas gingivalis(P. gingivalis) heat shock protein(hsp) 60 and T-cell and B-cell epitope specificities for P. gingivalis hsp60 in atherosclerosis. Anti-P, gingivalis IgG antibody titers were elevated in all patients. We could establish P. gingivalis hsp-specific T cell lines from the atheroma lesions, a mixture of $CD4^+$ and $CD8^+$ cells producing the cytokines characteristic of both Th1 and Th2 subsets. of 108 overlapping synthetic peptides spanning whole P. gingivalis hsp60 molecule, ten peptides with common epitopes specificities for both T-cell and B-cell were identified. it was concluded that P. gingivalis hsp60 might K involved in the immunoregulatory process of atherosclerotic diseases with epitope specificities.

• Journal of Periodontal and Implant Science
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• v.37 no.1
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• pp.11-21
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• 2007

Heat shock protein (HSP) is one of cellular protein commonly present in major periodontopathogenic bacteria as well as mammalian cells. The protein may play a role in the immunopathogenesis by modulating autoimmune reaction due to its high level of sequence homology between bacteria and human counterpart. Hence, identifying immunodomiant epitope of bacteria HSP that is cross-reactive to periodontopathogenic bacteria with a specificity to human HSP may comprise a critical strategy for development of a periodontal vaccine. The present study was performed to establish clones producing monoclonal antibody reactive to Porphyromonas gingivalis (p. gingivalis) HSP with a specificity to human HSP. 4 different hybridomas were cloned producing monoclonal IgG antibodies to P, gingivalis HSP and evaluated for their reactivity and specificity to other periodontopathogenic bacteria as well as to human HSP. These four monoclonal antibodies reacted with p. gingivalis HSP only with specificities to other bacteria tested and human HSP as well. The antigenic epitopes producing the 4 monoclonal antibody may be potentially developed as vaccine candidates. Further investigations are under way to identify more clones producing monoclonal antibodies reactive to P, gingivalis HSP and to other periodontopathogenic bacteria as well, while maintaining specificities to human counterpart.