• Journal of Microbiology and Biotechnology
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• 제17권6호
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• pp.993-1001
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• 2007

The technique of serological analysis of antigens by recombinant cDNA expression library (SEREX) uses autologous patient sera as a screening probe to isolate tumor-associated antigens for various tumor types. Isolation of tumor-associated antigens that are specifically reactive with patient sera, but not with normal sera, is important to avoid false-positive and autoimmunogenic antigens for the cancer immunotherapy. Here, we describe a selection methodology to isolate patient sera-specific antigens from a yeast surface-expressed cDNA library constructed from 15 patient lung tissues with non-small cell lung cancer (NSCLC). Several rounds of positive selection using patient sera alone as a screening probe isolated clones exhibiting comparable reactivity with both patient and normal sera. However, the combination of negative selection with allogeneic normal sera to remove antigens reactive with normal sera and subsequent positive selection with patient sera efficiently enriched patient sera-specific antigens. Using the selection methodology described here, we isolated 3 known and 5 unknown proteins, which have not been isolated previously, but and potentially associated with NSCLC.

• Tuberculosis and Respiratory Diseases
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• 제47권6호
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• pp.757-767
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• 1999

Background: Diagnosis by smear and/or cultures of the Mycobacterium tuberculosis from body fluid or biopsy specimen is "Gold standard". However the sensitivity of the direct microscopy is relatively low and culture of mycobacteria is time consuming. Despite an explosion in the techniques of rapid identification of mycobacteria by molecular genetic means, it is laborious and expensive and then rapid, inexpensive serodiagnosis is interested in diagnosis of tuberculosis. But sensitivity and specificity of known serologic antigen is not full sufficient level and then new antigen develop and combination cocktails of new developed antigens by ELISA are needed. Method: To compare the efficacy of different mycobacterial specific antigen and to assess the applicability of the combination of several different antigens in the diagnosis of tuberculosis, five ELISA tests derived 14KDa, 16KDa, 19KDa, 23KDa, 38KDa were evaluated in 57 active pulmonary patient and 24 inactive post-therapy follow up patient and 48 normal control. Results: The optical densities of ELISA test with 14KDa, 16KDa, 19KDa, 23KDa, 38KDa were significantly higher in active tuberculosis cases than in normal control(P<0.001, P<0.001, P<0.027, P<0.001, P<0.001) and those with 16KDa, 38KDa were significant higher in active tuberculosis cases than in inactive post-therapy follow up cases(P<0.01. P<0.001) and those of 14KDa, 16KDa, 23KDa, 38KDa were significant higher in inactive post-therapy follow up cases than in normal control(P<0.008. P<0.01. P<0.006. P<0.001). The sensitivity of 14KDa, 16KDa, 19KDa, 23KDa, 38KDa in active pulmonary patient cases was 42.1%, 43.9%, 15.8%, 28.0%, 70.2%, respectively and the specificity of 14KDa, 16KDa, 19KDa, 23KDa, 38KDa in active pulmonary patient cases was 95.8%, 95.8%, 91.7%, 89.6%, 93.8%, respectively. The sensitivity and specificity of combination 38KDa with 16KDa was 87% and 93.7%. Conclusion: The sensitivity and specificity of new antigens for serodiagnosis of the tuberculosis still remains limited at around 70%, which makes its a poor diagnostic tool for disease confirmation. A combination of cocktail antigens provided by cut-off value adjustment for serodiagnosis of tuberculosis some improved diagnostic yield than single antigen serologic test.

• 대한의생명과학회지
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• 제23권3호
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• pp.161-165
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• 2017

It has well reported that host immune system is closely related to cancer growth and eradication. Among cancer immunotherapy, cancer vaccine is focused on this review. Cancer vaccine is using host immune system against various tumor antigens to treat cancer. We discuss the classification and characteristics of the preventive vaccine, therapeutic vaccine and combination cancer immunotherapy.

• Parasites, Hosts and Diseases
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• 제39권2호
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• pp.171-176
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• 2001

Western blot analysis was performed to diagnose vivax malaria using stage-specific recombinant antigens. Genomic DNA from the whole blood of a malaria patient was used as templates to amplify the coding regions for the antigenic domains of circumsporozoite protein (CSP-1), meroxoite surface protein (MSP-1), apical merozoite antigen (AMA- 1), serine repeat antigen (SERA), and exported antigen (EXP- 1) of Plasmodium vivax. Each amplified DNA fragment was inserted into a pGEX-4T plasmid to induce the expression of GST fusion protein in Escherichia coli by IPTG. The bacterial cell extracts were separated on 10% SDS-PAGE followed by western blot analysis with patient sera which was confirmed by blood smear examination. When applied with patient sera, 147 (91.9%) out of 160 vivax malaria, 12 (92.3%) out of 13 falciparum malaria, and all 9 vivax/falciparum mixed malaria reacted with at least one antigen, while no reactions occurred with 20 normal uninfected sera. In the case of vivax malaria, CSP-1 reacted with 128 (80.0%) sera, MSP-1 with 102 (63.8%), AMA-1 with 128 (80.0%), SERA with 115 (71.9%), and EXP-1 with 89 (55.6%), respectively. We obtained higher detection rates when using S antigens (91.9%) rather than using each antigen solely (55.6 - 80%), a combination of 2 (76.3 - 87.5%), 3 (85.6 - 90.6%), or 4 antigens (89.4 - 91.3%). This method can be applied to serological diagnosis, mass screening in endemic regions, or safety test in transfusion of prevalent vivax malaria.

• Journal of Microbiology and Biotechnology
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• 제25권4호
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• pp.526-536
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• 2015

Leptospirosis is a worldwide zoonotic disease caused by pathogenic Leptospira, a genus of which more than 250 serovars have been identified. Commercial bacterin vaccines are limited in that they lack both cross-protection against heterologous serovars and long-term protection. This study investigated in mice the immunogenicity of an anti-leptospirosis vaccine, using the outer membrane proteins LipL32 and Loa22 as antigens. The immunogenicity of this vaccine formulation was compared with those induced by vaccines based on LipL32 or Loa22 alone. A DNA-encapsulated chitosan nanoparticle was used for in vivo DNA delivery. Using a unique DNA plasmid expressing both lipL32 and loa22 for vaccination, higher antibody responses were induced than when combining plasmids harboring each gene separately. Therefore, this formulation was used to test the immunogenicity when administered by a heterologous prime (DNA)-boost (protein) immunization regimen. The specific antibody responses against LipL32 (total IgG and IgG1) and Loa22 (IgG1) were higher in mice receiving two antigens in combination than in those vaccinated with a single antigen alone. Although no significant difference in splenic CD4⁺ T cell proliferation was observed among all groups of vaccinated mice, splenocytes from mice vaccinated with two antigens exhibited higher interferon-γ and IL-2 production than when using single antigens alone upon in vitro restimulation. Taken together, the immunogenicity induced by LipL32 and Loa22 antigens in a heterologous primeboost immunization regimen using chitosan as a DNA delivery system induces higher immune response, and may be useful for developing a better vaccine for leptospirosis.

• Parasites, Hosts and Diseases
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• 제55권1호
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• pp.15-20
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• 2017

The aim of this study was to identify antigens for a vaccine or drug target to control rabbit coccidiosis. A combination of 2-dimensional electrophoresis, immunoblotting, and mass spectrometric analysis were used to identify novel antigens from the sporozoites of Eimeria stiedae. Protein spots were recognized by the sera of New Zealand rabbits infected artificially with E. stiedae. The proteins were characterized by matrix-assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF/TOF-MS) analysis in combination with bioinformatics. Approximately 868 protein spots were detected by silver-staining, and a total of 41 immunoreactive protein spots were recognized by anti-E. stiedae sera. Finally, 23 protein spots were successfully identified. The proteins such as heat shock protein 70 and aspartyl protease may have potential as immunodiagnostic or vaccine antigens. The immunoreactive proteins were found to possess a wide range of biological functions. This study is the first to report the proteins recognized by sera of infected rabbits with E. stiedae, which might be helpful in identifying potential targets for vaccine development to control rabbit coccidiosis.

• Asian Pacific Journal of Cancer Prevention
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• 제15권6호
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• pp.2635-2640
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• 2014

Sera of cancer patients may contain antibodies that react with a unique group of autologous cellular antigens called tumor-associated antigens (TAAs). The present study aimed to determine whether a mini-array of multiple TAAs would enhance antibody detection and be a useful approach in esophageal cancer detection and diagnosis. Our mini-array of multiple TAAs consisted of eleven antigens, p53, pl6, Impl, CyclinB1, C-myc, RalA, p62, Survivin, Koc, CyclinD1 and CyclinE full-length recombinant proteins. Enzyme-linked immunosorbent assays (ELISA) were used to detect autoantibodies against eleven selected TAAs in 174 sera from patients with esophageal cancer, as well as 242 sera from normal individuals. In addition, positive results of ELISA were confirmed by Western blotting. In a parallel screening trial, with the successive addition of antigen to a final total of eleven TAAs, there was a stepwise increase in positive antibody reactions. The eleven TAAs were the best parallel combination, and the sensitivity and specificity in diagnosing esophageal cancer was 75.3% and 81.0%, respectively. The positive and negative predictive values were 74.0% and 82.0%, respectively, indicating that the parallel assay of eleven TAAs raised the diagnostic precision significantly. In addition, the levels of antibodies to seven antigens, comprising p53, Impl, C-myc, RalA, p62, Survivin, and CyclinD1, were significantly different in various stages of esophageal cancer, which showed that autoantibodies may be involved in the pathogenesis and progression of esophageal cancer. All in all, this study further supports our previous hypothesis that a combination of antibodies might acquire higher sensitivity for the diagnosis of certain types of cancer. A customized mini-array of multiple carefully-selected TAAs is able to enhance autoantibody detection in the immunodiagnosis of esophageal cancer and autoantibodies to TAAs might be reference indicators of clinical stage.

• 대한수의학회지
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• 제53권4호
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• pp.199-205
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• 2013

Avian pathogenic Escherichia coli (APEC) is a causative agent for a number of extra intestinal diseases and account for significant losses to the poultry industry. Since protective immunity against APEC is largely directed to virulence antigens, we have individually expressed four different viulence antigens, papA, papG, IutA, and CS31A, using an attenuated Salmonella Typhimurium and a plasmid pBB244. Following oral immunization of mice with combination of two or four of these strains, serum IgG and mucosal IgA responses were elicited against each antigen represented in the mixture. The antigen-specific mucosal IgA responses were significantly higher in the group of mice immunized with the heat-labile Escherichia coli enterotoxin B subunit (LTB) strain than those in the group of mice immunized without the LTB strain. While, there was no significant difference between these two groups in antigen-specific serum IgG responses. The results showed that LTB could act as mucosal immune adjuvant. To assess the nature of immunity, the distribution of antigen-specific IgG isotypes was analyzed. All groups promoted Th1-type immunity as determined by the IgG2a/IgG1 ratio. Thus, our findings provided evidence that immunization with a combination of several vaccine strains is one of the strategies of developing effective vaccines against APEC.

• 생명과학회지
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• 제28권4호
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• pp.496-507
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• 2018

톡소이드는 독성은 제거되고 항원성은 유지시킨 독소 단백질로써, 다양한 병원체의 감염 및 질병 예방을 위해 지속적으로 연구 되었다. 그러나, 톡소이드의 활성 감소 및 이와 함께 사용하는 어쥬번트의 부작용 등이 지속적으로 보고되면서, 면역성은 강화하고 어쥬번트의 사용은 줄일 수 있는 톡소이드 항원 전달 시스템이 필요하게 되었다. 따라서, 이러한 단점을 개선하고자 최근 새로운 백신과 약물 전달수송을 위해 다양한 분야에서 활용하고 있는 마이크로/나노 운반체를 톡소이드 항원에 도입하고 있다. 이와 같은 마이크로/나노 운반체는 미생물 자체를 이용하거나 미생물을 통해 생산해 낼 수도 있으며, 더 나아가 다양한 소재의 폴리머를 이용하여 제작할 수 있다. 본 총설에서는 톡소이드 항원 전달을 위한 마이크로/나노 운반체를 미생물 유래의 ghost cells (GCs), 그람 음성 세균이 분비하는 outer membrane vesicles (OMVs) 및 고분자 폴리머로 구성된 nanoparticles (NPs)으로 분류하였다. 마지막으로 각 운반체에 대한 톡소이드 항원의 전달 방식 및 이를 적용하였을 때 일어나는 면역반응에 대하여 서술하였으며, 이를 통해 향후 톡소이드의 효율 및 부작용이 개선되기를 기대한다.

• IMMUNE NETWORK
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• 제14권2호
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• pp.73-80
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• 2014

Because autoimmune diseases (AIDs) result from a complex combination of genetic and epigenetic factors, as well as an altered immune response to endogenous or exogenous antigens, systems biology approaches have been widely applied. The use of multi-omics approaches, including blood transcriptomics, genomics, epigenetics, proteomics, and metabolomics, not only allow for the discovery of a number of biomarkers but also will provide new directions for further translational AIDs applications. Systems biology approaches rely on high-throughput techniques with data analysis platforms that leverage the assessment of genes, proteins, metabolites, and network analysis of complex biologic or pathways implicated in specific AID conditions. To facilitate the discovery of validated and qualified biomarkers, better-coordinated multi-omics approaches and standardized translational research, in combination with the skills of biologists, clinicians, engineers, and bioinformaticians, are required.