Since 2010, variant strains of porcine epidemic diarrhea virus(PEDV) have caused disasters in the pork industry. The spike(S) protein, as the major immunity-eliciting antigen, has previously been used for serological ...Since 2010, variant strains of porcine epidemic diarrhea virus(PEDV) have caused disasters in the pork industry. The spike(S) protein, as the major immunity-eliciting antigen, has previously been used for serological testing and has been found to correlate significantly with the results of the serum neutralization(SN) test. However, further evaluation of this method is needed as new epidemic strains of PEDV emerge. Hence, the main objective of this study was to assess sow sera and determine the correlation between enzyme-linked immunosorbent assay(ELISA) results(involving a newly isolated GDS01 virus-based ELISA and ELISAs based on seven recombinant fragments comprising overlapping S1 and partial S2 sequences) and SN titers. Furthermore, we determined the reliability of the ELISAs based on receiver operating characteristics(ROC) curve analyses. For the most promising ELISA, i.e., the SP4 ELISA, the correlation coefficient(r) and the area under curve(AUC) were determined to be 0.6113 and 0.8538, respectively. In addition, we analyzed the homology of the SP4 sequences obtained from different strains(including vaccine strains) and found that various strains showed a high degree of homology in this region. Thus, we conclude that SP4 is a promising serological testing protein for use in the field.展开更多
AIM： To investigate the immunogenicity of H pylori proteins, to evaluate the production rate of anti H pylori IgG antibodies in relation to time and to demonstrate the fidelity of newly optimized in-house enzymelinke...AIM： To investigate the immunogenicity of H pylori proteins, to evaluate the production rate of anti H pylori IgG antibodies in relation to time and to demonstrate the fidelity of newly optimized in-house enzymelinked immunosorbent assay （ELISA） technique as an alternative for H pylori infection assay. METHODS： In the present study, 100 μg of formalinfixed H pylori whole cell antigens was injected into an experimental animal （New Zealand white female rabbit） intramuscularly on d 0, 16, 27 and 35. The first two doses were injected with adjuvants. On d 0, a serum sample was collected from the rabbit before immunization and this pre-immunized serum was used as a negative control for the whole study. To evaluate the immunogenic responses of the injected antigen, serum samples were collected from the rabbit at regular intervals up to d 42. The sera were analyzed using inhouse ELISA and Western blot techniques.RESULTS： The production of anti Hpylor/IgG antibodies in the rabbit in response to the injected antigen increased almost exponentially up to d 14 and after that it was maintained at the same level until the last day （d 42）. By analyzing the immune profiles of immunized sera, 11 proteins were identified to be immunogenic, among them 2 （approximately 100 kDa and 85 kDa） were most prominent. CONCLUSION： Analysis of the immune responses against pathogenic microorganisms like H py/ori is necessary for the development of various diagnostic and preventive approaches. The results of this experiment reveal that the formalin-fixed H pylori whole cell antigens injected into the rabbit are highly immunogenic. These prominent proteins （approximately 100 kDa and 85 kDa） might have higher immunogenic effects among humans infected with H pylori and some of these immunogenic proteins can be included in diagnostic approaches based on serology and also for vaccine formulation. The in- house ELISA is a promising alternative compared to invasive techniques.展开更多
To identify a cost-efficient altemative antibody testing strategy for screening and confirmation of HIV infection by rapid simple tests （RSTs） and enzyme-linked immunosorbent assays （ELISAs）. Methods Four RSTs （R...To identify a cost-efficient altemative antibody testing strategy for screening and confirmation of HIV infection by rapid simple tests （RSTs） and enzyme-linked immunosorbent assays （ELISAs）. Methods Four RSTs （RST1, RST2, RST3, and RST4 ） and five ELISAs （ELISA1, ELISA2, ELISA3, ELISA4, and ELISA5） were evaluated in two phases by using banked and serum specimens prospectively collected at regional hospitals and voluntary counseling and testing （VCT） centers in Beijing. A total of 200 banked serum specimens were included in the first phase, including 62 HIV-positive, 127 HIV-negative and 11 indeterminate specimens. All specimens were tested by four RSTs and five ELISAs respectively. The second phase involved prospective testing of 389 routine specimens, including 92 HIV-positive, 287 HIV-negative, and 10 indeterminate specimens. All the specimens were tested by two RSTs （RST2 and RST4） and three ELISAs （ELISA1, ELISA3, and ELISA4）, which were selected for their respective excellent sensitivity and/or specificity. Westem blot （WB） was used as a gold standard for confirming the reactivity of all the specimens. Results Sensitivity, specificity, and efficacy were calculated for each assay in two phases. In the first phase, four assays （ELISA4, RST2, RST3, and RST4） had a specificity of 100%. For the determination of efficacy, ELISA4, RST2, and RST4 were selected in the second phase. ELISA1 and ELISA3 which have a sensitivity of 95.9% and 93.2% respectively also entered this phase. In the second phase, all the five assays （ELISA1, ELISA3, ELISA4, RST2, and RST4） had a sensitivity and specifity of over 90%. ELISA1 had a sensitivity of 99% and ELISA4 a specificity of 99%. Conclusion The sensitivity ELISA1 and the specificit of ELISA4 are comparable to ELISA/WB standard strategy. Application of this alternative testing strategy provides a cost-effective method for determining HIV prevalence in Beijing.展开更多
基金supported by the National Key Research and Development Program(2016YFD0500101)
文摘Since 2010, variant strains of porcine epidemic diarrhea virus(PEDV) have caused disasters in the pork industry. The spike(S) protein, as the major immunity-eliciting antigen, has previously been used for serological testing and has been found to correlate significantly with the results of the serum neutralization(SN) test. However, further evaluation of this method is needed as new epidemic strains of PEDV emerge. Hence, the main objective of this study was to assess sow sera and determine the correlation between enzyme-linked immunosorbent assay(ELISA) results(involving a newly isolated GDS01 virus-based ELISA and ELISAs based on seven recombinant fragments comprising overlapping S1 and partial S2 sequences) and SN titers. Furthermore, we determined the reliability of the ELISAs based on receiver operating characteristics(ROC) curve analyses. For the most promising ELISA, i.e., the SP4 ELISA, the correlation coefficient(r) and the area under curve(AUC) were determined to be 0.6113 and 0.8538, respectively. In addition, we analyzed the homology of the SP4 sequences obtained from different strains(including vaccine strains) and found that various strains showed a high degree of homology in this region. Thus, we conclude that SP4 is a promising serological testing protein for use in the field.
基金Research Funds of Microbiology Department, Dhaka University
文摘AIM： To investigate the immunogenicity of H pylori proteins, to evaluate the production rate of anti H pylori IgG antibodies in relation to time and to demonstrate the fidelity of newly optimized in-house enzymelinked immunosorbent assay （ELISA） technique as an alternative for H pylori infection assay. METHODS： In the present study, 100 μg of formalinfixed H pylori whole cell antigens was injected into an experimental animal （New Zealand white female rabbit） intramuscularly on d 0, 16, 27 and 35. The first two doses were injected with adjuvants. On d 0, a serum sample was collected from the rabbit before immunization and this pre-immunized serum was used as a negative control for the whole study. To evaluate the immunogenic responses of the injected antigen, serum samples were collected from the rabbit at regular intervals up to d 42. The sera were analyzed using inhouse ELISA and Western blot techniques.RESULTS： The production of anti Hpylor/IgG antibodies in the rabbit in response to the injected antigen increased almost exponentially up to d 14 and after that it was maintained at the same level until the last day （d 42）. By analyzing the immune profiles of immunized sera, 11 proteins were identified to be immunogenic, among them 2 （approximately 100 kDa and 85 kDa） were most prominent. CONCLUSION： Analysis of the immune responses against pathogenic microorganisms like H py/ori is necessary for the development of various diagnostic and preventive approaches. The results of this experiment reveal that the formalin-fixed H pylori whole cell antigens injected into the rabbit are highly immunogenic. These prominent proteins （approximately 100 kDa and 85 kDa） might have higher immunogenic effects among humans infected with H pylori and some of these immunogenic proteins can be included in diagnostic approaches based on serology and also for vaccine formulation. The in- house ELISA is a promising alternative compared to invasive techniques.
基金As part of the AIDS Prevention Applied Research and Program of the Ministry of Health (WA2003-17)the present study is supportedby the Department of Reference,National Center for AIDS Control and Prevention,Chinese Center for Disease Control and Prevention.
文摘To identify a cost-efficient altemative antibody testing strategy for screening and confirmation of HIV infection by rapid simple tests （RSTs） and enzyme-linked immunosorbent assays （ELISAs）. Methods Four RSTs （RST1, RST2, RST3, and RST4 ） and five ELISAs （ELISA1, ELISA2, ELISA3, ELISA4, and ELISA5） were evaluated in two phases by using banked and serum specimens prospectively collected at regional hospitals and voluntary counseling and testing （VCT） centers in Beijing. A total of 200 banked serum specimens were included in the first phase, including 62 HIV-positive, 127 HIV-negative and 11 indeterminate specimens. All specimens were tested by four RSTs and five ELISAs respectively. The second phase involved prospective testing of 389 routine specimens, including 92 HIV-positive, 287 HIV-negative, and 10 indeterminate specimens. All the specimens were tested by two RSTs （RST2 and RST4） and three ELISAs （ELISA1, ELISA3, and ELISA4）, which were selected for their respective excellent sensitivity and/or specificity. Westem blot （WB） was used as a gold standard for confirming the reactivity of all the specimens. Results Sensitivity, specificity, and efficacy were calculated for each assay in two phases. In the first phase, four assays （ELISA4, RST2, RST3, and RST4） had a specificity of 100%. For the determination of efficacy, ELISA4, RST2, and RST4 were selected in the second phase. ELISA1 and ELISA3 which have a sensitivity of 95.9% and 93.2% respectively also entered this phase. In the second phase, all the five assays （ELISA1, ELISA3, ELISA4, RST2, and RST4） had a sensitivity and specifity of over 90%. ELISA1 had a sensitivity of 99% and ELISA4 a specificity of 99%. Conclusion The sensitivity ELISA1 and the specificit of ELISA4 are comparable to ELISA/WB standard strategy. Application of this alternative testing strategy provides a cost-effective method for determining HIV prevalence in Beijing.
