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1.
J Adv Res ; 36: 133-145, 2022 02.
Artigo em Inglês | MEDLINE | ID: mdl-35116173

RESUMO

Introduction: The COVID-19 global pandemic is far from ending. There is an urgent need to identify applicable biomarkers for early predicting the outcome of COVID-19. Growing evidences have revealed that SARS-CoV-2 specific antibodies evolved with disease progression and severity in COIVD-19 patients. Objectives: We assumed that antibodies may serve as biomarkers for predicting the clinical outcome of hospitalized COVID-19 patients on admission. Methods: By taking advantage of a newly developed SARS-CoV-2 proteome microarray, we surveyed IgG responses against 20 proteins of SARS-CoV-2 in 1034 hospitalized COVID-19 patients on admission and followed till 66 days. The microarray results were further correlated with clinical information, laboratory test results and patient outcomes. Cox proportional hazards model was used to explore the association between SARS-CoV-2 specific antibodies and COVID-19 mortality. Results: Nonsurvivors (n = 955) induced higher levels of IgG responses against most of non-structural proteins than survivors (n = 79) on admission. In particular, the magnitude of IgG antibodies against 8 non-structural proteins (NSP1, NSP4, NSP7, NSP8, NSP9, NSP10, RdRp, and NSP14) and 2 accessory proteins (ORF3b and ORF9b) possessed significant predictive power for patient death, even after further adjustments for demographics, comorbidities, and common laboratory biomarkers for disease severity (all with p trend < 0.05). Additionally, IgG responses to all of these 10 non-structural/accessory proteins were also associated with the severity of disease, and differential kinetics and serum positive rate of these IgG responses were confirmed in COVID-19 patients of varying severities within 20 days after symptoms onset. The area under curves (AUCs) for these IgG responses, determined by computational cross-validations, were between 0.62 and 0.71. Conclusions: Our findings might have important implications for improving clinical management of COVID-19 patients.


Assuntos
COVID-19 , Anticorpos Antivirais , Humanos , Imunoglobulina G , SARS-CoV-2 , Índice de Gravidade de Doença
2.
Curr Med Sci ; 41(6): 1052-1064, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34935114

RESUMO

The ongoing Coronavirus disease 19 pandemic has likely changed the world in ways not seen in the past. Neutralizing antibody (NAb) assays play an important role in the management of the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) outbreak. Using these tools, we can assess the presence and duration of antibody-mediated protection in naturally infected individuals, screen convalescent plasma preparations for donation, test the efficacy of immunotherapy, and analyze NAb titers and persistence after vaccination to predict vaccine-induced protective effects. This review briefly summarizes the various methods used for the detection of SARS-CoV-2 NAbs and compares their advantages and disadvantages to facilitate their development and clinical application.


Assuntos
Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Teste Sorológico para COVID-19/métodos , COVID-19/imunologia , Testes de Neutralização/métodos , SARS-CoV-2/imunologia , COVID-19/prevenção & controle , COVID-19/terapia , Teste Sorológico para COVID-19/tendências , Vacinas contra COVID-19/farmacologia , Humanos , Imunização Passiva , Testes de Neutralização/tendências , Pandemias/prevenção & controle , Soroterapia para COVID-19
3.
Curr Med Sci ; 41(6): 1081-1086, 2021 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-34741251

RESUMO

OBJECTIVE: The ongoing COVID-19 pandemic warrants accelerated efforts to test vaccine candidates. To explore the influencing factors on vaccine-induced effects, antibody responses to an inactivated SARS-CoV-2 vaccine in healthy individuals who were not previously infected by COVID-19 were assessed. METHODS: All subjects aged 18-60 years who did not have SARS-CoV-2 infection at the time of screening from June 19, 2021, to July 02, 2021, were approached for inclusion. All participants received two doses of inactivated SARS-CoV-2 vaccine. Serum IgM and IgG antibodies were detected using a commercial kit after the second dose of vaccination. A positive result was defined as 10 AU/mL or more and a negative result as less than 10 AU/mL. This retrospective study included 97 infection-naïve individuals (mean age 35.6 years; 37.1% male, 62.9% female). RESULTS: The seropositive rates of IgM and IgG antibody responses elicited after the second dose of inactivated SARS-CoV-2 vaccine were 3.1% and 74.2%, respectively. IgG antibody levels were significantly higher than IgM levels (P<0.0001). Sex had no effect on IgM and IgG antibody response after the second dose. The mean anti-IgG level in older persons (⩾42 years) was significantly lower than that of younger recipients. There was a significantly lower antibody level at > 42 days compared to that at 0-20 days (P<0.05) and 21-31 days (P<0.05) after the second dose. CONCLUSION: IgG antibody response could be induced by inactivated SARS-CoV-2 vaccine in healthy individuals (>18 years), which can be influenced by age and detection time after the second dose of vaccination.


Assuntos
Anticorpos Antivirais/sangue , Vacinas contra COVID-19/farmacologia , COVID-19/imunologia , COVID-19/prevenção & controle , SARS-CoV-2/imunologia , Vacinas de Produtos Inativados/farmacologia , Adolescente , Adulto , Fatores Etários , COVID-19/epidemiologia , Vacinas contra COVID-19/administração & dosagem , Vacinas contra COVID-19/imunologia , China/epidemiologia , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina M/sangue , Masculino , Pessoa de Meia-Idade , Pandemias , Estudos Retrospectivos , Vacinas de Produtos Inativados/administração & dosagem , Vacinas de Produtos Inativados/imunologia , Adulto Jovem
4.
Cell Discov ; 7(1): 67, 2021 Aug 17.
Artigo em Inglês | MEDLINE | ID: mdl-34400612

RESUMO

One of the best ways to control COVID-19 is vaccination. Among the various SARS-CoV-2 vaccines, inactivated virus vaccines have been widely applied in China and many other countries. To understand the underlying protective mechanism of these vaccines, it is necessary to systematically analyze the humoral responses that are triggered. By utilizing a SARS-CoV-2 microarray with 21 proteins and 197 peptides that fully cover the spike protein, antibody response profiles of 59 serum samples collected from 32 volunteers immunized with the inactivated virus vaccine BBIBP-CorV were generated. For this set of samples, the microarray results correlated with the neutralization titers of the authentic virus, and two peptides (S1-5 and S2-22) were identified as potential biomarkers for assessing the effectiveness of vaccination. Moreover, by comparing immunized volunteers to convalescent and hospitalized COVID-19 patients, the N protein, NSP7, and S2-78 were identified as potential biomarkers for differentiating COVID-19 patients from individuals vaccinated with the inactivated SARS-CoV-2 vaccine. The comprehensive profile of humoral responses against the inactivated SARS-CoV-2 vaccine will facilitate a deeper understanding of the vaccine and provide potential biomarkers for inactivated virus vaccine-related applications.

5.
Allergy ; 76(2): 551-561, 2021 02.
Artigo em Inglês | MEDLINE | ID: mdl-33040337

RESUMO

BACKGROUND: The missing asymptomatic COVID-19 infections have been overlooked because of the imperfect sensitivity of the nucleic acid testing (NAT). Globally understanding the humoral immunity in asymptomatic carriers will provide scientific knowledge for developing serological tests, improving early identification, and implementing more rational control strategies against the pandemic. MEASURE: Utilizing both NAT and commercial kits for serum IgM and IgG antibodies, we extensively screened 11 766 epidemiologically suspected individuals on enrollment and 63 asymptomatic individuals were detected and recruited. Sixty-three healthy individuals and 51 mild patients without any preexisting conditions were set as controls. Serum IgM and IgG profiles were further probed using a SARS-CoV-2 proteome microarray, and neutralizing antibody was detected by a pseudotyped virus neutralization assay system. The dynamics of antibodies were analyzed with exposure time or symptoms onset. RESULTS: A combination test of NAT and serological testing for IgM antibody discovered 55.5% of the total of 63 asymptomatic infections, which significantly raises the detection sensitivity when compared with the NAT alone (19%). Serum proteome microarray analysis demonstrated that asymptomatics mainly produced IgM and IgG antibodies against S1 and N proteins out of 20 proteins of SARS-CoV-2. Different from strong and persistent N-specific antibodies, S1-specific IgM responses, which evolved in asymptomatic individuals as early as the seventh day after exposure, peaked on days from 17 days to 25 days, and then disappeared in two months, might be used as an early diagnostic biomarker. 11.8% (6/51) mild patients and 38.1% (24/63) asymptomatic individuals did not produce neutralizing antibody. In particular, neutralizing antibody in asymptomatics gradually vanished in two months. CONCLUSION: Our findings might have important implications for the definition of asymptomatic COVID-19 infections, diagnosis, serological survey, public health, and immunization strategies.


Assuntos
Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , COVID-19/imunologia , Portador Sadio/imunologia , SARS-CoV-2/imunologia , Adulto , Idoso , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , COVID-19/sangue , COVID-19/diagnóstico , Teste para COVID-19/métodos , Portador Sadio/sangue , Portador Sadio/diagnóstico , Feminino , Humanos , Imunoglobulina G/sangue , Imunoglobulina G/imunologia , Imunoglobulina M/sangue , Imunoglobulina M/imunologia , Masculino , Pessoa de Meia-Idade
6.
Chin Med J (Engl) ; 120(14): 1220-5, 2007 Jul 20.
Artigo em Inglês | MEDLINE | ID: mdl-17697571

RESUMO

BACKGROUND: Tuberculosis remains the leading cause of human death. Currently, Bacillus Calmette-Guérin (BCG) is the only available vaccine against tuberculosis but its efficacy is highly variable. Thus, developing new tuberculosis vaccines becomes an urgent task. In this study, we evaluated in BALB/c mice the humoral and cellular immune responses of recombinant BCG expressing the antigen ESAT-6 from Mycobacterium tuberculosis. METHODS: Escherichia coli-BCG shuttle plasmid named pDE22-esat-6 was constructed by inserting the BamHI/EcoRI digested esat-6 gene PCR product into the similarly digested parental plasmid pDE22. BCG cells were transformed with pDE22-esat-6, which was named recombinant BCG (rBCG). BALB/c mice were immunized subcutaneously on the back with 100 microl normal saline containing 10(6) CFU of BCG or rBCG. They were sacrificed after 4 weeks to detect their humoral and cellular responses. RESULTS: There was no any significant differences in the growth characteristics between the conventional BCG and rBCG. In immunized mice, the IgG antibody titres of rBCG group were as high as 1:8000, which was significantly higher than that in BCG group (1:1400, P < 0.05). The elicited IFN-gamma level of rBCG group was (1993 +/- 106) pg/ml, which was also significantly higher than that in BCG group ((1463 +/- 105) pg/ml, P < 0.05). The splenocyte proliferation index of rBCG group reached 4.34 +/- 0.31, which was higher than that of BCG group (3.79 +/- 0.24, P < 0.05). CONCLUSION: rBCG secreted expressing antigen ESAT-6 stimulated stronger humoral and cellular immune responses than BCG did, and, therefore may be the better vaccine against mycobacterium tuberculosis.


Assuntos
Antígenos de Bactérias/genética , Vacina BCG/imunologia , Proteínas de Bactérias/genética , Mycobacterium tuberculosis/imunologia , Vacinas Sintéticas/imunologia , Animais , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Interferon gama/biossíntese , Ativação Linfocitária , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes/imunologia
7.
Acta Biochim Biophys Sin (Shanghai) ; 38(10): 683-90, 2006 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-17033714

RESUMO

The live vaccine Mycobacterium bovis bacillus Calmette-Guérin (BCG) provides variable efficacy against adult pulmonary tuberculosis (TB). Recombinant BCG, expressing either immunodominant antigens or Th1 cytokines, is a promising strategy for developing a new TB vaccine. However, not much is known about whether the introduction of cytokine and specific antigen genes concurrently into the BCG strain could improve the immunogenicity of BCG. In this study, a recombinant BCG strain (rBCG) expressing the fusion protein human interleukin (IL)-2 and ESAT-6 (early secreted antigenic target-6 kDa) antigen of Mycobacterium tuberculosis was constructed. Six weeks after BALB/c mice (H-2d) were immunized with 106 colony forming units (CFUs) BCG or rBCG, splenocyte proliferation was determined with MTT [3-(4,5-dimethylthiazolyl-2)-2, 5-diphenyltetrazolium bromide] assay, IL-4 and interferon (IFN)-gamma produced by splenocytes were tested by enzyme linked immunosorbent assay (ELISA,) and the cytotoxicity of splenocytes from immunized mice to P815 cells (H-2d) expressing ESAT-6 protein was measured using CytoTox 96 Non-Radioactive Cytotoxicity Assay. Compared with native BCG-vaccinated mice, rBCG induced stronger Th1 responses that were confirmed by high lymphoproliferative responses and IFN-gamma production to culture filtrate protein (CFP) or ESAT-6 protein. Moreover, rBCG induced significant enhanced CTL responses against P815-ESAT-6 cells. Results from rBCG-immunized mice demonstrated that introducing the il-2 and esat-6 genes into BCG could enhance Th1 type immune responses to ESAT-6. Further investigation is needed by introducing other Th1 cytokines and antigens into BCG to optimize the protective efficacy against TB.


Assuntos
Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Interleucina-2/imunologia , Mycobacterium bovis/genética , Proteínas Recombinantes de Fusão/imunologia , Animais , Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Células Cultivadas , Feminino , Humanos , Interleucina-2/genética , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium bovis/imunologia , Proteínas Recombinantes de Fusão/genética , Células Th1/imunologia
9.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 21(3): 287-9, 2005 May.
Artigo em Chinês | MEDLINE | ID: mdl-15862141

RESUMO

AIM: To investigate the effects of rBCG vaccination containing foreign antigen Der p2 in the form of lipoprotein on murine immune response. METHODS: 6 to 8 weeks old and newborn BALB/c mice were vaccined intraperitoneally with 10(6) CFU rBCG or BCG. At the same time, the control group was injected with saline. Six weeks later, all animals were injected with Der p2 (20 microg). After two weeks later, the concentrations of IL-4 and IFN-gamma in the serum and splenocyte culture supernatant (STLCS) were determined by ELISA, and Th subgroups were determined by double fluorescent staining and flow cytometry. RESULTS: After vaccination, the serum and STLCS from both rBCG-immunized and BCG-immunized group of adult and newborn BALB/c mice had significantly higher level of IFN-gamma and lower level of IL-4 than those from control groups. Besides, there was the larger percentage of CD4 (+) IFN-gamma (+) cells in spleen from rBCG-vaccined and BCG-vaccined mice than that from control group. However, the percentage of CD4 (+) IL-4 (+) cells in spleen cells from rBCG-vaccined and BCG-vaccined group was lower than that from control group. Moreover, the level of IFN-gamma in STLCS from rBCG-immunized was significantly higher, compared with that from BCG-immunized mice. At the same time, the percentage of CD4 (+) IFN-gamma (+) cells in spleen from rBCG-vaccined mice was larger than that from BCG-vaccined group. CONCLUSION: Both rBCG and BCG could stimulate Th1 predominant immune response, when injected intraperitoneally into adult or newborn BALB/c mice, The Der p2 expressed on the cell wall of BCG can work as the component of BCG and be recognized by the immune system of mice, therefore stimulates Der p2-specific Th1 predominant immune response. These data indicate that recombinant BCG-expressing antigens can be used as the antigen-specific vaccines against allergic diseases by regulating the balance of Th1/Th2.


Assuntos
Antígenos de Dermatophagoides/genética , Antígenos de Dermatophagoides/imunologia , Lipoproteínas/genética , Lipoproteínas/imunologia , Mycobacterium bovis/genética , Animais , Antígenos de Dermatophagoides/administração & dosagem , Parede Celular/genética , Expressão Gênica , Engenharia Genética , Imunização , Interferon gama/sangue , Interferon gama/metabolismo , Interleucina-4/sangue , Interleucina-4/metabolismo , Lipoproteínas/administração & dosagem , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium bovis/citologia , Baço/citologia , Baço/imunologia , Baço/metabolismo
10.
Zhonghua Jie He He Hu Xi Za Zhi ; 27(4): 249-52, 2004 Apr.
Artigo em Chinês | MEDLINE | ID: mdl-15144616

RESUMO

OBJECTIVE: To construct the E. coli.-BCG (Bacille Calmette-Guerin) shuttle vector expressing Mycobacterium tuberculosis secreted protein Ag85B-ESAT-6 on the surface of Mycobacterium vaccae. METHODS: The gene fragment containing 19 000 antigen (19-ss) were amplified by polymerase chain reaction (PCR) from the Mycobacterium tuberculosis H(37)Ra. We cloned the 19ss gene into the E. coli.-BCG shuttle vector pOLYG and named the pCW, which can shuttle and express exogenous antigen gene on cell wall of Mycobacterium. Then Mycobacterium tuberculosis secret protein Ag85B and ESAT-6 gene were cloned into the vector and determined by indirect immunofluorescence. RESULTS: The sequence of 19-ss gene was identified with Genbank reported by sequencing. The constructed E. coli.-BCG shuttle vector using 19ss gene had the function of shuttle between E. coli. and Mycobacteria. By indirect immunofluorescence technique the secreted protein Ag85B-ESAT-6 can be fused and expressed on surface of Mycobacterium vaccae. CONCLUSION: The E. coli.-BCG shuttle vector is constructed successfully which could express exogenous antigen gene as a chimeric exported membrane.


Assuntos
Aciltransferases/biossíntese , Antígenos de Bactérias/biossíntese , Vacina BCG , Proteínas de Bactérias/biossíntese , Vetores Genéticos , Mycobacterium/genética , Aciltransferases/genética , Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Escherichia coli/genética , Escherichia coli/metabolismo , Mycobacterium tuberculosis/genética , Plasmídeos , Proteínas Recombinantes de Fusão/biossíntese , Proteínas Recombinantes de Fusão/genética , Vacinas de DNA
11.
Zhonghua Jie He He Hu Xi Za Zhi ; 27(2): 89-92, 2004 Feb.
Artigo em Chinês | MEDLINE | ID: mdl-14990181

RESUMO

OBJECTIVE: To investigate the fused expression of secreted protein Ag85B-ESAT6 of Mycobacterium tuberculosis, and to provide a promising preventive subunit vaccine against tuberculosis. METHODS: The gene encoding Ag85B and ESAT6 protein was amplified by PCR from genome of Mycobacterium tuberculosis H(37)Rv strain, and inserted into cloning vector P(GEM)-T-easy. After sequence analysis, and digestion by restriction endonuclease, Ag85B-ESAT6 was cloned into corresponding sites of the expression vector P(PRO) EXHT, and the recombinant plasmid was transformed into expressive strain E. coli DH5 alpha, induced with IPTG and fusion protein was purified by Ni-NTA purification system. The specific antibody titer in the sera of BALB/c mouse immunized with two fusion protein was detected by ELISA. RESULTS: The sequences of Ag85B and ESAT6 by PCR amplification were identical to those reported by GenBank. The recombinant plasmid fused expression protein of Ag85B-ESAT6 with relative molecular mass (Mr) of 37,000, which was confirmed by Western-blot analysis with specific monoclonal antibody against 6 x HismAb. The fused expression protein was insoluble. It could be purified by Ni-NTA purification system. The specific antibody titer in the sera of BALB/c mouse immunized with fusion Ag85B-ESAT6 was 1:1000 and that of mouse immunized with fusion protein ESAT6-Ag85B was 1:5000. CONCLUSIONS: Secreted protein Ag85B-ESAT6 of Mycobacterium tuberculosis was successfully fused expressed in E. Coli DH5 alpha. It may become a new type of vaccine against tuberculosis.


Assuntos
Aciltransferases/genética , Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Proteínas Recombinantes de Fusão/biossíntese , Aciltransferases/imunologia , Animais , Anticorpos Antibacterianos/sangue , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Western Blotting , Eletroforese em Gel de Poliacrilamida , Interferon gama/biossíntese , Camundongos , Camundongos Endogâmicos BALB C , Proteínas Recombinantes de Fusão/imunologia , Proteínas Recombinantes de Fusão/isolamento & purificação
12.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 19(1): 90-2, 2003 Jan.
Artigo em Chinês | MEDLINE | ID: mdl-15132919

RESUMO

AIM: To explore protective efficacy of pTB30m and pTB30s encoding Ag85B protein against infection with M. tuberculosis H (37)R (v). METHODS: BALB/c mice were infected intravenously with 5x10(5) CFU of M. tuberculosisH (37)R (v) six weeks after the last vaccination of pTB30m and pTB30s. At the same time, normal BALB/c mice were injected intravenously with 5x10(6) T cells separated from immunized BALB/c mouse spleen through nylon wool column, and challenges were injected with 10(5) CFU of M. tuberculosis immediately intravenously. After 4 weeks, the CFU in spleens of infected mice were counted respectively. RESULTS: Vaccination with pTB30m and pTB30s produced significant protection against M. tuberculosis proliferation in the mouse spleens following challenge. As compared with the saline-injected mice, CFU in spleens of the mice vaccinated with pTB30m or pTB30s were reduced significantly, being 0.645(log(10) CFU, P<0.01) and 0.839(log(10) CFU, P<0.001), respectively. In contrast, bacterial load in the mice spleens vaccinated with empty plasmid only had little reduction. After adoptive immunization by T cells from the mice vaccinated with pTB30m and pTB30s, the mice might induce partial protection against M. tuberculosis proliferation in the mouse spleens following challenge. CONCLUSION: Protective efficacy of the mice immunized with pTB30s against challenge with M. tuberculosis virulent strain was better than that immunized with pTB30m.Thus, pTB30s is a promising DNA vaccine with respect to the prevention and treatment of tuberculosis.


Assuntos
Aciltransferases/imunologia , Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Mycobacterium tuberculosis/imunologia , Vacinas contra a Tuberculose/imunologia , Tuberculose/prevenção & controle , Vacinas de DNA/imunologia , Aciltransferases/genética , Transferência Adotiva , Animais , Antígenos de Bactérias/genética , Proteínas de Bactérias/genética , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Tuberculose/imunologia
13.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 19(2): 132-5, 2003 Mar.
Artigo em Chinês | MEDLINE | ID: mdl-15151748

RESUMO

AIM: To construct the E.coli-BCG shuttle vector carrying and expressing dust mite antigen gene Der p2 on cell wall of mycobacterium vaccae. METHODS: The gene fragment encoding 19 kDa antigen and the upstream control element (19-ss) was amplified by PCR from the mycobacteria tuberculosis H37Rv.Subsequently, the 19-ss gene was cloned into the E.coli-BCG shuttle vector pOLYG, which can schlepped and expressed exogenous antigen gene on cell wall of mycobacteria and containing the Der p2 gene. The expression of Der p2 gene in mycobacterium vaccae determined by indirect immunofluorescence staining. RESULTS: Sequencing proved that the cloned sequence of 19-ss gene was correct. The constructed E.coli-BCG shuttle vector (pCW) containing 19-ss gene had function of shuttle between E.coli and mycobacteria, and mediated the expression of antibiotic resistance gene. Indirect immunofluorescence staining indicated that the Der p2 gene was expressed in the form of lipoprotein on surface of the mycobacterium vaccae. CONCLUSION: E.coli-BCG shuttle vector has been constructed successfully, which can express exogenous antigen gene as a chimeric protein on cell wall.


Assuntos
Escherichia coli , Mycobacterium bovis , Parede Celular , Clonagem Molecular , Escherichia coli/genética , Vetores Genéticos , Lipoproteínas/genética , Mycobacterium bovis/metabolismo , Mycobacterium tuberculosis/genética
14.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 19(3): 260-2, 2003 May.
Artigo em Chinês | MEDLINE | ID: mdl-15155088

RESUMO

AIM: To investigate the effects of plasmid containing mouse IL-12 and human IL-18 genes on the humoral immune response of mice immunized by CFP10 gene of Mycobacterium tuberculosis (MTB) H(37)R(v) strain. METHODS: Human IL-18 cDNA was amplified from RNA of PBMCs by RT-PCR and cloned into the pGEM-Teasy vector. After sequencing it was subcloned into the the sites of BamH I and EcoR I digestion of pcDNA3.1. BALB/c mice were injected intramuscularly by eukaryotic expression plasmid pcmIL12 and pcIL18, together with MTB CFP10 DNA vaccine, respectively. The same immunization repeated three times at intervals of two weeks. Mouse sera were collected at two weeks after the each injection. The titer of anti-CFP10 antibody was detected by ELISA. RESULTS: IL-18 cDNA was amplified successfully from RNA of human PBMCs by RT-PCR and the result of sequencing was correct. The IL-18 gene was correctly inserted into the vector pcDNA3.1 by being confirmed with BamH I and EcoR I digestion analysis, positive plasmid was called pcIL18. After being immunized with pcCFP10 three times, the end point titer of anti-CFP10 was 1:4 000, while the titer obtained by being immunized with pcIL18 + pcCFP10 was 1:8 000, but yet, after being immunized with pcmIL12+pcCFP10, the end point titer of anti-CFP10 antibody was only 1:200. CONCLUSION: Combination of IL-18 gene with MTB CFP10 DNA vaccine can enhance the humoral immune responses to pcCFP10, whereas the immunization with IL-12 gene plus pcCFP10 made humoral immune response markedly descent. As for whether IL-18 gene plus MTB CFP10 DNA vaccine can induce markedly the cellular mediated immune response to CFP10 or not remains to be further investigated.


Assuntos
Formação de Anticorpos , Vacinas de DNA , Animais , Humanos , Imunização , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium tuberculosis/imunologia , Vacinas de DNA/imunologia
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