RESUMO
Chromosome 5q31 spans the T helper (Th) 2-related cytokine gene cluster, which is potentially important in Th1/Th2 immune responses. The chromosome 5q23.2-31.3 has been recently identified as a region with suggestive evidence of linkage to tuberculosis in the Asian population. With the aim of fine-mapping a putative tuberculosis susceptibility locus, we investigated a family-based association test between the dense single nucleotide polymorphism (SNP) markers within chromosome 5q31 and tuberculosis in 205 Thai trio families. Of these, 75 SNPs located within candidate genes covering SLC22A4, SLC22A5, IRF1, IL5, RAD50, IL13, IL4, KIF3A and SEPT8 were genotyped using the DigiTag2 assay. Association analysis revealed the most significant association with tuberculosis in haplotypes comprising SNPs rs274559, rs274554 and rs274553 of SLC22A5 gene (P(Global)=2.02 x 10(-6)), which remained significant after multiple testing correction. In addition, two haplotypes within the SLC22A4 and KIF3A region were associated with tuberculosis. Haplotypes of SLC22A5 were significantly associated with the expression levels of RAD50 and IL13. The results show that the variants carried by the haplotypes of SLC22A4, SLC22A5 and KIF3A region potentially contribute to tuberculosis susceptibility among the Thai population.
Assuntos
Cromossomos Humanos Par 5/genética , Predisposição Genética para Doença/genética , Cinesinas/genética , Proteínas de Transporte de Cátions Orgânicos/genética , Polimorfismo de Nucleotídeo Único/genética , Tuberculose/genética , Biologia Computacional , Feminino , Estudo de Associação Genômica Ampla , Genótipo , Haplótipos/genética , Humanos , Masculino , Linhagem , Membro 5 da Família 22 de Carreadores de Soluto , Simportadores , TailândiaRESUMO
Tuberculosis, a potentially fatal infectious disease, affects millions of individuals annually worldwide. Human protective immunity that contains tuberculosis after infection has not been clearly defined. To gain insight into host genetic factors, nonparametric linkage analysis was performed using high-throughput microarray-based single nucleotide polymorphism (SNP) genotyping platform, a GeneChip array comprised 59 860 bi-allelic markers, in 93 Thai families with multiple siblings, 195 individuals affected with tuberculosis. Genotyping revealed a region on chromosome 5q showing suggestive evidence of linkage with tuberculosis (Z(lr) statistics=3.01, logarithm of odds (LOD) score=2.29, empirical P-value=0.0005), and two candidate regions on chromosomes 17p and 20p by an ordered subset analysis using minimum age at onset of tuberculosis as the covariate (maximum LOD score=2.57 and 3.33, permutation P-value=0.0187 and 0.0183, respectively). These results imply a new evidence of genetic risk factors for tuberculosis in the Asian population. The significance of these ordered subset results supports a clinicopathological concept that immunological impairment in the disease differs between young and old tuberculosis patients. The linkage information from a specific ethnicity may provide unique candidate regions for the identification of the susceptibility genes and further help elucidate the immunopathogenesis of tuberculosis.
Assuntos
Povo Asiático/genética , Ligação Genética , Genoma Humano , Polimorfismo de Nucleotídeo Único , Tuberculose/genética , Idade de Início , Alelos , Criança , Cromossomos Humanos Par 17 , Cromossomos Humanos Par 20 , Cromossomos Humanos Par 5 , Família , Marcadores Genéticos , Haplótipos , Humanos , Escore Lod , Probabilidade , Irmãos , Estatísticas não Paramétricas , Tailândia , Tuberculose/imunologia , Adulto JovemRESUMO
The aim of this study was to assess the immunoglobulin (Ig)-subclass distribution of antimalarial antibody responses in 110 and 169 Thai patients with complicated and uncomplicated Plasmodium falciparum malaria, respectively. Antimalarial plasma IgG subclasses and IgE antibody levels against a crude malaria blood stages, and antigen preparation were determined using enzyme-linked immunosorbent assay (ELISA). On admission, the levels of anti-P. falciparum IgG1, IgG2 and IgG3 were significantly lower in patients with complicated malaria than uncomplicated malaria (IgG1, P < 0.0001; IgG2, P < 0.0001; IgG3, P < 0.0001). The levels of antimalarial IgE were slightly lower, but not statistically significant (P = 0.389) in the complicated malaria. After adjusting all antibody levels and age, anti-P. falciparum IgG3 levels remained significantly associated with complicated malaria. None of the other antibody concentrations showed statistically significant associations with complicated malaria. The anti-P. falciparum IgG3 levels were related to the IgG1 as well as IgG2 levels. A correlation between anti-P. falciparum IgG2 and IgE was observed in the complicated malaria group, and this may indicate their roles in the severity of disease. Our data suggest that anti-P. falciparum IgG3 is associated with a reduced risk of complicated malaria and that antimalarial Ig-subclasses are differently regulated in patients with complicated and uncomplicated malaria.
Assuntos
Anticorpos Antiprotozoários/sangue , Imunoglobulina E/sangue , Imunoglobulina G/sangue , Malária Falciparum/imunologia , Plasmodium falciparum/imunologia , Adolescente , Adulto , Idoso , Animais , Anticorpos Antiprotozoários/imunologia , Antígenos de Protozoários/imunologia , Feminino , Humanos , Imunoglobulina E/imunologia , Imunoglobulina G/imunologia , Malária Cerebral/imunologia , Malária Falciparum/parasitologia , Masculino , Pessoa de Meia-Idade , TailândiaRESUMO
The immunoglobulin G (IgG) subclass antibodies to Plasmodium falciparum blood stage antigens in the sera of 181 individuals living in malaria endemic area in Kanchanaburi Province, western Thailand, were determined by enzyme-linked immunosorbent assay (ELISA). In this study, IgG3 and IgG1 were shown to be the predominant subclasses. Generally, IgG2 were coexpressed with IgG1 and IgG3 while IgG4 was found to coexpress with other three IgG subclasses. The levels of specific IgG1, IgG2, and IgG3 increased significantly with age (r = 0.295, p = 0.000; r = 0.416, p = 0.000; r = 0.320, p = 0.000, respectively). The data indicate that the higher antibody production required continuous stimulation under natural condition. Furthermore, the levels of specific IgG1, IgG2 and IgG3 increased in immune individuals without clinical malaria, reported in adolescents and adults, were associated with malaria resistance. Similar results were found in children with different patterns of IgG subclasses in which the specific IgG2 and IgG3, but not IgG1 was related to resistance.
Assuntos
Antígenos de Protozoários/sangue , Imunoglobulina G/sangue , Malária Falciparum/imunologia , Adulto , Criança , Suscetibilidade a Doenças/imunologia , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulina G/classificação , Masculino , População Rural , TailândiaRESUMO
The total IgE and anti-Plasmodium falciparum IgE antibodies were determined by enzyme linked immunosorbent assay (ELISA) in 480 children and adults living in malaria endemic area along Thai-Myanmar border, Kanchanaburi Province, western Thailand. Approximately 73.13% of tested individuals had elevated levels of total IgE with a range of 160-998 ng/ml. 20.5% of these IgE were specific to P falciparum blood stage antigens, with a range of 78-353 microg/ml. However, the levels of total IgE were not significantly correlated with those of specific IgE (r = 0.083). The elevation of anti-P falciparum IgE antibodies seems to be age dependent. The prolonged or repeated exposure to malaria parasites is necessary for the induction of specific IgE response as indicated by the finding of a significant correlation between the levels of P falciparum specific IgE and the number of malaria attacks (r = 0.551, p = 0.01). Interestingly, among the specific IgE responders, 20 individuals naturally exposed to malaria but without clinical malaria reported had high levels of both total IgE and anti- P. falciparum IgE antibodies, with mean values of 418.67 mg/ml and 146.25 ng/ml, respectively. It is likely that the antibodies from such specific IgE responders could mediate phagocytosis in vitro.
Assuntos
Autoanticorpos/sangue , Imunoglobulina E/sangue , Malária Falciparum/imunologia , Plasmodium falciparum/imunologia , Adolescente , Adulto , Animais , Anticorpos Antiprotozoários/imunologia , Criança , Pré-Escolar , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulina E/imunologia , Malária Falciparum/epidemiologia , Masculino , Pessoa de Meia-Idade , Fagocitose , Tailândia/epidemiologiaRESUMO
Although HIV-1 subtype E associated with neurological dysfunction is common, the virological characteristics of HIV-1 isolated from the CNS for this subtype have not yet been identified. In this study, paired blood and CSF isolated from patients with AIDs-defining illnesses were cultured, sequenced and aligned. Phylogenetic tree and nucleotide-distances from both blood and CSF were investigated. Cytopathicity and co-receptor usage of paired blood and CSF isolates were compared to define the specific characteristics of CNS isolates. The results confirmed that CSF isolates showed less cytopathicity. It was found that both blood and CSF isolates used either CXCR4 or CXCR4 and CCR5 as co-receptors. Interestingly, one CSF isolate using CCR3 as a co-receptor was identified. By sequence analysis, the pair-wise distances of envelope gp 120 sequence and those of all variable regions (except V3 region) between blood and CSF isolates were significantly different. The genetic distances in V1/V2 regions of CSF isolates showed more diversity than those of blood isolates. These findings suggest that the evolution of V1/V2 regions of CSF isolates seems to be an advantage for HIV-1 in CNS infection. In contrast, the genetic distance in V4 and V5 regions of CSF isolates showed less diversity, suggesting that conservation in these regions might be necessary during the process of HIV-1 CNS infection.
Assuntos
Infecções Oportunistas Relacionadas com a AIDS/líquido cefalorraquidiano , HIV-1/genética , Sequência de Aminoácidos , Sequência de Bases , Linhagem Celular , Primers do DNA , Proteína gp120 do Envelope de HIV/química , Humanos , Macrófagos , Dados de Sequência Molecular , Fragmentos de Peptídeos/química , Fenótipo , Filogenia , Reação em Cadeia da Polimerase , Receptores de QuimiocinasRESUMO
Using an in vitro model of human lung endothelial cells, we studied different characteristics of Plasmodium falciparum isolates as potential factors for malaria severity in 2 Thai patient groups: 27 with complicated malaria and 42 with uncomplicated malaria. In regard to binding properties, no association existed between cytoadherence and rosette phenotypes (P = 0.1) and hypothrombocytemia increased the cytoadherence level (P = 0.007). Cytoadherence was significantly associated with malaria severity (P = 0.05) in contrast to rosette formation (P = 0.9). Intercellular adhesion molecule-1 and chondroitin-4-sulfate were major receptors of cytoadherence in those with complicated malaria compared with those with uncomplicated malaria (P < 10(-4)). Chondroitin-4-sulfate could act as a putative receptor for malaria complications in non-pregnant women. CD36 was the main receptor in patients with uncomplicated malaria (P < 10(-3)). Vascular cell adhesion molecule-1 and E-selectin played a minor role in 2 groups (P = 0.6). Qinghaosu derivatives were more efficient than other antimalarial drugs, but a positive correlation was observed between the 50% inhibitory concentrations of halofantrine and quinine and the number of adhesive parasitized red blood cells, suggesting their influence on cytoadherence.
Assuntos
Pulmão/patologia , Malária Falciparum/patologia , Plasmodium falciparum/química , Adolescente , Adulto , Animais , Anticorpos Monoclonais , Antimaláricos/farmacologia , Ligação Competitiva/fisiologia , Adesão Celular/fisiologia , Células Cultivadas , Sulfatos de Condroitina/fisiologia , Endotélio/patologia , Feminino , Humanos , Molécula 1 de Adesão Intercelular/fisiologia , Masculino , Pessoa de Meia-Idade , Parasitemia , Fenantrenos/farmacologia , Quinina/administração & dosagem , Quinina/uso terapêutico , Formação de Roseta , TailândiaRESUMO
Specific monoclonal antibodies (MAbs) to mefloquine conjugated to bovine serum albumin (mefloquine-BSA) were produced by hybridoma technology. The mefloquine-BSA was synthesized by converting mefloquine into hemisuccinate followed by convalently linked to bovine serum albumin (BSA) and coupling with N,N' disuccinimidyl carbonate (DSC). The conjugate was purified by Sephadex G-75 gel filtration using 0.01 M PBS pH 7.2. An average of 19.34 molecules of mefloquine were conjugated to each molecule of protein determined by differential UV absorption spectra of hapten and protein carrier. Sixteen monoclones producing antibody specific to mefloquine were screened by indirect ELISA using homologous antigens. The specificity of MAbs was determined by reacting with BSA and the structurally related antimalarial drug, quinine. Three, three, five and two MAbs belonged to IgG1, IgG2a, IgG2b and IgG3, respectively. Most of the MAbs slightly reacted with quinine-BSA due to the closely related structure of mefloquine to quinine. The selected MAb designated 11F9(G5)G9 which showed no cross reaction with quinine-BSA gave high reactivity with blood samples from malaria patients previously treated with mefloquine when compared to normal blood by indirect ELISA. The preliminary results indicated that such specific MAb could be used as antibody probe for detection of mefloquine in biological fluids.
Assuntos
Anticorpos Monoclonais/sangue , Antimaláricos/sangue , Antimaláricos/urina , Malária Falciparum/sangue , Mefloquina/sangue , Mefloquina/urina , Anticorpos Monoclonais/imunologia , Antimaláricos/imunologia , Cromatografia Líquida de Alta Pressão , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Humanos , Malária Falciparum/tratamento farmacológico , Mefloquina/imunologia , Sensibilidade e Especificidade , Soroalbumina Bovina/imunologia , TailândiaRESUMO
Cytotoxic T lymphocytes (CTLs) specific for epitope(s) within the circumsporozoite (CS) protein of malaria sporozoite have been shown to play an important role in protective immunity against malaria. Human CTLs against the potential epitope at the carboxy terminal region of CS protein of Plasmodium falciparum 7G8 strain (Pf7G8CS 368-390) were determined in thirty-six falciparum malaria patients and ten healthy controls. Four of 36 individuals and none of the healthy controls developed Pf7G8CS 368-390 specific CTL activity. The CTL activity was antigen specific and CD8+ T cell dependent. Although low CTL response has been determined, the study suggested that there was a correlation between initial parasitemia and the specific Pf7G8CS 368-390 CTL activity. A correlation between such CTL activity and anti-R32tet32 antibody levels among individuals with previous malaria experience was found, which was in contrast to those among individuals with recent malaria infection. All these 4 CTL positive individuals had at least two episodes of clinical malaria experience while all 25 individuals who were exposed to malaria for the first time did not have such a specific CTL response. These results showed that individuals with a history of natural endemic exposure to P. falciparum sporozoite developed low specific CTL responses to Pf7G8CS 368-390, so that previous but recent sporozoite exposure might be a prerequisite for generation of such CS protein specific CTL response.
Assuntos
Malária Falciparum/imunologia , Plasmodium falciparum/imunologia , Proteínas de Protozoários/imunologia , Linfócitos T Citotóxicos/imunologia , Adolescente , Adulto , Animais , Antígenos de Protozoários/imunologia , Ensaio de Imunoadsorção Enzimática , Epitopos , Feminino , Humanos , Malária Falciparum/epidemiologia , Masculino , Pessoa de Meia-Idade , Peptídeos/química , Peptídeos/imunologia , Tailândia/epidemiologiaRESUMO
Complicated malaria, caused by Plasmodium falciparum, is characterized by multiple organ dysfunction. The pathogenesis of complicated malaria involves complex host-parasite interactions that include polarized cytokine responses. Recently, correlates between Th1-like and Th2-like cytokines, especially interleukin-10 (IL), IL-12, and TNF-alpha, and specific types of organ dysfunction have been noted. Here, we measured IL-10, IL-12, and for the first time, IL-15, in 19 patients aged 16-55 years old with complicated malaria on days 0 (admission), 3, 7, and 14. For analysis, patients were grouped together or sub-categorized into hyperparasitemias or cerebral malaria (CM). For IL-10, a dramatic increase was noted on admission, followed by a reduction toward control values that closely paralleled parasite clearance. For IL-12, modest but persistent increases were noted over the entire 14 day period that did not correlate with parasitemia. In general, especially on days 0 and 3, hyperparasitemic patients had, in comparison with CM patients, higher IL-10 and IL-12 levels. In contrast, IL-15 was generally below detection in most samples. These results provide further insight into the pathogenesis of complicated malaria by strengthening the contention that cytokines such as IL-10 and IL-12 are involved in modulating the immune response to P. falciparum.
Assuntos
Interleucina-10/sangue , Interleucina-12/sangue , Interleucina-15/sangue , Malária Falciparum/sangue , Adolescente , Adulto , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Malária Falciparum/complicações , Masculino , Pessoa de Meia-IdadeRESUMO
Before field application of the direct agglutination test (DAT) for leishmaniasis, it was assessed as a diagnostic tool. Fifteen confirmed visceral leishmaniasis cases (bone marrow aspiration positive), 120 tuberculosis, 58 leprosy, 15 malaria, 26 intestinal parasitic infection cases, 24 endemic healthy controls from adjacent to the study area, and 18 controls from Kathmandu (who had never visited the VL endemic areas) were tested for anti-leishmanial antibody agglutination titers. Two of the tuberculosis cases were positive for anti-leishmanial agglutinating antibodies at 1:800. All the visceral leishmaniasis confirmed cases were reactive to anti-leishmanial antibody at > or = 1:3,200. Other specimens were negative for serology. The sensitivity of the direct agglutination test was 100% and the specificity was 99.2%. The direct agglutination test had positive and negative predictive values of 100% and 99.2% respectively. The direct agglutination test has been found to be simple, rapid, reliable, economic, safe and adaptable to micro-techniques using microtiter plates. It is specific and sensitive. The direct agglutination test is simple enough for it to be performed in a field laboratory.
Assuntos
Testes de Aglutinação/métodos , Leishmaniose Visceral/diagnóstico , Testes de Aglutinação/normas , Estudos de Avaliação como Assunto , Humanos , Leishmaniose Visceral/imunologia , Nepal/epidemiologia , Valor Preditivo dos Testes , Sensibilidade e EspecificidadeRESUMO
Monoclonal antibodies (MAbs) to quinine conjugated to a carrier protein were produced. Quinine was converted into a hemisuccinate prior to covalently linked to bovine serum albumin (BSA) by reacting with N,N'-disuccinimidyl carbonate (DSC). Coupling ratio of quinine-BSA was 13:1 calculated by spectrophotometry and 14:1 by calculation from quinine standard curve. This immunogen was used for both monoclonal antibody production and for screening test, indirect ELISA. The specificity of quinine-BSA MAbs was examined by checking the cross reactivity with BSA and the structurally related antimalarial drug, mefloquine. Six MAbs belonging to IgG1 were obtained. These MAbs slightly reacted with mefloquine-BSA because of closely related structure of mefloquine to quinine and similar conjugate preparation procedure used for conjugation. One selected MAb against quinine-BSA, showed higher reactivity with blood samples from patients previously treated with quinine when compared to normal blood. This preliminary test indicated that MAbs obtained may be useful to be used as the probe for detection of quinine in biological fluids.
Assuntos
Anticorpos Monoclonais/biossíntese , Antimaláricos/imunologia , Quinina/imunologia , Animais , Anticorpos Monoclonais/química , Anticorpos Monoclonais/imunologia , Especificidade de Anticorpos , Antimaláricos/sangue , Reações Cruzadas , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Hibridomas , Imunoglobulina G/biossíntese , Imunoglobulina G/química , Imunoglobulina G/imunologia , Mefloquina/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Quinina/sangue , Soroalbumina Bovina/química , Soroalbumina Bovina/imunologiaRESUMO
Fifty-eight monoclonal antibodies (MAbs) raised against the erythrocytic stages of Plasmodium vivax were selected for typing of 501 P. vivax isolates from different geographic locations throughout Thailand. Based on their reactivities in the indirect fluorescent antibody test, these MAbs were classified into five groups: group I MAbs showing generalized staining of all blood stages; group II MAbs reacting with merozoites and their organelles; group III MAbs reacting with the surface membrane of merozoites; Group V MAbs reacting with the surface membrane of trophozoites and schizonts; and group VII MAbs reacting with internal components of the parasites. Sixteen MAbs reacted with more than 95% of the isolates; the epitopes recognized by these MAbs were considered as being invariant. The remaining MAbs reacted with 30-90% of the isolates, and the epitopes recognized by these MAbs were regarded as being variable. The variant epitopes were associated with > 200-, 135-, and 100-kilodalton (kDa) molecules of all blood stages, the 95-kDa molecule on merozoite organelles, the 200-kDa molecule on the surface of trophozoites and schizonts, and the 85-kDa molecule of the parasite internal components. Antigenic diversity occurred among the P. vivax population in the endemic areas of Thailand and was shown to vary from place to place and was highest in the area with the highest rate of transmission along the Myanmar border in western Thailand and along the Cambodian border in eastern Thailand, including Trat (48.4%), Tak (41.7%), Chantaburi (36.5%), and Mae Hong Son (36.4%). Demonstration of antigenic diversity of P. vivax parasites signals a note of caution in the development of vaccines for vivax malaria. The vaccines should be directed against protective, conserved and not against variant epitopes.
Assuntos
Antígenos de Protozoários/genética , Plasmodium vivax/imunologia , Animais , Anticorpos Monoclonais/imunologia , Variação Antigênica , Antígenos de Protozoários/imunologia , Eletroforese em Gel de Poliacrilamida , Epitopos , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Plasmodium vivax/classificação , Plasmodium vivax/genética , Polimorfismo Genético , Especificidade da Espécie , TailândiaRESUMO
Relapse infections are an important obstacle to the successful treatment and control of Plasmodium vivax malaria, but little is known about the nature of the relapse. To provide insight into the antigenic disparity of the parasites causing initial clinical symptoms and causing relapse, a panel of 58 monoclonal antibodies (MAbs) against erythrocytic stages of Plasmodium vivax was tested by indirect fluorescent antibody test in five relapse cases. The initial and relapse strains from three patients (R3, R4, and R5) exhibited similar IFA reactivity with all MAbs tested, whereas the isolates from two relapse cases (R1 and R2) showed different patterns of reactivity and were seen only with 15 MAbs In case R1, different IFA reactivities were observed with 12 MAbs, nine of which reacted with the initial (RPV261) but not the relapse (RPV393) isolates, whereas the other three MAbs reacted only with the relapse isolates. With regards to the second relapse case (R2) in whom two relapses occurred, different IFA reactivities were demonstrated with seven MAbs that reacted only with the initial isolate (RPV 182) and with the isolate from the first relapse (RPV 240) but not with the isolate from the second relapse (RPV 300). The antibody responses from patients who developed primary clinical symptoms and relapse were detected by Western immunoblotting. In cases R3, R4 and R5, there was no difference in the spectrum of antigens from initial and relapse sera recognized by the antibodies. In contrast, in cases R1 and R2, the molecules recognized by antibodies in initial and relapse sera were markedly altered. In case R1, the series of molecules of P. vivax antigens recognized by initial (RPV 261) and relapse (RPV 393) sera were 21, 25, 31, 39, 42, 61, 95, 115, 200, > 200 kDa and 21, 24, 31, 35, 57, 75, 200, > 200 kDa, respectively. In case R2, the initial serum (RPV 182) recognized P. vivax antigens with molecular weights of 23, 30, 52, 57, 68, 75, 85, 95, 115, and 195 kDa while the first relapse (RPV 240) and the second relapse sera recognized P. vivax antigens with molecular weights of 23, 30, 52, 85, 95,115 kDa and 30, 57, 68, 75, 85,195 kDa, respectively.
Assuntos
Antígenos de Protozoários/classificação , Malária Vivax/imunologia , Plasmodium vivax/imunologia , Animais , Anticorpos Monoclonais , Variação Antigênica , Antígenos de Protozoários/genética , Antígenos de Protozoários/imunologia , Western Blotting , Eletroforese em Gel de Poliacrilamida , Epitopos , Técnica Indireta de Fluorescência para Anticorpo , Humanos , Malária Vivax/sangue , Plasmodium vivax/classificação , Recidiva , TailândiaRESUMO
Thirty-one hybridomas producing monoclonal antibodies (MAbs) against structural proteins of RSV subgroup A (Long strain) and RSV subgroup B (Japanese wild strain) were produced and separated into three groups by their reactivities with RSV-A and RSV-B using IFA. Group I was specific to RSV-A, Group II was specific to RSV-B and group III was specific to both subgroups. Characterization of selected two MAbs from each group indicated that three MAbs recognized phosphoprotein (P) and the others recognized fusion protein (F). All of the selected MAbs were IgG1 and carried kappa light chain. These selected MAbs can be used to detect the presence of RSV from NPAs and classify them into two subgroups. The infection rates of RSV in Thai children are very low and most of them were RSV subgroup A.
Assuntos
Anticorpos Monoclonais/imunologia , Vírus Sinciciais Respiratórios/imunologia , Animais , Western Blotting , Ensaio de Imunoadsorção Enzimática , Camundongos , Camundongos Endogâmicos BALB C/imunologiaRESUMO
BALB/c mice immunized with irradiated Plasmodium yoelii sporozoites produce antibodies and cytotoxic T lymphocytes against the circumsporozoite protein and against a 140-kDa protein, sporozoite surface protein 2 (PySSP2). Approximately 50% of mice immunized with P815 cells transfected with the gene encoding PySSP2 are protected against malaria, and this protection is reversed by in vivo depletion of CD8+ T cells. To determine if CD8+ T cells against PySSP2 are adequate to protect against malaria in the absence of other malaria-specific immune responses, we produced three CD8+ T-cell clones by stimulating spleen cells from mice immunized with irradiated P. yoelii sporozoites with a mitomycin-treated P815 cell clone transfected with the PySSP2 gene. Adoptive transfer of clone TSLB7 protected 100% of mice against P. yoelii. The second clone protected 58% of mice, and the third clone provided no protection. Clone TSLB7 protected even when administered 3 h after sporozoite inoculation at a time when sporozoites had entered hepatocytes, suggesting that it is recognizing and eliminating infected hepatocytes. These studies demonstrate that cytotoxic T lymphocytes against PySSP2 can protect against P. yoelii sporozoite challenge in the absence of other parasite-specific immune responses.
Assuntos
Imunoterapia Adotiva/métodos , Malária/prevenção & controle , Plasmodium yoelii/imunologia , Proteínas de Protozoários/imunologia , Linfócitos T Citotóxicos/imunologia , Animais , Antígenos CD8/imunologia , Células Clonais , Feminino , Interferon gama/biossíntese , Interleucinas/biossíntese , Malária/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Proteínas de Protozoários/genética , Proteínas Recombinantes/imunologiaRESUMO
A two-site pan-species monoclonal antibody sandwich ELISA (MAb-MAb ELISA) was developed to detect both Plasmodium vivax and P. falciparum antigens in whole blood impregnated on filter paper. In this assay, the plates were coated with pan-species MAb 3F9 and another pan-species MAb M26-32 conjugated with alkaline phosphatase was used for detection of bound antigen. The sensitivity of this assay was 5, 10 and 10 parasites per 10(6) erythrocytes for cultured P. falciparum, patient-derived P. vivax and P. falciparum, respectively. The coincidence rates for this assay were 93% (92/99) with healthy individuals and 93% (42/45) with microscopically confirmed vivax malaria cases. After two weeks treatment, 77.7% (14/18) of vivax malaria were still positive by this assay but with diminished level of reactivities [corrected].
Assuntos
Anticorpos Monoclonais , Anticorpos Antiprotozoários , Antígenos de Protozoários/sangue , Ensaio de Imunoadsorção Enzimática/métodos , Malária Falciparum/diagnóstico , Malária Vivax/diagnóstico , Plasmodium falciparum/imunologia , Plasmodium vivax/imunologia , Animais , Humanos , Malária Falciparum/imunologia , Malária Vivax/imunologia , Plasmodium cynomolgi/imunologia , Sensibilidade e EspecificidadeRESUMO
Two systems of sandwich enzyme-linked immunosorbent assay (ELISA), a two-site monoclonal antibody sandwich ELISA MAb-MAb sandwich ELISA) and a two site polyclonal-monoclonal antibody sandwich ELISA (PAb-MAb sandwich ELISA) for the detection of Plasmodium vivax antigens were developed. The assays showed good correlation with the level of parasitemia when tested against serially diluted P. vivax parasites (r = 0.937, and 0.997 for MAb-MAb and PAb-MAb sandwich ELISA, respectively), with the ability to detect as few as 6.68 parasites/10(6) erythrocytes and 2.69 parasites/10(3) erythrocytes, respectively. The MAb-MAb sandwich ELISA was specific, since it was positive only with P. vivax-infected erythrocytes from vivax malaria patients and negative when erythrocytes from 34 healthy individuals and 30 falciparum malaria cases were tested. In contrast, cross-reaction was found in the PAb-MAb sandwich ELISA when the plates were coated with polyclonal IgG and tested against the serially diluted P. falciparum SO strain antigen prepared from in vitro cultures. Comparison between the two systems of two-site sandwich ELISA showed that the MAb-MAb sandwich ELISA was superior to the PAb-MAb sandwich ELISA: (1) it gave a higher sensitivity when tested with serially diluted P. vivax antigen preparations from vivax malaria patients; (2) it gave a higher specificity when tested with the SO strain of P. falciparum from in vitro cultures, (3) it gave a lower absorbance value when tested with erythrocytes from healthy individuals. All 281 cases of vivax malaria already proven by microscopic examination were positive by MAb-MAb sandwich ELISA.(ABSTRACT TRUNCATED AT 250 WORDS)
Assuntos
Anticorpos Monoclonais , Anticorpos Antiprotozoários , Antígenos de Protozoários/sangue , Ensaio de Imunoadsorção Enzimática/métodos , Malária Vivax/diagnóstico , Plasmodium vivax/imunologia , Animais , Estudos de Avaliação como Assunto , Humanos , Malária Vivax/imunologia , Reprodutibilidade dos Testes , Sensibilidade e EspecificidadeRESUMO
A Balb/c mouse was immunized with a crude soluble antigen of Opisthorchis viverrini adult worms (OVAA) over a period of 7 months. Spleen cells from the immune mouse were fused with Sp2/0 myeloma cells. Among the 264 tissue culture wells containing the fused cells, cells of 96 wells (36%) produced antibodies to the immunizing agent. Antibodies produced by cells in several wells reacted with antigens from other species of parasite. Cells of 17 wells produced antibodies specific only to OVAA, thus cells from three representative wells were cloned by limiting dilution. Hybrids obtained produced antibodies which could be classified according to their tissue specificities into three groups. The first group of antibodies reacted strongly to the worm integument and weakly with the muscles while those belonging to the second group reacted only to muscles of the worms. The monoclonal antibodies of the third group gave a positive reaction to both muscles and tegument.