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Synthetic peptides mimic gp75 from Paracoccidioides brasiliensis in the diagnosis of paracoccidioidomycosis.

Caldini CP, Xander P, Kioshima ÉS, Bachi AL, de Camargo ZP, Mariano M, Lopes JD - Mycopathologia (2012)

Bottom Line: There was significant recognition of P2 by sera of untreated PCM patients when compared with normal human sera.These data indicate a potential use of P2 as diagnostic tool in PCM.Its application for serological diagnosis of PCM may contribute to the development and standardization of simpler, faster and highly reproducible immunodiagnostic tests at low cost.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Microbiologia, Imunologia e Parasitologia, Universidade Federal de São Paulo-Escola Paulista de Medicina, Disciplina de Imunologia, Rua Botucatu, 862, 4º andar, São Paulo, 04023-900, Brazil.

ABSTRACT
Paracoccidioidomycosis (PCM) is a systemic granulomatous disease, endemic in Latin America, caused by the thermal dimorphic fungus Paracoccidioides brasiliensis. Although some fungal antigens have already been characterized and used for serological diagnosis, cross-reactions have been frequently observed. Thus, the examination of fungal forms in clinical specimens or isolation of P. brasiliensis by culture is still the most frequent method for the diagnosis of this mycosis. In this study, a random peptide phage display library was used to select mimotopes of P. brasiliensis, which were employed as antigens in an indirect enzyme-linked immunosorbent assay. The protective monoclonal antibody against experimental PCM (anti-gp75) was used as molecular target to screen a phage display library. That approach led to a synthetic peptide named P2, which was synthesized and tested against PCM patients' sera to check whether it was recognized. There was significant recognition of P2 by sera of untreated PCM patients when compared with normal human sera. Sera from treated PCM group, patients with other mycosis or co-infected with HIV had much lower recognition of P2 than untreated patient group. The test showed a sensitivity of 100 and 94.59% of specificity in relation to human sera control. These data indicate a potential use of P2 as diagnostic tool in PCM. Its application for serological diagnosis of PCM may contribute to the development and standardization of simpler, faster and highly reproducible immunodiagnostic tests at low cost.

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Related in: MedlinePlus

Antigenic potential of P2. a Five BALB/c mice were immunized with P2 peptide emulsified in adjuvant. Fifteen days after last immunization, ELISA was performed to verify the production of polyclonal antibodies by animals. The sera of immunized mice reacted with P2, showing antigenic potential of synthetic peptide. b Recognition of gp75 by anti-P2 polyclonal antibodies. Nitrocellulose membrane with exoantigen proteins immobilized on the surface was tested with anti-P2 polyclonal antibodies. The band with molecular weight of 75 kDa was recognized by sera containing anti-P2 antibodies. Anti-gp43 were used as reaction control
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Fig3: Antigenic potential of P2. a Five BALB/c mice were immunized with P2 peptide emulsified in adjuvant. Fifteen days after last immunization, ELISA was performed to verify the production of polyclonal antibodies by animals. The sera of immunized mice reacted with P2, showing antigenic potential of synthetic peptide. b Recognition of gp75 by anti-P2 polyclonal antibodies. Nitrocellulose membrane with exoantigen proteins immobilized on the surface was tested with anti-P2 polyclonal antibodies. The band with molecular weight of 75 kDa was recognized by sera containing anti-P2 antibodies. Anti-gp43 were used as reaction control

Mentions: Humoral immune response induced by P2 synthetic peptide was evaluated by subcutaneous immunization of BALB/c mice. After three immunizations, reactivity of the mice sera was detected by ELISA. Serial dilutions of a pool of sera from mice after the last immunization were employed, and the recognition of P2 by polyclonal sera was observed (Fig. 3a). Sera from control mice injected only with adjuvant did not react with P2 synthetic peptide.Fig. 3


Synthetic peptides mimic gp75 from Paracoccidioides brasiliensis in the diagnosis of paracoccidioidomycosis.

Caldini CP, Xander P, Kioshima ÉS, Bachi AL, de Camargo ZP, Mariano M, Lopes JD - Mycopathologia (2012)

Antigenic potential of P2. a Five BALB/c mice were immunized with P2 peptide emulsified in adjuvant. Fifteen days after last immunization, ELISA was performed to verify the production of polyclonal antibodies by animals. The sera of immunized mice reacted with P2, showing antigenic potential of synthetic peptide. b Recognition of gp75 by anti-P2 polyclonal antibodies. Nitrocellulose membrane with exoantigen proteins immobilized on the surface was tested with anti-P2 polyclonal antibodies. The band with molecular weight of 75 kDa was recognized by sera containing anti-P2 antibodies. Anti-gp43 were used as reaction control
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3368115&req=5

Fig3: Antigenic potential of P2. a Five BALB/c mice were immunized with P2 peptide emulsified in adjuvant. Fifteen days after last immunization, ELISA was performed to verify the production of polyclonal antibodies by animals. The sera of immunized mice reacted with P2, showing antigenic potential of synthetic peptide. b Recognition of gp75 by anti-P2 polyclonal antibodies. Nitrocellulose membrane with exoantigen proteins immobilized on the surface was tested with anti-P2 polyclonal antibodies. The band with molecular weight of 75 kDa was recognized by sera containing anti-P2 antibodies. Anti-gp43 were used as reaction control
Mentions: Humoral immune response induced by P2 synthetic peptide was evaluated by subcutaneous immunization of BALB/c mice. After three immunizations, reactivity of the mice sera was detected by ELISA. Serial dilutions of a pool of sera from mice after the last immunization were employed, and the recognition of P2 by polyclonal sera was observed (Fig. 3a). Sera from control mice injected only with adjuvant did not react with P2 synthetic peptide.Fig. 3

Bottom Line: There was significant recognition of P2 by sera of untreated PCM patients when compared with normal human sera.These data indicate a potential use of P2 as diagnostic tool in PCM.Its application for serological diagnosis of PCM may contribute to the development and standardization of simpler, faster and highly reproducible immunodiagnostic tests at low cost.

View Article: PubMed Central - PubMed

Affiliation: Departamento de Microbiologia, Imunologia e Parasitologia, Universidade Federal de São Paulo-Escola Paulista de Medicina, Disciplina de Imunologia, Rua Botucatu, 862, 4º andar, São Paulo, 04023-900, Brazil.

ABSTRACT
Paracoccidioidomycosis (PCM) is a systemic granulomatous disease, endemic in Latin America, caused by the thermal dimorphic fungus Paracoccidioides brasiliensis. Although some fungal antigens have already been characterized and used for serological diagnosis, cross-reactions have been frequently observed. Thus, the examination of fungal forms in clinical specimens or isolation of P. brasiliensis by culture is still the most frequent method for the diagnosis of this mycosis. In this study, a random peptide phage display library was used to select mimotopes of P. brasiliensis, which were employed as antigens in an indirect enzyme-linked immunosorbent assay. The protective monoclonal antibody against experimental PCM (anti-gp75) was used as molecular target to screen a phage display library. That approach led to a synthetic peptide named P2, which was synthesized and tested against PCM patients' sera to check whether it was recognized. There was significant recognition of P2 by sera of untreated PCM patients when compared with normal human sera. Sera from treated PCM group, patients with other mycosis or co-infected with HIV had much lower recognition of P2 than untreated patient group. The test showed a sensitivity of 100 and 94.59% of specificity in relation to human sera control. These data indicate a potential use of P2 as diagnostic tool in PCM. Its application for serological diagnosis of PCM may contribute to the development and standardization of simpler, faster and highly reproducible immunodiagnostic tests at low cost.

Show MeSH
Related in: MedlinePlus