title: FC-TRIPLEX Chagas/Leish IgG1: a multiplexed flow cytometry method for differential serological diagnosis of chagas disease and leishmaniasis.
abstract: Differential serological diagnosis of Chagas disease and leishmaniasis is difficult owing to cross-reactivity resulting from the fact that the parasites that cause these pathologies share antigenic epitopes. Even with optimized serological assays that use parasite-specific recombinant antigens, inconclusive test results continue to be a problem. Therefore, new serological tests with high sensitivity and specificity are needed. In the present work, we developed and evaluated the performance of a new flow cytometric serological method, referred to as FC-TRIPLEX Chagas/Leish IgG1, for the all-in-one classification of inconclusive tests. The method uses antigens for the detection of visceral leishmaniasis, localized cutaneous leishmaniasis, and Chagas disease and is based on an inverted detuned algorithm for analysis of anti-Trypanosomatidae IgG1 reactivity. First, parasites were label with fluorescein isothiocyanate or Alexa Fluor 647 at various concentrations. Then serum samples were serially diluted, the dilutions were incubated with suspensions of mixed labeled parasites, and flow cytometric measurements were performed to determine percentages of positive fluorescent parasites. Using the new method, we obtained correct results for 76 of 80 analyzed serum samples (95% overall performance), underscoring the outstanding performance of the method. Moreover, we found that the fluorescently labeled parasite suspensions were stable during storage at room temperature, 4 °C, and -20 °C for 1 year. In addition, two different lots of parasite suspensions showed equivalent antigen recognition; that is, the two lots showed equivalent categorical segregation of anti-Trypanosomatidae IgG1 reactivity at selected serum dilutions. In conclusion, we have developed a sensitive and selective method for differential diagnosis of Chagas disease, visceral leishmaniasis, and localized cutaneous leishmaniasis.
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Andréa
authors: A
authors: Teixeira-Carvalho
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Fernanda Magalhães Freire
authors: FM
authors: Campos
authors: Laboratório de Helmintoses Intestinais, Departamento de Parasitologia, Instituto de Ciências Biológicas, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brasil.
authors: Stefan Michael
authors: SM
authors: Geiger
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Roberta Dias Rodrigues
authors: RD
authors: Rocha
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Fernanda Fortes
authors: FF
authors: de Araújo
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Danielle Marquete
authors: DM
authors: Vitelli-Avelar
authors: Departamento de Fisiopatologia, Centro de Ciências Biológicas e da Saúde, Universidade Estadual de Montes Claros, Montes Claros, Brasil.
authors: Mariléia Chaves
authors: MC
authors: Andrade
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Márcio Sobreira Silva
authors: MS
authors: Araújo
authors: Núcleo de Doenças Infecciosas, Centro Biomédico, Universidade Federal do Espírito Santo, Vitória, Brasil.
authors: Elenice Moreira
authors: EM
authors: Lemos
authors: Serviço de Pesquisa, Fundação Centro de Hematologia e Hemoterapia de Minas Gerais-HEMOMINAS, Belo Horizonte, Minas Gerais, Brasil.
authors: Anna Bárbara
authors: AB
authors: de Freitas Carneiro Proietti
authors: Departamento de Doenças Infecciosas, Instituto de Medicina Tropical, Universidade de São Paulo-USP, São Paulo, Brasil.
authors: Ester Cerdeira
authors: EC
authors: Sabino
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Rafaella Gaiotti
authors: RG
authors: Caldas
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Carolina Renata Camargos
authors: CR
authors: Freitas
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Ana Carolina
authors: AC
authors: Campi-Azevedo
authors: Departamento de Pós-Graduação em Patologia Geral, Faculdade de Medicina, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brasil; Departamento de Propedêutica Complementar, Faculdade de Medicina, Universidade Federal de Minas Gerais, Belo Horizonte, Minas Gerais, Brasil.
authors: Silvana Maria
authors: SM
authors: Elói-Santos
authors: Laboratório de Biomarcadores de Diagnóstico e Monitoração, Centro de Pesquisas René Rachou, Fundação Oswaldo Cruz, Belo Horizonte, Minas Gerais, Brasil.
authors: Olindo Assis
authors: OA
authors: Martins-Filho
mesh_terms: D014355:Chagas Disease / Q000097:blood* / Q000276:immunology / Q000469:parasitology; D003937:Diagnosis, Differential; D005434:Flow Cytometry; D006801:Humans; D007074:Immunoglobulin G / Q000097:blood* / Q000276:immunology; D007892:Leishmania braziliensis / Q000276:immunology / Q000472:pathogenicity; D016773:Leishmaniasis, Cutaneous / Q000097:blood* / Q000276:immunology / Q000469:parasitology; D012698:Serologic Tests; D014349:Trypanosoma cruzi / Q000276:immunology / Q000472:pathogenicity
chemical_list: D007074:Immunoglobulin G