Double-blind study to evaluate flow cytometry analysis of anti-live trypomastigote antibodies for monitoring treatment efficacy in cases of human Chagas' disease
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2002-09
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Resumo
The validation of flow cytometry analysis of anti-live trypomastigote antibodies (FC-ALTA) to monitor cure
after treatment of Chagas’ disease was evaluated with serum samples from treated and nontreated chagasic
patients. After optimization of the original technique, toward better sensitivity and applicability to field
surveys, we design a double blind study of 94 coded samples classified into the following categories: patients
not treated (NT) and patients treated but not cured (TNC), both presenting positive conventional serology and
xenodiagnosis; patients treated and cured (TC), showing negative serology and xenodiagnosis; and patients
treated under evaluation (TUE), who presented positive or oscillating conventional serology (CSA) but negative
xenodiagnosis. Coded samples, diluted 1:256, were assayed by incubation with live cell culture trypomastigotes,
which were subsequently stained with fluorescein isothiocyanate-conjugated anti-human immunoglobulin G,
with prior fixation and analysis by flow cytometry. The results were expressed as the percentages of positive
fluorescent parasites (PPFP) for each individual sample, establishing 20% PPFP as the cutoff between negative
and positive results. Our data demonstrated that all NT and TNC presented positive results while all but one
TC had a PPFP lower than 20%. Analysis of TUE demonstrated a wide degree of reactivity, with PPFP values
that were negative (PPFP < 20%), low positive (20% < PPFP < 50%), and high positive (PPFP > 50%). As TUE
with negative PPFP presented negative xenodiagnosis and positive or oscillating CSA, they were classified as
dissociated according to the criteria of Krettli and Brener (J. Immunol. 128:2009-2012, 1982) and could indeed
be considered cured after chemotherapy. This study demonstrates and validates the use of FC-ALTA to easily
identify anti-live trypomastigote membrane-bound antibodies, offering another approach for investigating and
monitoring the efficacy of specific chemotherapy in cases of human Chagas’ disease.
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Trypomastigote antibodies, Trypanosoma cruzi, Chagas' disease, Double-blind study
Citação
MARTINS FILHO, Olinto Assis; SANTOS, Silvana Maria Eloi; CARVALHO, Andréa Teixeira; OLIVEIRA, Rodrigo Corrêa; RASSI, Anis; LUQUETTI, Alejandro Ostermayer; RASSI, Gustavo Gabriel; BRENER, Zigman. Double-blind study to evaluate flow cytometry analysis of anti-live trypomastigote antibodies for monitoring treatment efficacy in cases of human Chagas' disease. Clinical and Diagnostic Laboratory Immunology, Washington, v. 9, n. 5, p. 1107-1113, Sept. 2002.