IQUIR   05412
INSTITUTO DE QUIMICA ROSARIO
Unidad Ejecutora - UE
artículos
Título:
Favorably orienting recombinant proteins to develop amperometric biosensors to diagnose Chagas’ disease
Autor/es:
BELLUZO, M. S.; RIBONE, M. E.; CAMUSSONE, C.; MARCIPAR, I. S.; LAGIER, C. M.
Revista:
ANALYTICAL BIOCHEMISTRY
Editorial:
ACADEMIC PRESS INC ELSEVIER SCIENCE
Referencias:
Año: 2010
ISSN:
0003-2697
Resumen:
Clinical immunoassays often display suitable sensitivity but some lack of specificity or vice versa.As a tradeoff between specificity improvement and sensitivity loss, biosensors were designed to perform indirect immunoassays with amperometric detection using tailor-made chimeric receptors to react with the analyte, specific anti-Trypanosoma cruzi immunoglobulin G (IgG). Recombinant chimeras were designed to favor their oriented covalent attachment. This allows the chimeras to properly expose their epitopes, to efficiently capture the analyte, and towithstand severe chemical treatment to reuse the biosensors. By further binding the secondary antibody, horseradish peroxidase-labeled anti-human IgG, in the presence of the solublemediator and the enzyme substrate, a current that increasedwith the analyte concentrationwasmeasured. Biosensors using the chimeric constructions showed 100% specificitywith samples that had revealed false-positive results when using other bioreceptors. A protein bearing a poly-Lys chain and thioredoxin as directing elements displayed the highest signal-to-noise ratio (P < 0.05). The limit of detection was 62 ng ml-1, which is eight times lower than that obtained with a currently used commercial Chagas enzyme-linked immunosorbent assay (ELISA) kit. Reusability of the biosensor was assessed. The signal was approximately 80% of the original one after performing 10 consecutive determinations.