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Doctoral Thesis
DOI
https://doi.org/10.11606/T.42.2014.tde-27112014-092411
Document
Author
Full name
Gustavo Cabral de Miranda
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2014
Supervisor
Committee
Gidlund, Magnus Ake (President)
Brito, Hermi Felinto de
Câmara, Niels Olsen Saraiva
Oliveira, Rodrigo Corrêa
Silva, Wilmar Dias da
Title in Portuguese
Novos biossensores baseados em anticorpos naturais e sintéticos para detecção de LDL oxidada (oxLDL) usada como biomarcador de aterosclerose.
Keywords in Portuguese
Anticorpo plástico
Aterosclerose
Biosensor
Imunossensor
oxLDL
Abstract in Portuguese
Vários estudos que tentam compreender a gênese da aterosclerose têm demonstrado evidências que a oxLDL é peça importante para o desenvolvimento da doença, tornando-a um importante marcador. Os objetivos deste trabalho foram propor um novo imunossensor empregando anticorpos monoclonais anti-oxLDL e desenvolver um processo inovador de obtenção de anticorpos plásticos anti-oxLDL. Para construção do imunossensor, a região Fc do anticorpo foi ligada ao eletrodo de trabalho do dispositivo AuSPE (Screen-Printed Gold Electrodes), utilizando cisteamina. Posteriormente, foi adicionado BSA como bloqueador de possíveis regiões livres. Após o bloqueio, o imunossensor foi testado com oxLDL e outros antígenos, como forma de garantir a especificidade. Em relação aos anticorpos plásticos, chamados MAPS, estes foram desenvolvidos contruindo uma camada impressa de forma invertida ao anticorpo plástico SPAN, referência deste trabalho. Após obtenção dos MAPSs, estes passaram por diversos testes, similar ao imunossensor. Os resultados demonstraram excelente sensibilidade e especificidade às moléculas de oxLDL com detecção em tempo real em ambas as metodologias.
Title in English
New biosensors based on natural and synthetic antibodies for the detection of oxidized LDL (oxLDL) used as a biomarker for atherosclerosis.
Keywords in English
Atherosclerosis
Biosensor
Immunosensor
oxLDL
Plastic antibody
Abstract in English
Increased levels of plasma oxLDL are associated with atherosclerosis, and the subsequent development of severe cardiovascular diseases that are today a major cause of death in modern countries. It is therefore important to find a reliable and fast assay to determine oxLDL. A new Immunosensor employing three monoclonal antibodies against oxLDL and a backside protein-surface imprinting process are proposed in this work. To generate the Immunosensor the mAbs were set-up by cysteamine on a gold layer of a disposable screen-printed electrode. BSA was immobilized further as bloker. All steps were followed by various characterization techniques such as electrochemical impedance spectroscopy and square wave voltammetry. To generate specific synthetic antibody materials, called MAPS, these were developed with a backside protein-surface imprinting process of the plastic antibody SPAN, the reference of the work. The devices were successfully applied to determine the oxLDL fraction in real serum, without prior dilution or necessary chemical treatment. Overall, these were promising results with the possibility to apply on the practical use of clinical.
 
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Publishing Date
2014-11-28
 
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