Imunossensor impedimétrico para determinação de rituximabe aplicado ao controle qualidade e monitoramento clínico

• Autor(a) principal: Rodrigues, Edson Silvio Batista
• Data de Publicação: 2023
• Tipo de documento: Tese
• Idioma: por
• Título da fonte: Repositório Institucional da UFG
• Texto Completo: http://repositorio.bc.ufg.br/tede/handle/tede/12938
• topic: Imunoensaio; Proteína CD20; Espectroscopia de impedância eletroquímica; Monitoramento terapêutico de fármacos; Testes on-site; Immunoassay; CD20 protein; Electrochemical impedance spectroscopy; Therapeutic drug monitoring; On-site testing; CIENCIAS DA SAUDE::FARMACIA::FARMACOTECNIA

Introduction: Cancer is an umbrella term for a large group of diseases that can affect any part of the body. Rituximab (RTX) is a chimeric monoclonal antibody (mAb) formed by the mouse/human combination, which specifically binds to the CD20 cell marker that is expressed in normal and malignant B lymphocytes, exerting significant antitumor activity. In this context, RTX emerged as a therapeutic alternative in the treatment of various pathologies, including autoimmune diseases, inflammatory diseases and various types of cancer in which sensitive and low-cost methods are required for therapeutic drug monitoring (TDM) and quality control (QC) of this drug. Objectives: To develop a selective electrochemical immunosensor for the detection and quantification of RTX in urine and blood. Methodology: The ploliconal antibody (pAb) was purified from chicken egg, used as a biorecognition element to create an immunosensor for the detection and quantification of RTX. The immunosensor was constructed from the immobilization of the RTX IgY antibody on the glassy carbon electrode surfaces. The characterization of the immunosensor was obtained through monitoring at each stage via cyclic voltammetry and electrochemical impedance spectroscopy, where selectivity is obtained by specifically binding pAb and RTX and casein is used to block possible nonspecific interactions of other proteins. After the development of the immunosensor, the validation of the method was carried out, evaluating the selectivity, accuracy, precision, linearity and limits of detection (LOD) and quantification (LOQ). Finally, the RTX was determined in urine and blood samples. Results and discussion: Linearity was demonstrated between charge transfer resistance and RTX from 2 to 14 μg mL−1 (r2 of 0.99), with LOD and LOQ of 130 and 400 ng mL−1 respectively. The method showed adequate precision with a RPD of 0.96% for researcher 1 and 1.27% for researcher 2, and accuracy with a RPD of 1.30%, with a confidence interval between 99.0% and 101.0 %. Selectivity was achieved by demonstrating the binding specificity between the antibody and RTX, where casein blocks non-specific binding when the immunosensor is applied to blood and urine samples. Conclusion: The developed immunosensor presented precision and accuracy for the detection of RTX with results in approximately 20 minutes. In this way, low-cost, marker-sensitive electrochemical immunosensors can help in TDM, QC and extended stability monitoring of different drugs, in a simple way and with versatile assays.