Desenvolvimento de imunossensor para detecção de hemorragia feto – materna (HFM)
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Data
2019-02-27
Autores
Nishio, Suelen de Souza Assunpção
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Universidade Estadual Paulista (Unesp)
Resumo
A Hemorragia feto-materna (HFM) se dá pela transferência do sangue fetal para o compartimento intravascular materno, devido à ruptura na membrana vásculo-sincicial da placenta. A HFM pode ser responsável pela alosensibilização do sistema imune materno, levando a morbidade e mortalidade de gravidez corrente e/ou de futura, assim como também constitui a base da etiopatogenia de várias afecções, como se verifica na doença hemolítica perinatal que expõe a complicações fetais como, hidropsia, danos cerebrais hipóxicos e morte fetal. Detectar e quantificar hemácias fetais ajuda a prevenir as consequências devida a ocorrência da HFM. Entre os testes diagnósticos utilizados na detecção e quantificação têm - se os mais sensíveis o teste quantitativo de Kleihauer e Betke e o de Citometria de fluxo, sendo o primeiro considerado padrão ouro. Tendo em vista a importância do diagnóstico laboratorial da HFM foi desenvolvida a proposta da construção de imunossensor para detecção de amostras de sangue fetal na corrente sanguínea materna. Foi realizada a produção de anticorpo monoclonal com objetivo de produzir anticorpos contra antígenos de superfícies de hemácias fetais humana e também foram realizadas a construção de unidades sensoriais para detecção de sinal biológico de imunoafinidade entre anticorpos que detectam hemácias fetais e a hemoglobina fetal presente nestas células, a fim de se diagnosticar a ocorrência da HFM. Foram obtidos como resultados a produção de anticorpo monoclonal que detecta hemácias humanas tanto adulta quanto fetal e a construção de imunossensores que detectam reações de imunoafinidade utilizando anticorpos monoclonais comerciais purificados anti-CD71 e anti-HbF adsorvido a matrizes poliméricas em superfícies de eletrodo impresso de carbono e eletrodo interdigitado de ouro pela técnica Layer-by-layer (LbL). Os sistemas sensoriais montados foram analisados através de medidas eletroquímicas e de impedância elétrica, sendo obtida uma melhor resposta por medida de impedância elétrica, portanto obteve-se resultados satisfatórios em eletrodos interdigitado de ouro.
Fetomaternal hemorrhage (FMH) occurs through the transfer of fetal blood into the maternal intravascular compartment, due to rupture in the placenta-syncytial membrane. FMH may be responsible for the alosensitization of the maternal immune system, leading to morbidity and mortality of current and / or future pregnancies, as well as being the basis of the etiopathogenesis of various conditions, as in perinatal haemolytic disease that exposes to fetal complications such as hydrops, hypoxic brain damage, and fetal death. Detecting and quantifying fetal erythrocytes helps to prevent the consequences due to the occurrence of FMH. Among the diagnostic tests used in the detection and quantification are the most sensitive the quantitative test of Kleihauer and Betke and the one of Flow cytometry, being the first considered gold standard. Considering the importance of the laboratory diagnosis of FMH, the proposal of the construction of immunosensor for the detection of fetal blood samples in the maternal blood was developed. The production of a monoclonal antibody was carried out with the objective of producing antibodies against antigens on human fetal red blood cells and also the construction of sensorial units for the detection of biological signal of immunoaffinity between antibodies that detect fetal red blood cells and the fetal hemoglobin present in these cells, in order to diagnose the occurrence of FMH. The results obtained are the production of monoclonal antibody that detects both adult and fetal human erythrocytes and the construction of immunoassay detectors that detect immunoaffinity reactions using purified commercial anti-CD71 and anti-HbF monoclonal antibodies adsorbed to polymer matrices on carbon printed electrode surfaces and interdigitated gold electrode by the Layer-by-layer (LbL) technique. The sensory systems assembled were analyzed by means of electrochemical and electrical impedance measurements, and a better response was obtained by electrical impedance measurement, so satisfactory results were obtained on interdigitated gold electrodes
Fetomaternal hemorrhage (FMH) occurs through the transfer of fetal blood into the maternal intravascular compartment, due to rupture in the placenta-syncytial membrane. FMH may be responsible for the alosensitization of the maternal immune system, leading to morbidity and mortality of current and / or future pregnancies, as well as being the basis of the etiopathogenesis of various conditions, as in perinatal haemolytic disease that exposes to fetal complications such as hydrops, hypoxic brain damage, and fetal death. Detecting and quantifying fetal erythrocytes helps to prevent the consequences due to the occurrence of FMH. Among the diagnostic tests used in the detection and quantification are the most sensitive the quantitative test of Kleihauer and Betke and the one of Flow cytometry, being the first considered gold standard. Considering the importance of the laboratory diagnosis of FMH, the proposal of the construction of immunosensor for the detection of fetal blood samples in the maternal blood was developed. The production of a monoclonal antibody was carried out with the objective of producing antibodies against antigens on human fetal red blood cells and also the construction of sensorial units for the detection of biological signal of immunoaffinity between antibodies that detect fetal red blood cells and the fetal hemoglobin present in these cells, in order to diagnose the occurrence of FMH. The results obtained are the production of monoclonal antibody that detects both adult and fetal human erythrocytes and the construction of immunoassay detectors that detect immunoaffinity reactions using purified commercial anti-CD71 and anti-HbF monoclonal antibodies adsorbed to polymer matrices on carbon printed electrode surfaces and interdigitated gold electrode by the Layer-by-layer (LbL) technique. The sensory systems assembled were analyzed by means of electrochemical and electrical impedance measurements, and a better response was obtained by electrical impedance measurement, so satisfactory results were obtained on interdigitated gold electrodes
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Palavras-chave
Anticorpo monoclonal, Biossensor, Hemorragia feto – materna, Imunossensor, Monoclonal antibody, Biosensor, Fetomaternal hemorrhage, Immunosensor