ABSTRACT
Abstract The quantification of viral nucleic acids in serum by real-time PCR plays an important role in diagnosing hepatitis B virus and hepatitis C virus infection. In this study, we developed an assay using specific primers and probes to quantify hepatitis B virus DNA or hepatitis C virus RNA in serum from infected patients. For standardization and validation of the assay, an international panel of hepatitis B virus/hepatitis C virus and standard plasmids was used. A correlation coefficient of 0.983 and 0.963 for hepatitis B virus and hepatitis C virus, respectively, was obtained based on cycle threshold values and concentrations of DNA or RNA. The standard curve showed a linear relationship from 19 IU/mL to 1.9 × 109 IU/mL of serum, with a coefficient of determination (r2) of 0.99. In sera from patients infected with hepatitis B virus or hepatitis C virus viral loads (19 IU/mL and 1.9 × 109 IU/mL), we quantified viral loads with a detection limit of 1.9 × 102 IU/mL. The real-time quantitative PCR assay developed in this study provides an ideal system for routine diagnosis and confirmation of indeterminate serological results, especially in immunosuppressed patients.
Subject(s)
Humans , Hepatitis B virus/genetics , Hepatitis C/diagnosis , Hepatitis C/virology , Hepacivirus/genetics , Viral Load , Hepatitis B/diagnosis , Hepatitis B/virology , DNA, Viral , RNA, Viral , Reproducibility of Results , Sensitivity and Specificity , Real-Time Polymerase Chain ReactionABSTRACT
A hereditariedade autossômica dominante da neoplasia endócrina múltipla tipo 2 (NEM 2) relaciona-se à ativação do proto-oncogene RET, através de mutações missense. As mutações do RET são encontradas em 95% dos casos índices de NEM 2 e apresentam relação direta entre sua localização codon específica e os diversos fenótipos desenvolvidos, dentre eles, carcinoma medular datireóide, feocromocitoma e/ou hiperparatireoidismo. Baseando-se em análises bioquímicas e genéticas, é possível efetuar um diagnósticoprematuro, viabilizando a intervenção cirúrgica em tempo hábil. A periodicidade da monitorização bioquímica é ditada pelo fenótipopresente, pelas manifestações clínicas familiares e pelo genótipo RET. A recomendação da análise genética deve ser feita a todos indivíduos afetados e também a seus ascendentes e descendentes diretos, caso alguma mutação esteja presente; permitindo identificar os portadores de mutações RET, previamente ao início da sintomatologia. Neste trabalho, serão discutidos os aspectos molecularesdos diversos fenótipos da NEM 2, bem como a importância da identificação genotípica do proto-oncogene RET e sua interação com os testes bioquímicos visando o diagnóstico precoce, prevenção, monitorização, screening familiar e, portanto, maior sobrevidado paciente.
The dominant autossomic hereditarity of the multiple endocrine neoplasia type 2 (MEN 2) is related to RET proto-oncogene activation, through mutations missense. RET mutations are found in 95% of MEN 2 index cases and present direct relation between its specific localization codon and the diverse developed phenotypes, among them, medullary thyroid carcinoma, pheochromocytoma and/or hyperparathyroidism. Being based on biochemists and genetics analyses, it is possible to perform a premature diagnosis, making possible a surgical intervention in the right time. The biochemist monitoring regularity is determined by present phenotype, the familiar clinical manifestations and RET genotype. The recommendation of the genetic analysis must be made to all affected individuals and also their ascendants and descendants, in case some mutation is present, allowing to identify the RET mutations carriers previously to the beginning of the symptomatology. In this work, the molecular aspects of MEN 2 diverse phenotypes will be discussed, as well as the importance of the RET proto-oncogene genotypic identification and its interaction with the biochemists tests aiming the precocious diagnosis, prevention, monitoring, familiar screening e, therefore, the patients longer survival.