Uso de una reacción de polimerasa en cadena simple combinada con un ensayo de movilidad de heteroduplexes en la caracterización de la región 5ï no codificante del virus de la hepatitis C / Use of a simple polymerase chain reaction combined with a heteroduplex mobility assay to characterize the non coding 5ï region of hepatitis C virus

RESUMO

Background: One of the most used methods for the characterization of hepatitis C virus strains is the use of a nested polymerase chain reaction (PCR) with a restriction fragment length polymorphism (RFLP) assay. Sometimes, RFLP results do not differentiate new strains. There are other more complex methods and only the sequencing of the PCR fragment allows a correct characterization of the strain. Aim: To report the detection of hepatitis C virus using a single PCR assay of the 5' non codifying region. Material and methods: Thirty five serum samples coming from patients with chronic hepatitis or blood donors were assayed for hepatitis C virus. Results: The reported method increases the PCR sensitivity through the combination of polyacrylamide gel electrophoresis and silver staining of amplified products. This allowed the semi quantitative estimation of viral load and the characterization of amplified products through their electrophoretic motility. These PCR products were used in a heteroduplex motility assay, allowing the discrimination between sequences of different genotypes. Conclusions: Heteroduplex assays can be used to characterize the 5' non codifying region of the hepatitis C virus for routine laboratory purposes: