Data/hora: 01/06/2024 01:30:11

Provedor de dados: | 55
País: | Brazil
Título: | Duplex realtime PCR method for Epstein-Barr virus and human DNA quantification: its application for post-transplant lymphoproliferative disorders detection
Autores: | Fellner,María Dolores; Durand,Karina; Rodriguez,Marcelo; Irazu,Lucía; Alonio,Virginia; Picconi,María Alejandra.
Data: | 2014-06-01
Ano: | 2014
Palavras-chave: | EBV; Real-time PCR; Viral load; PTLD.
Resumo: | INTRODUCTION: The quantification of circulating Epstein-Barr virus (EBV) DNA is used to monitor transplant patients as an early marker of Post-Transplant Lymphoproliferative Disorders (PTLD). So far no standardized methodology exists for such determination.OBJECTIVE: Our purpose was to develop and validate a real-time PCR assay to quantify EBV DNA in clinical samples from transplant recipients.METHODS: A duplex real-time PCR method was developed to amplify DNA from EBV and from a human gene. The EBV load was determined in peripheral blood mononuclear cells (PBMC), plasma and oropharyngeal tissue from 64 non-transplanted patients with lymphoid-hypertrophy (Non-Tx), 47 transplant recipients without PTLD (Tx), 54 recipients with PTLD (Tx-PTLD), and 66 blood donors (BD). WinPEPI, version 11.14 software was used for statistical analysis.RESULTS:Analytical validation: the intra and inter-assays variation coefficients were less than 4.5% (EBV-reaction) and 3% (glyceraldehyde 3-phosphate dehydrogenase - GAPDH reaction). Linear ranges comprised 107-10 EBV genome equivalents (gEq) (EBV-reaction) and 500,000-32 human gEq (GAPDH-reaction). The detection limit was 2.9 EBV gEq (EBV-reaction). Both reactions showed specificity. Application to clinical samples: higher levels of EBV were found in oropharyngeal tissue from transplanted groups with and without PTLD, compared to Non-Tx (p &lt; 0.05). The EBV load in PBMC from the groups of BD, Non-Tx, Tx and Tx-PTLD exhibited increasing levels (p &lt; 0.05). In BD, PBMC and plasma, EBV loads were undetectable.CONCLUSIONS: The performance of the assay was suitable for the required clinical application. The assay may be useful to monitor EBV infection in transplant patients, in particular in laboratories from low-income regions that cannot afford to use commercial assays.
Tipo: | Info:eu-repo/semantics/article
Idioma: | Inglês
Identificador: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1413-86702014000300271
Editor: | Brazilian Society of Infectious Diseases
Relação: | 10.1016/j.bjid.2013.07.011
Formato: | text/html
Fonte: | Brazilian Journal of Infectious Diseases  v.18 n.3 2014
Fonte: | info:eu-repo/semantics/openAccess