| Registro completo |
| Provedor de dados: |
BJID
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| País: |
Brazil
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| Título: |
Duplex realtime PCR method for Epstein-Barr virus and human DNA quantification: its application for post-transplant lymphoproliferative disorders detection
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| Autores: |
Fellner,María Dolores
Durand,Karina
Rodriguez,Marcelo
Irazu,Lucía
Alonio,Virginia
Picconi,María Alejandra
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| Data: |
2014-06-01
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| Ano: |
2014
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| Palavras-chave: |
EBV
Real-time PCR
Viral load
PTLD
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| 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 < 0.05). The EBV load in PBMC from the groups of BD, Non-Tx, Tx and Tx-PTLD exhibited increasing levels (p < 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.
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| Tipo: |
Info:eu-repo/semantics/article
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| Idioma: |
Inglês
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| Identificador: |
http://www.scielo.br/scielo.php?script=sci_arttext&pid=S1413-86702014000300271
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| Editor: |
Brazilian Society of Infectious Diseases
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| Relação: |
10.1016/j.bjid.2013.07.011
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| Formato: |
text/html
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| Fonte: |
Brazilian Journal of Infectious Diseases v.18 n.3 2014
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| Direitos: |
info:eu-repo/semantics/openAccess
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