Volume 11 Supplement 1

16th Interntional Conference on Human Retroviruses: HTLV and Related Viruses

Open Access

Development and validation of a Multiplex Real-Time PCR for HTLV-1/2 confirmatory diagnosis

  • Maurício Cristiano Rocha Júnior1, 2,
  • Rodrigo Haddad3,
  • Virginia Mara de Deus Wagatsuma1,
  • Oswaldo Massaiti Takayanagui4,
  • Svetoslav Nanev Slavov1,
  • Katia Kaori Otaguiri1,
  • Evandra Strazza Rodrigues1, 2,
  • Dimas Tadeu Covas1, 4 and
  • Simone Kashima1, 2
Retrovirology201411(Suppl 1):P105


Published: 7 January 2014

Accurate diagnostic tests are powerful tool to control the spread of the human T-lymphotropic virus (HTLV) infection. In Brazil, the currently applied diagnostic algorithm for HTLV-1/2 screening is based on serological tests (enzyme-linked immunosorbent assay followed by Western Blot). However this algorithm is unsuitable due to its high cost and the elevated rate of Western Blot (WB) indeterminate results. Nevertheless, molecular techniques such as real-time PCR (qPCR) can overlap these drawbacks because of their high sensitivity and specificity. Several studies have described different qPCR protocols for HTLV-1/2 diagnosis but they lack suitable validation. For this reason, we developed and validated a qualitative multiplex qPCR platform for simultaneous detection and discrimination of the HTLV-1/2 infection in a single reaction tube. A total of 73 HTLV-positive samples and 100 samples obtained from non-infected individuals were tested. The detection limit was one copy/reaction for HTLV-1 and 10 copies/reaction for HTLV-2. We observed a high and similar efficiency between the single and multiplex format as well as the cycle treshold and r 2 values. In addition, this molecular platform reached 100% of sensitivity and specificity. In conclusion, the developed method using one tube multiplex qPCR (confirmatory and discriminatory) for HTLV-1/2 was validated. It showed low cost and high sensitivity and specificity compared to previously described assays and to the traditional confirmatory method (WB). Therefore, this platform can be a supportive tool for the current confirmatory methods adopted.

Authors’ Affiliations

Hemocentro de Ribeirão Preto, Universidade de São Paulo (USP)
Faculdade de Ciências Farmacêuticas de Ribeirão Preto - Universidade de São Paulo (USP) Ribeirão Preto
Universidade de Brasília, Faculdade de Ceilândia – UNB/FCE, Brasília
Faculdade de Medicina de Ribeirão Preto - Universidade de São Paulo (USP) Ribeirão Preto


© Rocha Júnior et al; licensee BioMed Central Ltd. 2014

This article is published under license to BioMed Central Ltd. This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated.