期刊
JOURNAL OF IMMUNOLOGICAL METHODS
卷 448, 期 -, 页码 9-20出版社
ELSEVIER SCIENCE BV
DOI: 10.1016/j.jim.2017.05.002
关键词
Heparin; Ecteola-cellulose; PRNT; Vaccinology; Yellow fever virus
资金
- Fundacao de Amparo a Pesquisa do Estado de Minas Gerais (FAPEMIG) [2016]
- Conselho Nacional de Desenvolvimento Cientifico e Tecnologico (CNPq) [2015]
- Programa Nacional de Imunizacoes (PNI) [277/2013]
- Secretaria de Vigilancia em Saude (SVS) [2013-2017]
- Bio-Manguinhos/FIOCRUZ [016/2013]
- PROEP/CPqRR/FIOCRUZ [401998/20126]
- PDTIS [2013-2017]
- CAPES [2013-2107]
- Conselho Nacional de Desenvolvimento Cientifico e Tecnologico-CNPq-Brasil
Technological innovations in vaccinology have recently contributed to bring about novel insights for the vaccine induced immune response. While the current protocols that use peripheral blood samples may provide abundant data, a range of distinct components of whole blood samples are required and the different anticoagulant systems employed may impair some properties of the biological sample and interfere with functional assays. Although the interference of heparin in functional assays for viral neutralizing antibodies such as the functional plaque reduction neutralization test (PRNT), considered the gold-standard method to assess and monitor the protective immunity induced by the Yellow fever virus (YFV) vaccine, has been well characterized, the development of pre analytical treatments is still required for the establishment of optimized protocols. The present study intended to optimize and evaluate the performance of pre-analytical treatment of heparin-collected blood samples with ecteolacellulose (ECT) to provide accurate measurement of anti-YFV neutralizing antibodies, by PRNT. The study was designed in three steps, including: I. Problem statement; II. Pre-analytical steps; III. Analytical steps. Data confirmed the interference of heparin on PRNT reactivity in a dose-responsive fashion. Distinct sets of conditions for ECT pre-treatment were tested to optimize the heparin removal. The optimized protocol was pre-validated to determine the effectiveness of heparin plasma:ECT treatment to restore the PRNT titers as compared to serum samples. The validation and comparative performance was carried out by using a large range of serum vs heparin plasma:ECT 1:2 paired samples obtained from unvaccinated and 17DD-YFV primary vaccinated subjects. Altogether, the findings support the use of heparin plasma:ECT samples for accurate measurement of anti-YFV neutralizing antibodies. (C) 2017 Elsevier B.V. All rights reserved.
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