In vivo assessment of specific cytotoxic T lymphocyte killing

In vivo assessment of specific cytotoxic T lymphocyte killing

Author Clemente, T. Google Scholar
Dominguez, M. R. Autor UNIFESP Google Scholar
Vieira, N. J. Google Scholar
Rodrigues, M. M. Autor UNIFESP Google Scholar
Amarante-Mendes, G. P. Google Scholar
Institution Universidade de São Paulo (USP)
Universidade Federal de São Paulo (UNIFESP)
Abstract The direct killing of target cells by cytotoxic T lymphocytes (CTLs) plays a fundamental role in protective immunity to viral, bacterial, protozoan and fungi infections, as well as to tumor cells. in vivo cytotoxic assays take into account the interaction of target and effector cells in the context of the proper microenvironment making the analysis biologically more relevant than in vitro cytotoxic assays. Thus, the development, improvement and validation of in vivo methods are necessary in view of the importance of the results they may provide. We describe and discuss in this manuscript a method to evaluate in vivo specific cytotoxic T lymphocyte killing. We used as model system mice immunized with human recombinant replication-deficient adenovirus 5 (HAd5) containing different transgenes as the trigger of a CTL-mediated immune response. To these mice, we adoptively transferred syngeneic cells labeled with different vital fluorescent dyes. Donor cells were pulsed (target) or not (control non-target) with distinct CD8 T-cell epitopes, mixed in a 1:1 ratio and injected iv. into immunized or non-immunized recipient mice. After 18-24 h, spleen cells are collected and analysed by flow cytometry. A deviation from the 1:1 ratio of control and target cell populations indicates antigen specific lysis of target cells. (C) 2013 Elsevier Inc. All rights reserved.
Keywords In vivo cytotoxic assay
Human recombinant replication-deficient
adenovirus 5
Adeno beta gal
Language English
Date 2013-06-01
Published in Methods. San Diego: Academic Press Inc Elsevier Science, v. 61, n. 2, p. 105-109, 2013.
ISSN 1046-2023 (Sherpa/Romeo, impact factor)
Publisher Elsevier B.V.
Extent 105-109
Access rights Closed access
Type Article
Web of Science ID WOS:000321234700004

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