University of Oulu

Myllymäki, H., Niskanen, M., Oksanen, K., Rämet, M. Immunization of Adult Zebrafish for the Preclinical Screening of DNA-based Vaccines. J. Vis. Exp. (140), e58453, doi:10.3791/58453 (2018).

Immunization of adult zebrafish for the preclinical screening of DNA-based vaccines

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Author: Myllymäki, Henna1; Niskanen, Mirja1; Oksanen, Kaisa1,2;
Organizations: 1BioMediTech Institute, Faculty of Medicine and Life Sciences, University of Tampere
2MedEngine Oy
3Department of Pediatrics, Tampere University Hospital
4Department of Children and Adolescents, Oulu University Hospital
5PEDEGO Research Unit, Medical Research Center, University of Oulu
Format: article
Version: published version
Access: open
Online Access: PDF Full Text (PDF, 1 MB)
Persistent link:
Language: English
Published: MyJoVE Corporation, 2018
Publish Date: 2020-03-26


The interest in DNA-based vaccination has increased during the past two decades. DNA vaccination is based on the cloning of a sequence of a selected antigen or a combination of antigens into a plasmid, which enables a tailor-made and safe design. The administration of DNA vaccines into host cells leads to the expression of antigens that stimulate both humoral and cell-mediated immune responses. This report describes a protocol for the cloning of antigen sequences into the pCMV-EGFP plasmid, the immunization of adult zebrafish with the vaccine candidates by intramuscular microinjection, and the subsequent electroporation to improve intake. The vaccine antigens are expressed as green fluorescent protein (GFP)-fusion proteins, which allows the confirmation of the antigen expression under UV light from live fish and the quantification of expression levels of the fusion protein with ELISA, as well as their detection with a western blot analysis. The protective effect of the vaccine candidates is tested by infecting the fish with Mycobacterium marinum five weeks postvaccination, followed by the quantification of the bacteria with qPCR four weeks later. Compared to mammalian preclinical screening models, this method provides a cost-effective method for the preliminary screening of novel DNA-based vaccine candidates against a mycobacterial infection. The method can be further applied to screening DNA-based vaccines against various bacterial and viral diseases.

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Series: Journal of visualized experiments
ISSN: 1940-087X
ISSN-E: 1940-087X
ISSN-L: 1940-087X
Volume: 140
Article number: e58453
DOI: 10.3791/58453
Type of Publication: A1 Journal article – refereed
Field of Science: 1182 Biochemistry, cell and molecular biology
Funding: This work was supported by Jane ja Aatos Erkon Säätiö (Jane and Aatos Erkko Foundation; to M.R.), Sigrid Juséliuksen Säätiö (Sigrid Juselius Foundation; to M.R.), the Competitive State Research Financing of the Expert Responsibility area of Tampere University Hospital (to M.R.), Tampereen Tuberkuloosisäätiö (Tampere Tuberculosis Foundation; to M.R., H.M., and M.N.), Suomen Kulttuurirahasto (Finnish Cultural Foundation; to H.M.), Suomen Tuberkuloosin Vastustamisyhdistyksen Säätiö (Finnish Anti-Tuberculosis Foundation; to H.M.), Väinö ja Laina Kiven säätiö (Väinö and Laina Kivi Foundation; to M.N.) and the Tampere City Science Foundation (to M.N.).
Copyright information: © 2018 Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License.