Use of a murine secreted alkaline phosphatase as a non-immunogenic reporter gene in mice.

Autor: Maelandsmo GM; Department of Tumor Biology, Norwegian Radium Hospital, 0310 Oslo, Norway., Ross PJ, Pavliv M, Meulenbroek RA, Evelegh C, Muruve DA, Graham FL, Parks RJ
Jazyk: angličtina
Zdroj: The journal of gene medicine [J Gene Med] 2005 Mar; Vol. 7 (3), pp. 307-15.
DOI: 10.1002/jgm.666
Abstrakt: Background: The development of any vector system as a gene delivery system requires its optimization in vitro and in vivo. Preliminary studies frequently involve the use of a reporter gene, which allows for the rapid and simple assay of vector function through monitoring expression levels of the reporter gene. However, evaluation of vector efficacy can be compromised by immune responses directed against immunogenic reporter proteins.
Methods: We have cloned a murine secreted alkaline phosphatase (mSEAP), and explored its use as a reporter gene in the context of an early region 1 (E1)-deleted adenovirus (Ad) vector. Studies involved characterization of gene expression in vitro and in vivo, and immunological responses after gene delivery to mice.
Results: In tissue culture, we show that mSEAP is easily measured quantitatively using a sensitive, commercially available chemiluminescent assay, or visualized directly using histological staining. The level of transgene expression from AdmSEAP was similar to that observed for an Ad vector encoding the human placental secreted alkaline phosphatase (hSEAP). After intravenous administration in mice, AdmSEAP continued to express at high levels for the duration of the experiment (1 month), whereas expression from AdhSEAP declined to background levels over the course of the experiment. Although cytotoxic T-lymphocytes were not detected against either the murine or human SEAP proteins in mice, antibodies were readily detected against the human protein. No antibodies were detected to mSEAP.
Conclusions: Taken together, these data illustrate that mSEAP is a sensitive, non-immunogenic reporter gene for preclinical mouse studies.
(Copyright (c) 2004 John Wiley & Sons, Ltd.)
Databáze: MEDLINE
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