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Therefore, the sole variables were transgene chromosomal insertion site and copy number. We then adapted and optimized a technique that tests, at the single cell level, persistence of stem cell\u2010mediated transgene expression in vivo based on correlating the presence of the transgene in a given NSC's nucleus (by fluorescence in situ hybridization [FISH]) with the frequency of that transgene's product within the same cell (by combined immunohistochemistry [IHC]). Under the above\u2010stated conditions, insertion site is likely the most contributory variable dictating transgene downregulation in an NSC after 3 months in vivo. We also observed that this obstacle could be effectively and safely counteracted by simple serial infections (as few as three) inserting redundant copies of the transgene into the prospective donor NSC. (The preservation of normal growth control mechanisms and an absence of tumorigenic potential can be readily screened and ensured ex vivo prior to transplantation.) The combined FISH\/IHC strategy employed here for monitoring the dynamics of transgene expression at the single cell level in vivo may be used for other types of therapeutic and housekeeping genes in endogenous and exogenous stem cells of many organs and lineages. J. Comp. Neurol. 515:83\u201392, 2009. \u00a9 2009 Wiley\u2010Liss, Inc.<\/jats:p>","DOI":"10.1002\/cne.21957","type":"journal-article","created":{"date-parts":[[2009,3,20]],"date-time":"2009-03-20T17:01:02Z","timestamp":1237568462000},"page":"83-92","source":"Crossref","is-referenced-by-count":8,"title":["The dynamics of long\u2010term transgene expression in engrafted neural stem cells"],"prefix":"10.1002","volume":"515","author":[{"given":"Jean\u2010Pyo","family":"Lee","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"David J.","family":"Tsai","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Kook","family":"In Park","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Alan R.","family":"Harvey","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Evan 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