Methods in Molecular Biology Volume 411, 2007, pp 25-34

Detection of Reporter Gene Expression in Murine Airways

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We have shown that to overcome the low levels of expression from gene transfer vector-mediated β-galactosidase expression in lung, it is essential to replace the cytoplasmic β-galactosidase gene with a nuclear targeted β-galactosidase gene. We found that lung should be sectioned and fixed prior to staining for β-galactosidase expression and that en bloc staining of intact lung is inefficient at staining positively transduced cells located deeper in the lung spaces. For GFP fluorescence, it is important to inflate the lungs with fixative prior to freezing and subsequent sectioning. For processing of nasal tissues for β-galactosidase expression, we expand on a protocol used in previously reported gene transfer studies.