Bioluminescence imaging for assessment and normalization in transfected cell arrays.

@article{citeulike:2997404, abstract = {Transfected cell arrays (TCAs) represent a high-throughput technique to correlate gene expression with functional cell responses. Despite advances in TCAs, improvements are needed for the widespread application of this technology. We have developed a TCA that combines a two-plasmid system and dual-bioluminescence imaging to quantitatively normalize for variability in transfection and increase sensitivity. The two-plasmids consist of: (i) normalization plasmid present within each spot, and (ii) functional plasmid that varies between spots, responsible for the functional endpoint of the array. Bioluminescence imaging of dual-luciferase reporters (renilla, firefly luciferase) provides sensitive and quantitative detection of cellular response, with minimal post-transfection processing. The array was applied to quantify estrogen receptor alpha (ERalpha) activity in MCF-7 breast cancer cells. A plasmid containing an ERalpha-regulated promoter directing firefly luciferase expression was mixed with a normalization plasmid, complexed with cationic lipids and deposited into an array. ER induction mimicked results obtained through traditional assays methods, with estrogen inducing luciferase expression 10-fold over the antiestrogen fulvestrant or vehicle. Furthermore, the array captured a dose response to estrogen, demonstrating the sensitivity of bioluminescence quantification. This system provides a tool for basic science research, with potential application for the development of patient specific therapies.}, address = {Department of Interdepartmental Biological Sciences, Northwestern University, Evanston, Illinois.}, author = {Pannier, A. K. and Ariazi, E. A. and Bellis, A. D. and Bengali, Z. and Jordan, V. C. and Shea, L. D. }, citeulike-article-id = {2997404}, doi = {http://dx.doi.org/10.1002/bit.21477}, issn = {1097-0290}, journal = {Biotechnology and bioengineering}, keywords = {estrogen, fluorescence, tma}, month = {October}, number = {2}, pages = {486--497}, posted-at = {2008-07-13 17:32:42}, priority = {2}, title = {Bioluminescence imaging for assessment and normalization in transfected cell arrays.}, url = {http://dx.doi.org/10.1002/bit.21477}, volume = {98}, year = {2007} }

TY - JOUR ID - citeulike:2997404 L3 - citeulike-article-id:2997404 TI - Bioluminescence imaging for assessment and normalization in transfected cell arrays. JF - Biotechnology and bioengineering VL - 98 IS - 2 SP - 486 EP - 497 AD - Department of Interdepartmental Biological Sciences, Northwestern University, Evanston, Illinois. SN - 1097-0290 N2 - Transfected cell arrays (TCAs) represent a high-throughput technique to correlate gene expression with functional cell responses. Despite advances in TCAs, improvements are needed for the widespread application of this technology. We have developed a TCA that combines a two-plasmid system and dual-bioluminescence imaging to quantitatively normalize for variability in transfection and increase sensitivity. The two-plasmids consist of: (i) normalization plasmid present within each spot, and (ii) functional plasmid that varies between spots, responsible for the functional endpoint of the array. Bioluminescence imaging of dual-luciferase reporters (renilla, firefly luciferase) provides sensitive and quantitative detection of cellular response, with minimal post-transfection processing. The array was applied to quantify estrogen receptor alpha (ERalpha) activity in MCF-7 breast cancer cells. A plasmid containing an ERalpha-regulated promoter directing firefly luciferase expression was mixed with a normalization plasmid, complexed with cationic lipids and deposited into an array. ER induction mimicked results obtained through traditional assays methods, with estrogen inducing luciferase expression 10-fold over the antiestrogen fulvestrant or vehicle. Furthermore, the array captured a dose response to estrogen, demonstrating the sensitivity of bioluminescence quantification. This system provides a tool for basic science research, with potential application for the development of patient specific therapies. KW - estrogen KW - fluorescence KW - tma AU - Pannier, AK AU - Ariazi, EA AU - Bellis, AD AU - Bengali, Z AU - Jordan, VC AU - Shea, LD PY - 2007/10/01/ UR - http://dx.doi.org/10.1002/bit.21477 ER -