Abstract：
INTRODUCTION:Affymetrix oligonucleotide microarrays are widely used in basic and applied research (Lander, E.S., (1999). Array of hope. Nature Genetics 21, 3-4; Lockhart, D.J. & Winzeler, E.A. (2000) Genomics, gene expression and DNA arrays. Nature 405, 827-836.) The need for a significant amount of starting RNA has limited its use in applications where the amount of RNA is limiting, such as with Laser Captured Microdissection (LCM), small biopsies, or peripheral blood in rodent models. To overcome this limitation, various RNA amplification and labeling methods have been described, however, further optimization and validation of these methods are needed. METHODS:Here we reported using the Arcturus technology to optimize amplification and labeling of small amounts of RNA for Affymetrix microarray studies. We assessed the technical feasibility and variation introduced by differences in starting RNA quantity and differences in technical performance by microarray hybridization. RESULTS:We demonstrated that the current approach is reliable to amplify as little as 40 ng total RNA, and it is suitable for Affymetrix studies yielding satisfactory quantitative chip performance. We also showed that differences in labeling methods contribute more to variation than the differences in starting RNA quantity per se. As a model, we studied the well-documented TNF-induced inflammatory responses in cultured human vascular endothelial cells. We were able to recapitulate the TNF-induced responses using small RNA sample profiling.
journal_name
J Pharmacol Toxicol Methodsjournal_title
Journal of pharmacological and toxicological methodsauthors
Shou J,Qian HR,Lin X,Stewart T,Onyia JE,Gelbert LMdoi
10.1016/j.vascn.2005.02.004keywords:
subject
Has Abstractpub_date
2006-03-01 00:00:00pages
152-9issue
2eissn
1056-8719issn
1873-488Xpii
S1056-8719(05)00021-3journal_volume
53pub_type
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journal_title：Journal of pharmacological and toxicological methods
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doi：10.1016/j.vascn.2006.02.009
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doi：10.1016/j.vascn.2009.03.005
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journal_title：Journal of pharmacological and toxicological methods
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journal_title：Journal of pharmacological and toxicological methods
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journal_title：Journal of pharmacological and toxicological methods
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journal_title：Journal of pharmacological and toxicological methods
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