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Hoefig, K.P. ; Heissmeyer, V.

Measuring microRNA expression in size-limited FACS-sorted and microdissected samples.

Methods Mol. Biol. 667, 47-63 (2010)
DOI PMC
Open Access Green möglich sobald Postprint bei der ZB eingereicht worden ist.
MicroRNAs (miRNAs) are small noncoding RNAs of an average length of 22 nucleotides, which repress translation of a large number of target mRNAs. The particular importance of this group of small RNAs arises from the ever growing evidence that they control many biological processes, such as differentiation, proliferation, and apoptosis and that deregulation of individual miRNAs frequently results in cancer. The expression of miRNAs is spatially and temporarily fine-tuned and expression levels can reach more than 50,000 copies of one miRNA within a single cell. It is well documented that the comparison of miRNA signatures of normal and diseased tissues results in a small number of differentially expressed miRNAs, which are consequently of high diagnostic value. However, measuring miRNA expression can easily produce false-positive results, due to the high sequence similarity of the miRNAs within families and because biologically inactive pre-miRNAs as well as contaminating bystander cells may falsify the signal. The application of a quantitative PCR-based method is described here to specifically and reliably detect miRNA expression levels from as little as 50 cells. Pure cell populations were either derived from fluorescence-activated cell sorting (FACS) or laser capture microdissection (LCM). Importantly, a combination of quantitative PCR and LCM can also be applied to measure miRNA expression of cells obtained from formalin-fixed, paraffin-embedded (FFPE) tissues, thereby giving experimental access to archives with large numbers of routinely collected normal and diseased tissue samples.
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Publikationstyp Artikel: Journalartikel
Dokumenttyp Wissenschaftlicher Artikel
Herausgeber Monticelli, S.*
Schlagwörter Cell Separation, Flow Cytometry, Humans, Lasers, Mice, Mice, Inbred BALB C, MicroRNAs, Microdissection, Tissue Fixation, methods, genetics, metabolism
Sprache englisch
Veröffentlichungsjahr 2010
HGF-Berichtsjahr 2010
ISSN (print) / ISBN 1064-3745
e-ISSN 1940-6029
Konferenztitel MicroRNAs and the Immune System : Methods and Protocols
Zeitschrift Methods in Molecular Biology
Quellenangaben Band: 667, Heft: 1, Seiten: 47-63 Artikelnummer: , Supplement: ,
Verlag Springer
Verlagsort Berlin [u.a.]
Begutachtungsstatus Peer reviewed
Institut(e) Institute of Molecular Immunology (IMI)
Research Unit Molecular Immune Regulation (AMIR)
POF Topic(s)
Forschungsfeld(er)
PSP-Element(e) G-551000-001
DOI 10.1007/978-1-60761-811-9_4
PubMed ID 20827526
Erfassungsdatum 2010-12-31
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