Cummings, M, Mappa, G and Orsi, N (2018) Laser Capture Microdissection and Isolation of High-Quality RNA from Frozen Endometrial Tissue. Methods of Molecular Biology, 1723. pp. 155-166. ISSN 1064-3745
Abstract
Laser capture microdissection (LCM) allows expression profiling of specific cell populations within tissues. However, isolation of high-quality RNA from laser capture microdissected frozen tissue is beset by problems arising from intrinsic tissue RNase activity. Herein, we describe an optimized staining/LCM/RNA extraction protocol developed for the isolation of epithelial RNA from frozen tissue sections using human endometrial cancer as a model tissue. This method combines excellent, reproducible visualization of tissue morphology with the isolation of high-integrity RNA suitable for downstream applications such as expression microarray analysis. We present quantitative and qualitative RNA data obtained from >200 endometrial epithelial samples (normal, hyperplastic, and cancerous), where 92% of samples had RIN values of 7 and above and highlight common pitfalls faced by investigators. This method should also be broadly applicable to a range of other tissue types.
Metadata
Item Type: | Article |
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Authors/Creators: |
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Copyright, Publisher and Additional Information: | © Springer Science+Business Media, LLC 2018. This is an author produced version of a paper published in Methods in Molecular Biology. The final publication is available at Springer via https://doi.org/10.1007/978-1-4939-7558-7_8. Uploaded in accordance with the publisher's self-archiving policy. |
Keywords: | Frozen tissue; Laser capture microdissection; Cresyl violet; RNA integrity; RIN; Endometrium; Epithelium |
Dates: |
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Institution: | The University of Leeds |
Academic Units: | The University of Leeds > Faculty of Medicine and Health (Leeds) > School of Medicine (Leeds) > Leeds Institute of Cancer and Pathology (LICAP) > Pathology & Tumour Biology (Leeds) |
Depositing User: | Symplectic Publications |
Date Deposited: | 30 Jan 2018 10:47 |
Last Modified: | 18 Jan 2019 01:38 |
Status: | Published |
Publisher: | Humana Press |
Identification Number: | 10.1007/978-1-4939-7558-7_8 |
Open Archives Initiative ID (OAI ID): | oai:eprints.whiterose.ac.uk:126713 |