Laser Capture Microdissection and Isolation of High-Quality RNA from Frozen Endometrial Tissue.

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.

• Authors: Michele Cummings, Georgia Mappa, Nicolas M Orsi
• Journal: Methods in molecular biology (Clifton, N.J.) (Methods Mol Biol) Vol. 1723 Pg. 155-166 ( 2018) ISSN: 1940-6029 [Electronic] United States
• PMID: 29344859 (Publication Type: Journal Article)

Chemical References

• RNA

Topics

• Endometrium (metabolism)
• Female
• Frozen Sections
• Gene Expression Profiling
• Humans
• Laser Capture Microdissection (methods)
• RNA (analysis, genetics, isolation & purification)