mRNA transcript quantification in archival samples using multiplexed, color-coded probes

A recently developed probe-based technology, the NanoString nCounter™ gene expression system, has been shown to allow accurate mRNA transcript quantification using low amounts of total RNA. We assessed the ability of this technology for mRNA expression quantification in archived formalin-fixed, para...

Full description

Saved in:

Bibliographic Details
Published in:	BMC biotechnology 2011-05, Vol.11 (1), p.46-46, Article 46
Main Authors:	Reis, Patricia P, Waldron, Levi, Goswami, Rashmi S, Xu, Wei, Xuan, Yali, Perez-Ordonez, Bayardo, Gullane, Patrick, Irish, Jonathan, Jurisica, Igor, Kamel-Reid, Suzanne
Format:	Article
Language:	eng
Subjects:	Carcinoma, Squamous Cell - genetics Carcinoma, Squamous Cell - metabolism Color Fluorescent Dyes - chemistry Gene expression Gene Expression Profiling Humans Messenger RNA Methodology Molecular Probe Techniques Mouth Neoplasms - genetics Mouth Neoplasms - metabolism Nanotechnology - methods Nucleic Acid Probes - chemistry Organic Chemicals - chemistry Paraffin Embedding Physiological aspects Polymerase chain reaction Reverse Transcriptase Polymerase Chain Reaction - methods RNA, Messenger - analysis
Online Access:	Get full text
Tags:	Add Tag No Tags, Be the first to tag this record!

Description
Summary:	A recently developed probe-based technology, the NanoString nCounter™ gene expression system, has been shown to allow accurate mRNA transcript quantification using low amounts of total RNA. We assessed the ability of this technology for mRNA expression quantification in archived formalin-fixed, paraffin-embedded (FFPE) oral carcinoma samples. We measured the mRNA transcript abundance of 20 genes (COL3A1, COL4A1, COL5A1, COL5A2, CTHRC1, CXCL1, CXCL13, MMP1, P4HA2, PDPN, PLOD2, POSTN, SDHA, SERPINE1, SERPINE2, SERPINH1, THBS2, TNC, GAPDH, RPS18) in 38 samples (19 paired fresh-frozen and FFPE oral carcinoma tissues, archived from 1997-2008) by both NanoString and SYBR Green I fluorescent dye-based quantitative real-time PCR (RQ-PCR). We compared gene expression data obtained by NanoString vs. RQ-PCR in both fresh-frozen and FFPE samples. Fresh-frozen samples showed a good overall Pearson correlation of 0.78, and FFPE samples showed a lower overall correlation coefficient of 0.59, which is likely due to sample quality. We found a higher correlation coefficient between fresh-frozen and FFPE samples analyzed by NanoString (r = 0.90) compared to fresh-frozen and FFPE samples analyzed by RQ-PCR (r = 0.50). In addition, NanoString data showed a higher mean correlation (r = 0.94) between individual fresh-frozen and FFPE sample pairs compared to RQ-PCR (r = 0.53). Based on our results, we conclude that both technologies are useful for gene expression quantification in fresh-frozen or FFPE tissues; however, the probe-based NanoString method achieved superior gene expression quantification results when compared to RQ-PCR in archived FFPE samples. We believe that this newly developed technique is optimal for large-scale validation studies using total RNA isolated from archived, FFPE samples.
ISSN:	1472-6750 1472-6750