title: Systematic evaluation of medium-throughput mRNA abundance platforms.
abstract: Profiling of mRNA abundances with high-throughput platforms such as microarrays and RNA-seq has become an important tool in both basic and biomedical research. However, these platforms remain prone to systematic errors and have challenges in clinical and industrial applications. As a result, it is standard practice to validate a subset of key results using alternate technologies. Similarly, clinical and industrial applications typically involve transitions from a high-throughput discovery platform to medium-throughput validation ones. These medium-throughput validation platforms have high technical reproducibility and reduced sample input needs, and low sensitivity to sample quality (e.g., for processing FFPE specimens). Unfortunately, while medium-throughput platforms have proliferated, there are no comprehensive comparisons of them. Here we fill that gap by comparing two key medium-throughput platforms--NanoString's nCounter Analysis System and ABI's OpenArray System--to gold-standard quantitative real-time RT-PCR. We quantified 38 genes and positive and negative controls in 165 samples. Signal:noise ratios, correlations, dynamic range, and detection accuracy were compared across platforms. All three measurement technologies showed good concordance, but with divergent price/time/sensitivity trade-offs. This study provides the first detailed comparison of medium-throughput RNA quantification platforms and provides a template and a standard data set for the evaluation of additional technologies.
authors.affiliation: Informatics and Biocomputing Platform, Ontario Institute for Cancer Research, Toronto, Ontario M5G 0A3, Canada.
authors.forename: Stephenie D
authors.initials: SD
authors.lastname: Prokopec
authors.forename: John D
authors.initials: JD
authors.lastname: Watson
authors.forename: Daryl M
authors.initials: DM
authors.lastname: Waggott
authors.forename: Ashley B
authors.initials: AB
authors.lastname: Smith
authors.forename: Alexander H
authors.initials: AH
authors.lastname: Wu
authors.forename: Allan B
authors.initials: AB
authors.lastname: Okey
authors.forename: Raimo
authors.initials: R
authors.lastname: Pohjanvirta
authors.forename: Paul C
authors.initials: PC
authors.lastname: Boutros
mesh_terms: D000818:Animals; D020869:Gene Expression Profiling / Q000379:methods; D059014:High-Throughput Nucleotide Sequencing / Q000191:economics / Q000295:instrumentation / Q000379:methods*; D008297:Male; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods; D000072317:Polychlorinated Dibenzodioxins / Q000633:toxicity; D012333:RNA, Messenger / Q000032:analysis* / Q000187:drug effects; D051381:Rats; D020318:Rats, Long-Evans; D017208:Rats, Wistar; D060888:Real-Time Polymerase Chain Reaction / Q000191:economics / Q000295:instrumentation / Q000379:methods*; D015203:Reproducibility of Results; D059629:Signal-To-Noise Ratio
chemical_list: D000072317:Polychlorinated Dibenzodioxins; D012333:RNA, Messenger