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August 01, 2009 (Vol. 29, No. 14)

High-Throughput Nanovolume qPCR

Next-Generation Platform Combines Microarray Throughput with Sensitivity

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    Advantages of SmartChip real-time PCR system for expression analysis

    Gene-expression profiling is an integral part of biomarker discovery as well as discovery and validation of new therapeutic targets to support drug development. Initially, profiling the intracellular range of transcript abundance relied on hybridization-based technology, such as microarrays or spotted cDNA arrays, or sequencing-based technology such as SAGE. Due to inherent technical limitations, neither method provides an accurate quantification of the broad range of transcript levels present in a given sample. 

    This is particularly true in the case of low-abundance transcripts due to the fact that signal-to-noise ratios decrease exponentially with the decrease of transcript number. In contrast, the signal-to-noise ratio is constant in real-time PCR facilitating the accurate measurement of low-abundance transcripts. Typically, large changes in gene expression can be underestimated or entirely missed by hybridization-based technologies.

    Quantitative PCR (qPCR)-based gene-expression profiling is the current gold standard for the precise monitoring of a selected set of genes to validate results of a hybridization-based microarray experiment. However, the overall limitations of qPCR and hybridization-based technologies have created a need for a next-generation platform capable of combining the advantages of these two technologies, while addressing their shortcomings, in a single system.

    WaferGen Biosystems has developed a platform for rapid quantitative gene-expression analysis. This technology, referred to as the SmartChip™ Real-Time PCR System, is equipped with a high-density, rapid-cycling configuration capable of providing high-throughput levels, while offering discovery and validation capabilities in a single step. 

    The platform performs thousands of nanoscale quantitative PCR assays in a single run, essentially combining the screening capability of microarrays with the sensitivity of qPCR. This enables life scientists to identify a broad spectrum of expressed genes in a single day, while still discriminating small changes in expression. The SmartChips can be configured to analyze large numbers of genes on a single sample, or interrogate many targeted sets of genetic markers on many samples at once. 

    Whole-genome assay time may be significantly reduced with the SmartChip system as compared to several days or months with microarrays or real-time PCR, respectively. Additionally, it is estimated that this platform will generate expression results 48 times faster and at 50% less cost than conventional methods.

    The SmartChip system also incorporates a number of ease-of-use features including content-ready, high-density chips containing 5,000–30,000 nano-wells with gene panels optimized for cancer, toxicology, and whole genome. The platform requires a small sample size as compared to other technologies and offers real-time detection and sophisticated read-out options while assuring detection sensitivity and temperature uniformity across chips.

    The SmartChip platform consists of content-ready SmartChips, a sample dispenser, a real-time PCR instrument, and analysis software. To date, 5,184-well and primer loaded SmartChips have been fabricated and used to measure ~1,000 genes in total RNA samples derived from matched normal and lung tumor tissue (Table).

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