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NanoString nCounter Technology

NanoString Technologies

Genomic and Transcriptomic Analysis Without the Use of Enzyme – also from FFPE Tissue

NanoString is a fully automated technology ensuring precise and reproducible quantification of either RNA or DNA, making it compatible with clinical research. The technology is robust across many sample types and qualities, both tissues of poor and varying quality can be used, including FFPE tissue sections.

The nCounter® Analysis System from NanoString Technologies offers single molecule detection with no amplification, using unique fluorescent barcodes. The barcodes enable direct, digital detection of hundreds (≤800) of different target molecules in a single run allowing targeted gene expression profiling, gene fusion and CNV analysis.

NanoString can detect target molecules as small as 100 base pairs, and also even miRNAs, and only requires a limited input amount from 25 ng of RNA. This enables multiplex gene expression profiling with detection sensitivity down to 1 copy per cell.

Digital Quantification of DNA and RNA

Each assay consists of two probes of 50 bp, which are complementary to the target: a capture probe linked to biotin for immobilization and purification, and a reporter probe containing a unique fluorescent barcode.

The technology is reliable, due to the simplicity of the method. Once the target-probe complexes are hybridized, the complexes are captured using the biotin tag on the capture probe on a streptavidin-coated surface, where they are aligned and immobilized for imaging, and excess probe is removed.

A scanner then digitally counts each fluorescent barcode hybridized to an RNA or DNA molecule and put together the counts for a total count of the target in the sample.

Thus, one count equal one molecule. This ensures a very low false positive rate (0.1%) compared to analogue technologies that measure fluorescent intensities.

The data from the digital reads are normalized to internal controls and a set of reference genes. This will generate highly precise relative counts that can be interpreted in terms of relative change across samples. Absolute counts are not calculated using this technology.

Digital gene expression technology using NanoString nCounter

Single molecule barcode technology for digital detection of DNA, RNA and proteins.

Targeted Gene Expression Profiling

Analysis of the transcriptome can be useful as a first line of investigation in explorative studies, like drug target discovery or in search of biomarkers for a specific disease. The transcriptome changes rapidly as response to the external or internal environment in the body, and thus may be able to predict certain conditions.

At Bioxpedia we offer gene expression profiling service, using the NanoString panels. NanoString nCounter is a robust and reproducible technology for targeted gene expression profiling. Multiple panels ranging from app. 200-770 genes have been developed within different research areas, including immunology, oncology and neurology. Most gene expression panels come in both a human and mouse version.

Below are listed some of the panels currently available:

  • Breast Cancer 360
  • PanCancer IO 360
  • CAR-T Characterization
  • Metabolic Pathways
  • Cancer metabolism
  • PanCancer Immune Profiling
  • PanCancer Pathways
  • PanCancer Progression
  • Autoimmune Profiling
  • Autoimmune Discovery
  • Fibrosis
  • DNA Damage and repair
  • Human Organ Transplant
  • Immunology v2
  • Inflammation v2
  • Innate Immunity
  • Cellular signalling
  • Myeloid Innate Immunity
  • Neuroinflammation
  • Neuropathology
  • Alzheimer’s Disease
  • .. more are available!

For information about other panels or of the genes included in the panels, don’t hesitate to ask us. We are also happy to help choosing the right panel for your project. See also NanoStrings website for more panels.

miRNA Expression Profiling

In addition to gene expression panels, miRNA expression panels have also been developed for both human, mouse and rat miRNA profiling. BioXpedia offers sample analysis using NanoStrings miRNA panels.

To be able to detect the small ~22 nucleotide miRNAs, the technology has been enhanced with the use of additional specific oligos. In addition to the capture and reporter probe, a target specific bridge oligo and miRTag is included in the miRNA assays. Thus, the miRNA length is extended with the miRTag by the use of the bridge oligo, and the miRNA:miRTag molecule can then hybridize to the two probes and be counted as usual.

The miRNA expression panels contain 827, 577 and 423 miRNA assays for human, mouse and rat, respectively, and app. 100 ng of total RNA is required for the analysis.

Copy Number Variation and Gene Fusion Assays

Genomic alterations can indeed lead to disruption or modification of specific processes and cause serious diseases like cancer or other genetic disorders.

NanoString has assays for Copy Number Variation analysis and Gene Fusion Assays which are especially suited for FFPE tissue samples. BioXpedia offers sample analysis using the CNV and Gene Fusion panels from NanoString.

Copy Number Variation (CNV) is known as a number of repeated elements in the genome, which can vary in the population and can result in e.g. protein misfolding and inactivity giving rise to disease.

NanoString has developed the panel v2 Cancer CN Assay for detection of copy number variation of 87 genes commonly amplified or deleted in cancer. The input amount for the analysis is 300 ng genomic DNA.

Gene Fusion as the name indicates, refers to the fusion of two separate genes by chromosomal rearrangement resulting in one hybrid gene. Gene fusions are often found to be drivers for cancer, and the resulting gene is referred to as an oncogene. NanoString has developed two gene fusion panels:

  • Leukaemia Gene Fusion Panel
  • Lung Gene Fusion Panel

The RNA input for Gene Fusion assays is app. 100 ng of total RNA, but up to 300 ng if the RNA source is FFPE tissue.