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NanoString nCounter

NanoString nCounter Sample Preparation and Submission

NanoString nCounter Service Fees

NanoString nCounter Assay Resources

NanoString nCounter

The Genomics Division operates the NanoString nCounter MAX system to measure or assess nucleic acids.  This system uses an amplification-free technology that measures RNA or DNA content by hybridizing fluorescently color-coded barcode-labelled probes to the target molecules and directly counting the number of probes of each barcode that binds to its target (see illustration below).  An overview of the technology can be found here.

These assays are capable of measuring up to 800 targets in a single reaction and the platform cartridge can accommodate up to 12 samples/run. A notable advantage of the technology is the ability to analyze RNA samples of poor quality (such as FFPE samples), however higher amounts of input material are typically required. A listing of the pre-designed gene expression panels and other NanoString assays can be found here.

Sample Preparation and Submission

  1. Acquire your MAX Cartridge and Assay Code Set from NanoString.
    • Your Assay Code set includes the Reporter Code Set, Capture Code Set, and USB drive with the RLF file.
    • See NanoString Assay Resources below for more information
  2. Download the NanoString nSolver software. Training videos are available on the NanoString website. Please note you won't need this until after you receive your data.
  3. Download and complete the PDF iconNanoString Sample Submission Form.
  4. Contact the Genomics Division to arrange for a date and time to bring the NanoString assay, samples, and completed PDF iconNanoString Sample Submission Form to the lab for processing. Please note this instrument is not available for self-serve, we will run the assay.
  5. Prepare your samples
    • For expression profiling, miRNA and miRGE assays, please provide 20 μl of 75 - 350 ng of RNA as measured by a spectrophotometer (e.g., NanoDrop) or fluorometer (e.g., Qubit) in a 0.5 ul microfuge tube suspended in the purification kit elution buffer (e.g., RNAse free water, TE, or Tris buffer).

      *For the best results, preliminary sample QC of the purified RNA sample quality via a spectrophotometer by measuring absorbance at 230 nm (A230), 260 nm (A260) and 280 nm (A280) should be evaluated by the investigator prior submission. The A260/ A280 ratio can help identify contamination with proteins, whereas the A260/A230 ratio can help identify contamination with organic compounds, such as phenol, and guanidinium salts. NanoString recommends a 260/280 ratio of 1.9 or greater and a 260/230 ratio of 1.8 or greater for optimal results.
       
    • For expression profiling using cell lysates, please provide no less than 15,000 cells in a volume of 5 μl in a 0.5 ml tube. Cell lysates should be prepared in a Guanidinium-based (GITC) lysis buffer such as QiagenTM buffer RLT. Cell concentration should be approximately 2,000 – 10,000 cells/μl. At higher cell/buffer ratios, cell lysis and denaturation is inhibited and the solution too viscous to pipette effectively. Note: Cell lysates are not compatible with the miRNA and miRGE assays.
       
    • For miRNA & miRGE assays, please provide at least 200 μl plasma or serum.  Note: The use of plasma, serum or other biofluids may require optimization.
    • Contact the Genomics Division for sample submission for other applications.
  6. Bring your samples and assay components to the Genomics Division on your reserved date and time.

Genomics Division NanoString Workflow:
After you submit your samples to us, wewill perform the following:

  1. Evaluate QA/QC
    • Evaluate sample integrity using an Agilent Bioanalyzer.
    • Quantify your samples for input normalization across the sample set using the Trinean DropSense
  2. Prepare samples for hybridization
  3. Run hybridization
  4. Load cartridges and scan the saples on the NanoString nCounter Sprint System
  5. Data will be transferred to the USB drive containing the RLF file that was provided as part of the sample submission. The data package will include a compressed folder containing Reporter Code Count (RCC) and Reporter Library File (RLF) files for upload into the NanoString nSolver software for analysis.

NanoString nCounter Service Fees

Application Cost per Sample
Expression Profiling Panels

Standard: $39

Low Input: $59

MicroRNA Panels $44
miRGE $44
Other Applications Contact Genomics Division

 

 

 

 

 

 

 

NanoString Assay Resources:

  • Expression Profiling - mRNA and/or lncRNA:  Use total RNA or cell lysate. Validated modifications for single cell analysis.
  • MicroRNA Profiling – miRNA:  Off the shelf panels are available for human, mouse, rat, and fly. miRNA profiling of >800 miRNA sequences.
  • Expression and miRNA Profiling – miRGE (miRNA, mRNA, lncRNA): Analyze expression levels of both message and micro RNA simultaneously in a single reaction
  • Copy Number Variation – CNV:  Measure DNA copy number in up to 800 regions.
  • Protein/RNA expression - Study complex cellular mechanisms with the simultaneous analysis of human gene expression and protein response.
  • ChIP/String - Measure dsDNA targets in immunoprecipitation enriched chromatin.  Can be used to validate ChIP-Seq.
  • Custom Panels - CodeSets designed to your specifications. Include up to 800 targets of interest and designed to any organism with known sequence. Minimum custom order is 48 reactions.