Data generation and QC

From assay to actionable data with Olink

Please note that the data analysis steps below will be taken care of for you when you send your samples to one of our dedicated, highly skilled sample analysis service labs, who will provide you with a comprehensive Normalized Protein eXpression (NPX) data/quality control report after your study is completed. If more extensive data analysis services are required, our team of Olink biostatisticians can help you with customized statistical analysis – see our fee-for-service statistical services..

If you intend to run our panels in your own lab, or simply want to know more details, please read on below!

Data generation with Olink® Explore

Please note, details related to the Olink Target platform can be found further down this page – go there directly

NPX is Olink’s relative protein quantification unit on a log2 scale. NPX values from Olink Explore are calculated from the number of matched counts (= i.e. the number of  sequence reads for each specific combination of sample and assay) from the NovaSeq 6000 or NextSeq 2000 run.

Running the assay

Detection of the digital PEA signal and sample analysis for Olink® Explore are performed via NGS, using Illumina’s NovaSeq 6000 or NextSeq 2000 instruments*. These high-throughput systems enable true multiplexing, with simultaneous detection and readout of 384 analytes & controls in 96/384 samples (36 864/147 456 data points) in each lane. The instruments can process four lanes in parallel, generating 147 456/589 824  data points from a single run.

Olink Explore workflow, from immunoassay to NGS readout

 

About sample volumes:

  • Olink Explore technically requires 2.8 µL of plasma or serum to measure 1536 analytes.
  • Another 7 µL are needed to compensate for dead-volumes in the automated liquid handling system
  • Analysis Service requests you actually send 40 µL of each sample in case re-runs are required.

The sample preparation steps take about 24 hours, while the final NGS readout takes less than 12 hours to complete.

Data generation and quality control

Data from the NovaSeq 6000 run is uploaded to the MyData cloud. After this, Olink MyData cloud software processes the data, performs QC-analysis, calculates NPX values and produces a Certificate of Analysis (CoA). The cloud software provides Normalized Protein eXpression (NPX) values for further data analysis. This is an entirely new software specially developed for Olink Explore, although most of the functionality is based on the existing NPX Manager Software that was developed for Olink Target 96 panels.

Olink has developed a built-in QC system using internal controls, enabling full control over the technical performance of assays and samples. Three internal controls are spiked into every sample for each dilution and panel, and are designed to monitor the quality of assay performance, as well as the quality of individual samples:

  1. Immuno control (incubation step)
  2. Extension control (extension and pre-amplification step)
  3. Amplification control (amplification step)

Each sample plate also includes a control strip with the following controls:

  1. Two Sample controls used to estimate the precision (intra- and inter-CVs)
  2. Three Negative controls used to set the background levels for each protein assay and to calculate the limit of detection (LOD)
  3. Three Plate controls used to compensate for potential variation between runs and plates

For more details, see our white paper.

Due to the huge amount of data generated by NovaSeq 6000 instruments, all data storage and analysis is handled by a secure AWS cloud service.

Our Analysis Service labs provide customers with project reports generated using the MyData Cloud software. Examples of the documents you will receive from our Analysis Service once your samples have been run are available via the links below:

  • Example Certificate of Analysis (CoA) – download PDF
  • Example of Result File – download zip file (right-click and “save as” to your device)

For an overview of the practical details involved in ordering a study, sending samples and receiving results using Olink Explore 1536 from our in-house Analysis Service labs in Boston MA or Uppsala, Sweden, PLEASE READ OUR GUIDE.

Statistical analysis

If more extensive data analysis services are required, our team of Olink data scientists can help you with customized statistical analysis – see our statistical analysis services.

You may also try our new post-assay statistical analysis app, Olink® Insights Stat Analysis, where you can perform basic statistical analyses and generate heatmap, sample distribution or Principal Component Analysis (PCA) plots yourself.

Example of Principle Component Analysis (PCA) plot

 

Contact us

Questions about Olink Explore?

*”Illumina” and “NovaSeq” are trademarks and the property of Illumina Inc. or their respective owners.

Data generation with Olink® Target

Detection and sample analysis is performed by high-throughput real-time PCR analysis using the Fluidigm® BioMark™ HD System. This PCR platform is a truly high capacity system which enables simultaneous detection of 96 analytes in 96 samples creating 9,216 datapoints from a single run. The BioMark HD platform uses microfluidic distribution of sample and assays requiring only 7 nL reactions and takes less than 3 hours to complete. The BioMark HD platform consists of a real-time PCR instrument, 96×96 Dynamic Array™ IFC for multiplexing the samples and biomarkers, and an IFC loader to load the 96×96 Dynamic Array IFC. More information can be found on the Fluidigm website.

(Fluidigm, BioMark, and Dynamic Array are trademarks or registered trademarks of Fluidigm Corporation).

Data generation and quality control with Olink® NPX Manager

For easier data analysis Olink Proteomics has developed a software, Olink NPX Manager, which can be used for data generated with the Olink panels and the Fluidigm BioMark real-time PCR instrument. This is an easy-to-use tool that lets you import,  quality-control and normalize your Olink data, providing you with Normalized Protein eXpression (NPX) values for further data analysis.

If you want to purchase an Olink NPX Manager license, please contact your Business Development Manager at Olink Proteomics, or email sales@olink.com

Customers who run Olink studies through one of our Analysis Service labs or authorized service providers will receive project reports that are generated from NPX Manager.

You can see dummy examples of the post-analysis documents you will receive by clicking the links below:

Certificate of Analysis for project

Results file explanation

Results file with NPX/LOD data  (Excel file)

2947

Biomarker assays

~475 million

Protein data points generated

662

Publications listed on website

35

Certified Core Labs

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