Using Omics Playground to Analyze Olink Proteomic Data

Published on August 6th, 2025
⏱ 3 min read

Introduction

Olink Target proteomics data are protein measurements obtained using Olink’s Proximity Extension Assay (PEA) technology. PEA technology generates relative protein quantification provided
as Normalized Protein eXpression (NPX) units, allowing comparison of protein levels across samples. 

Among the available Olink technologies, the Target panel is frequently used: it allows multiplexing, with each panel measuring up to 92 biomarker proteins simultaneously from a minimal volume of plasma or serum sample.

Olink has emerged as a research- and disease-focused technology:

  1. It offers over 10 panels covering various biological processes and disease areas, including oncology, cardiovascular, neurology, and inflammation;
  2. It has high specificity and sensitivity, and also offer great reproducibility across experiments and labs due to its highly standardized assay. Researchers use Olink Target proteomics data for various applications, including biomarker discovery, drug development, and investigating protein-protein interactions in specific disease areas or biological processes.

This white paper aims to provide guidance on best practices for analyzing Olink proteomics data in Omics Playground, ensuring the accuracy, reproducibility, and consistency provided by the OlinkAnalyze R package.

 

Olink Proteomics AB. (2025). Olink’s Proximity Extension Assay (PEA) .
Figure 1. Olink Proteomics AB. (2025). Olink’s Proximity Extension Assay (PEA) .

Distribution of Olink Normalized Protein Expression (NPX) values

At BigOmics Analytics, we assessed the distribution of Olink proteomics data in publicly-available datasets. 

We observe that NPX-normalized proteomics intensities range from -10 or -5 to 10-15 in the log2 scale. We used the two datasets available a part of the Olink Analyze R package, and Feyaerts et al. (PMID: 35839768), Carlyle et al. (PMID: 35734478), Tomalin et al. (PMID: 31705874), Bao et al. (doi: 10.3389/fnmol.2023.1185021), Wang et al. (PMID: 38045696).

As a distinctive feature of Olink Target Panel(s) data, only a minority of values are negative.

On the contrary, Olink Explore HT proteomics data are 0 centered, with about 50% of negative values.

Distribution of NPX values across multiple Olink Target. panel datasets, showing typical ranges and data patterns.
Figure 2. Distribution of NPX values across multiple Olink Target. panel datasets, showing typical ranges and data patterns.

Comparison between Olink Analyze R package and Omics Playground (Welch t-test)

Figure 3. Comparison of Log₂ fold-change and FDR values between Omics Playground and OlinkAnalyze

Conclusion

As per the Olink documentation, NPX are normalized and log2-transformed proteomic intensities.

The presented comparisons used NPX data as it is. No normalization was performed in the Omics Playground platform.

Our analyses indicate a very good concordance in analysis of Olink proteomic data between Omics Playground and OlinkAnalyze R package.

The recommended steps for analysis of Olink proteomic data in Omics Playground are:

  • Skip normalization. Alternatively, you could try out one of our proteomics- tailored methods, including MaxMedian or MaxSum normalization.
  • If interested in getting results consistent with OlinkAnalyze R package, select Welch t-test for differential protein expression testing.

Omics Playground features and capabilities for Olink Proteomics data

Feature comparison of Omics Playground analysis modules available for Olink Targeted Panels, Olink HT Explore, and LC/MS datasets.
Figure 4. Feature comparison of Omics Playground analysis modules available for Olink Targeted Panels, Olink HT Explore, and LC/MS datasets.

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