Proteomics data preprocessing

A number of internal and external controls were added to the plasma samples for quality control to monitor protein–antibody reactions, the DNA extension step, and detection quality of the qPCR in order to estimate the background signal and to calculate the limit of detection (LOD) for Olink panels. Proteins below LOD were imputed with LOD²⁷. Protein concentration was represented in arbitrary units on a log₂ scale and termed Normalized Protein eXpression (NPX), i.e., a one NPX difference means a doubling of protein concentration. The NPX value represented a relative quantification so that the data for a specific protein can be compared across different samples. Reference samples run on plates from different batches were included for batch-effect correction. The adjustment factor at protein level for each batch was calculated as median NPX of the bridging samples and subtracted from the NPX values of each sample. Batch-corrected log- transformed NPX was used in subsequent analyses (termed normalized NPX). We compared the reproducibility of the bridging samples using Pearson correlation. Supplementary Figure 1 shows the high reproducibility of the Olink panels across six representative sets of technical duplicates, with a mean correlation r = 0.97.