Background: With the emergence of COVID-19, the urgency to understand the immune response towards infections has expanded. Severe cases have shown the involvement of a wide range of inflammatory modulators, where the detection of low-abundance effector proteins might be challenging. Here, we assess the detectability of immune-related markers using two high-throughput multiplexed proteomics immunoassays.

Methods: We deployed the Olink and NULISAseq-based proteomics platforms to blood samples from healthy controls (n=44) and critically ill COVID-19 patients (n=83), using three Olink panels (Inflammation, CVDII, CVDIII; 255 unique proteins) and the NULISAseq inflammation panel (203 proteins). In common, Olink and NULISAseq panels share 94 analytes. Statistical comparisons were performed using FDR-adjusted linear models to construct volcano and ratio plots, heatmaps and KEGG pathways.

Findings: NULISAseq platform showed higher dynamic range of detectability and higher CV values across samples than Olink according to the shared proteins (n =94). High analyte accordance between both technologies was achieved, where 71 proteins showed strong to very strong correlations. All analytes showed a positive correlation (Spearman rho=0.767), except for two targets (KITLG and TSLP). Differential protein expression showed similar patterns of up- and down-regulation with high significance (-log10 FDR p-value up to 50), such as IL6, CCL7, CXCL8, and CXCL10. 62 proteins shared significant values on both platforms; of which 55 showed concordant effect directionality, but 7 showed opposite directionality (IL6R, IL1R2, IL15RA, KITLG, TSLP, IL17C, and IL4R). KEGG Pathway analysis revealed enrichment of common infection-related pathways such as cytokine-cytokine interaction and viral protein-cytokine interaction concordantly. Additionally, Olink and NULISAseq data yielded 14 and 16 solely significant proteins among the shared proteins, plus 9 and 36 additional significant cytokines among 161 and 109 exclusive proteins, respectively.

Interpretation: Both Olink and NULISAseq techniques effectively detected low-abundance proteins with high sensitivity and multiplexing ability using a low volume of samples. NULISAseq outperformed Olink in sensitivity and dynamic range. Nevertheless, both techniques shared comparable differential expression and enrichment findings and provided unique cytokines profile, which need further validation and verification. Therefore, we propose a complementary approach for a comprehensive understanding of the immune response to COVID-19.