Significant correlation between endothelin-1 levels in human plasma from coronary arteries measured by enzyme linked immunosorbent assay and olink proteomics proximity extension assay

Abstract Introduction New endothelin antagonists have recently been approved for the treatment of cardiovascular conditions including drug resistant hypertension and cerebral vasospasm associated with subarachnoid haemorrhage, where elevated endothelin-1 (ET-1) levels have been shown to contribute to increased vasoconstriction (1). ET-1 is also linked to coronary microvascular dysfunction and may be a future treatment target. The Olink Proteomics platform is widely used for the simultaneous analysis of multiple plasma proteins in very small sample volumes of 10 µL, including ET-1. The cross-reactivity of antibodies detecting ET-1 in plasma has not yet been disclosed, and it is unclear how Olink-derived values relate to biologically active ET-1. Purpose Our aim was to compare ET-1 values obtained via the Olink Explore platform with those measured using an ET-1-specific ELISA in paired coronary artery plasma samples. Method Plasma samples aspirated from the coronary artery ostium of 29 patients with coronary artery disease were analysed. ET-1 was measured in duplicate using the Quantikine sandwich ELISA, which does not cross-react with other ET peptides released from the vascular endothelium, such as big ET-1. These measurements were compared with proteomic analysis using the Olink platform, which quantifies up to 3,072 proteins. The Olink assay employs dual antibodies conjugated to DNA oligonucleotides, forming detectable double stranded DNA upon protein binding, which is then amplified and quantified via next-generation sequencing. Pearson’s correlation, linear regression analysis and Bland-Altman plot was used to assess the relationship and agreement between the assays. Results The cohort was 80% male, with a median age of 64 (IQR 40-69); 13% had diabetes. Olink intra- and inter-assay coefficients of variation (CoV) were 2.2% and 2.4%, respectively. ELISA intra- and inter-assay CoV were 6.3% and 7.3%, respectively. Olink ET-1 concentrations correlated with ELISA ET-1 levels (r = 0.53, p = 0.003), and Olink values significantly predicted ELISA results (p = 0.003). Secondary analysis showed a significant correlation between Olink ET-1 and ETB receptor concentrations (r = 0.40, p 0.05). Bland-Altman plot showed good agreement between the 2 assays and no systematic bias. Conclusion The Olink ET-1 assay demonstrated a significant correlation with the ELISA. These results suggest that the Olink assay can be used to identify changes in plasma levels of the biologically active peptide associated with pathophysiological conditions and as a biomarker for ET-1 release in clinical trials.Figure 1 Figure 2