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. 2025:2929:129-142.
doi: 10.1007/978-1-0716-4595-6_10.

Proteomics by qPCR Using the Proximity Extension Assay (PEA)

Julio Manuel Martínez-Moreno  1 , Adrián Llamas-Urbano  1 , Nuria Barbarroja  1   2 , Carlos Pérez-Sánchez  3   4   5
Affiliations

Proteomics by qPCR Using the Proximity Extension Assay (PEA)

Julio Manuel Martínez-Moreno et al. Methods Mol Biol. 2025.

Abstract

The Proximity Extension Assay (PEA) is an innovative technology developed by Olink Proteomics that has revolutionized proteomics research. This method enables the simultaneous detection of hundreds of proteins using a minimal sample volume (1 microliter), combining the specificity of antibody-based immunoassays with the sensitivity of quantitative PCR (qPCR). The PEA process relies on the binding of two antibodies, each conjugated with unique DNA sequences, to a target protein. Upon proximity, these DNA sequences hybridize, forming a molecular barcode that is subsequently amplified by PCR, ensuring high specificity and minimizing cross-reactivity-an issue commonly encountered in multiplexed immunoassays. This chapter provides a detailed overview of the technical and methodological aspects of PEA, including the incubation, extension, amplification, and detection steps, as well as the quality control measures implemented in the Olink® NPX Signature software for data analysis. This technology has been applied in more than 2000 peer-reviewed studies, demonstrating its potential for biomarker discovery in diagnostics, prognostics, and personalized medicine across various diseases. The integration of PEA into biomedical research continues to enhance the precision and reproducibility of proteomics studies.

Keywords: Olink; PEA; Protein array; Proteomics; Proximity extension assay; qPCR.

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References

    1. Cuesta-López L, Escudero-Contreras A, Hanaee Y, Pérez-Sánchez C, Ruiz-Ponce M, Martínez-Moreno JM, Pérez-Pampin E, González A, Plasencia-Rodriguez C, Martínez-Feito A, Balsa A, López-Medina C, Ladehesa-Pineda L, Rojas-Giménez M, Ortega-Castro R, Calvo-Gutiérrez J, López-Pedrera C, Collantes-Estévez E, Arias-de la Rosa I, Barbarroja N (2024) Exploring candidate biomarkers for rheumatoid arthritis through cardiovascular and cardiometabolic serum proteome profiling. Front Immunol 14(15):1333995 - DOI
    1. Ruiz-Ponce M, Cuesta-López L, López-Montilla MD, Pérez-Sánchez C, Ortiz-Buitrago P, Barranco A, Gahete MD, Herman-Sánchez N, Lucendo AJ, Navarro P, López-Pedrera C, Escudero-Contreras A, Collantes-Estévez E, López-Medina C, Arias-de la Rosa I, Barbarroja N (2023) Decoding clinical and molecular pathways of liver dysfunction in psoriatic arthritis: impact of cumulative methotrexate doses. Biomed Pharmacother 168:115779 - DOI - PubMed
    1. Arias-de la Rosa I, Ruiz-Ponce M, Cuesta-López L, Pérez-Sánchez C, Leiva-Cepas F, Gahete MD, Navarro P, Ortega R, Cordoba J, Pérez-Pampin E, González A, Lucendo AJ, Collantes-Estévez E, López-Pedrera C, Escudero-Contreras A, Barbarroja N (2023) Clinical features and immune mechanisms directly linked to the altered liver function in patients with rheumatoid arthritis. Eur J Intern Med 118:49–58 - DOI - PubMed
    1. Perez-Sanchez C, Escudero-Contreras A, Cerdó T, Sánchez-Mendoza LM, Llamas-Urbano A, la Rosa IA, Pérez-Rodriguez M, Muñoz-Barrera L, Del Carmen Abalos-Aguilera M, Barbarroja N, Calvo J, Ortega-Castro R, Ruiz-Vilchez D, Moreno JA, Burón MI, González-Reyes JA, Collantes-Estevez E, Lopez-Pedrera C, Villalba JM (2023) Preclinical characterization of pharmacologic NAD+ boosting as a promising therapeutic approach in rheumatoid arthritis. Arthritis Rheumatol 75(10):1749–1761 - DOI - PubMed
    1. Barbarroja N, López-Montilla MD, Cuesta-López L, Pérez-Sánchez C, Ruiz-Ponce M, López-Medina C, Ladehesa-Pineda ML, López-Pedrera C, Escudero-Contreras A, Collantes-Estévez E, Arias-de la Rosa I (2023) Characterization of the inflammatory proteome of synovial fluid from patients with psoriatic arthritis: potential treatment targets. Front Immunol 14:1133435 - DOI - PubMed - PMC

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