Rapid, Automated, and Specific Immunoassay to Directly Measure Matrix Metalloproteinase-9-Tissue Inhibitor of Metalloproteinase-1 Interactions in Human Plasma Using AlphaLISA Technology: A New Alternative to Classical ELISA

• Published in: Frontiers in immunology 2017-07, Vol.8, p.853-853
• Main Authors: Pulido-Olmo, Helena, Rodríguez-Sánchez, Elena, Navarro-García, José Alberto, Barderas, María G, Álvarez-Llamas, Gloria, Segura, Julián, Fernández-Alfonso, Marisol, Ruilope, Luis M, Ruiz-Hurtado, Gema
• Format: Article
• Language: English
• Subjects: AlphaLISA; immunoassay; Immunology; matrix metalloproteinase-9; protein interaction; tissue inhibitor of metalloproteinase-1
• ISSN: 1664-3224; 1664-3224
• DOI: 10.3389/fimmu.2017.00853

The protocol describes a novel, rapid, and no-wash one-step immunoassay for highly sensitive and direct detection of the complexes between matrix metalloproteinases (MMPs) and their tissue inhibitor of metalloproteinases (TIMPs) based on AlphaLISA technology. We describe two procedures: (i) one approach is used to analyze MMP-9-TIMP-1 interactions using recombinant human MMP-9 with its corresponding recombinant human TIMP-1 inhibitor and (ii) the second approach is used to analyze native or endogenous MMP-9-TIMP-1 protein interactions in samples of human plasma. Evaluating native MMP-9-TIMP-1 complexes using this approach avoids the use of indirect calculations of the MMP-9/TIMP-1 ratio for which independent MMP-9 and TIMP-1 quantifications by two conventional ELISAs are needed. The MMP-9-TIMP-1 AlphaLISA assay is quick, highly simplified, and cost-effective and can be completed in less than 3 h. Moreover, the assay has great potential for use in basic and preclinical research as it allows direct determination of native MMP-9-TIMP-1 complexes in circulating blood as biofluid.