title: Immunochemical Determination of Pyocyanin and 1-Hydroxyphenazine as Potential Biomarkers of Pseudomonas aeruginosa Infections.
abstract: A novel immunochemical approach to diagnose Pseudomonas aeruginosa infections is reported, which is based on the quantification of relevant and specific virulence factors secreted by this microorganism. Specific antibodies have been raised using hapten PC1 (a 1:1 mixture of 9-hydroxy- and 6-hydroxy-phenazine-2-carobxylic acids), designed to recognize 1-hydroxyphenazine (1-OHphz), which is the main metabolite of pyocyanin (PYO). PYO is one of the most important virulence factors produced by nearly all P. aeruginosa strains, and other species do not produce this factor. With these antibodies, an immunochemical analytical procedure able to quantify both 1-OHphz and PYO in complex clinical samples has been developed. 1-OHphz can be directly measured in solubilized sputum samples diluted 20 times with the assay buffer. Quantification of PYO is accomplished after conversion to 1-OHphz in just 20 min under basic conditions. A LOD of 0.60 ± 0.01 nM (4.80 ± 0.08 nmol kg(-1) sputum) is reached for both biomarker targets under the conditions established, a value that is much below the reported concentrations on sputum samples obtained from infected patients (up to 100 μM). The assay is robust, reproducible, accurate, can be run in about 2 h, and many samples can be measured simultaneously. The present reported assay could represent a significant improvement in the diagnosis of infectious diseases caused by this pathogen.
authors.affiliation: Nanobiotechnology for Diagnostics (Nb4D), IQAC-CSIC , ‡Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Spain, Jordi Girona, 18-26, 08034 Barcelona, Spain.
authors.forename: Carme
authors.initials: C
authors.lastname: Pastells
authors.affiliation: Nanobiotechnology for Diagnostics (Nb4D), IQAC-CSIC , ‡Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Spain, Jordi Girona, 18-26, 08034 Barcelona, Spain.
authors.forename: Nuria
authors.initials: N
authors.lastname: Pascual
authors.affiliation: Nanobiotechnology for Diagnostics (Nb4D), IQAC-CSIC , ‡Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Spain, Jordi Girona, 18-26, 08034 Barcelona, Spain.
authors.forename: Francisco
authors.initials: F
authors.lastname: Sanchez-Baeza
authors.affiliation: Nanobiotechnology for Diagnostics (Nb4D), IQAC-CSIC , ‡Networking Research Center on Bioengineering, Biomaterials and Nanomedicine (CIBER-BBN), Spain, Jordi Girona, 18-26, 08034 Barcelona, Spain.
authors.forename: M-Pilar
authors.initials: MP
authors.lastname: Marco
mesh_terms: D000818:Animals; D000906:Antibodies / Q000737:chemistry / Q000276:immunology; D015415:Biomarkers / Q000032:analysis; D005260:Female; D007120:Immunochemistry; D008958:Models, Molecular; D015394:Molecular Structure; D010619:Phenazines / Q000032:analysis* / Q000276:immunology; D011552:Pseudomonas Infections / Q000175:diagnosis* / Q000276:immunology; D011550:Pseudomonas aeruginosa / Q000737:chemistry* / Q000276:immunology*; D011710:Pyocyanine / Q000032:analysis* / Q000276:immunology; D011817:Rabbits
chemical_list: D000906:Antibodies; D015415:Biomarkers; D010619:Phenazines; C050093:1-hydroxyphenazine; D011710:Pyocyanine