title: Rapid and sensitive determination of protein-nitrotyrosine by ELISA: Application to human plasma.
abstract: 3-Nitrotyrosine (3NT) is known as an important indicator of nitrosative stress and has been linked to various diseases. Our aim was to develop an indirect ELISA (enzyme-linked immunosorbent assay) method suitable for the detection of protein-bound 3NT in clinical plasma and serum samples. Nitrated protein standards and reduced protein standards were prepared. Limit of detection was determined for standards; recovery and reproducibility were determined for human plasma samples. The limit of detection for this method is 1.82±0.56 pmol/mg protein. Mean recovery of standards was 95%. 3NT concentration in plasma samples of obese and normal weight subjects was determined to be between 2 pmol/mg and 19 pmol/mg. No time-consuming sample preparation or expensive laboratory equipment is required, and applied antibodies are commercially available. Sensitivity, rapid analysis time, possibilities of high throughput applications and small sample volumes make this ELISA attractive for use in clinical laboratories.
authors.affiliation: Institute of Nutrition, Department of Nutritional Toxicology, Friedrich Schiller University of Jena, Dornburger Strasse 24, Jena, Germany.
authors.forename: Daniela
authors.initials: D
authors.lastname: Weber
authors.forename: Nicole
authors.initials: N
authors.lastname: Kneschke
authors.forename: Stefanie
authors.initials: S
authors.lastname: Grimm
authors.forename: Ina
authors.initials: I
authors.lastname: Bergheim
authors.forename: Nicolle
authors.initials: N
authors.lastname: Breusing
authors.forename: Tilman
authors.initials: T
authors.lastname: Grune
mesh_terms: D000293:Adolescent; D000328:Adult; D000368:Aged; D000369:Aged, 80 and over; D000818:Animals; D001798:Blood Proteins / Q000032:analysis*; D002417:Cattle; D003902:Detergents / Q000494:pharmacology; D004797:Enzyme-Linked Immunosorbent Assay / Q000379:methods*; D005260:Female; D006801:Humans; D057230:Limit of Detection; D008875:Middle Aged; D009765:Obesity / Q000097:blood; D010084:Oxidation-Reduction; D030421:Peroxynitrous Acid / Q000494:pharmacology; D010949:Plasma; D011136:Polysorbates / Q000494:pharmacology; D011309:Preservation, Biological; D055550:Protein Stability; D012015:Reference Standards; D012680:Sensitivity and Specificity; D044967:Serum; D012710:Serum Albumin, Bovine / Q000737:chemistry; D013696:Temperature; D013997:Time Factors; D014443:Tyrosine / Q000031:analogs & derivatives* / Q000032:analysis; D055815:Young Adult
chemical_list: D001798:Blood Proteins; D003902:Detergents; D011136:Polysorbates; D030421:Peroxynitrous Acid; D012710:Serum Albumin, Bovine; C002744:3-nitrotyrosine; D014443:Tyrosine