Bovine Nitrotyrosine (NT) ELISA Kit

Nitrotyrosine is a stable biochemical marker formed when reactive nitrogen species nitrate tyrosine residues on proteins, serving as a direct indicator of nitric oxide-mediated oxidative stress and peroxynitrite injury. This competitive ELISA from Assay Genie is calibrated for bovine serum and plasma samples and is widely applied in cardiovascular, inflammatory, neurodegenerative, and toxicology research to quantify protein nitration levels. The 96-well microplate format supports standard curve generation alongside duplicate sample measurements in a single run. Reconstituted standards and samples should be aliquoted to limit freeze-thaw cycles. The kit is stored at 2-8 °C between uses, the colorimetric enzymatic reaction is read at 450 nm with a microplate reader, and the assay sensitivity typically reaches the low nmol/L range. All components are supplied for research use only and are not intended for diagnostic procedures.

The competitive ELISA format uses a pre-coated microplate where sample nitrotyrosine competes with a conjugated reference for binding sites, producing an inverse absorbance relationship. Each 96-well plate from Assay Genie accommodates up to 40 unknown samples run in duplicate alongside an 8-point standard curve typically spanning 0.5-100 nmol/L. Sample preparation requires 50 µL of serum or plasma per well, with EDTA plasma preferred over heparinized samples to minimize assay interference. The complete workflow includes an initial incubation step, wash cycles, and a 30-minute substrate reaction with TMB, producing a blue color that turns yellow upon stop solution addition. All reagents should equilibrate to room temperature before use, and the supplied wash buffer concentrate must be diluted 1:25 with deionized water. Beyond the cardiovascular and neurodegenerative research noted in the product overview, this assay is commonly deployed in bovine models of mastitis, endotoxemia, and ruminal acidosis to quantify nitrosative stress, and is often paired with immunohistochemistry for 3-nitrotyrosine in fixed tissue sections. Reproducibility is supported by intra-assay CV under 10% and inter-assay CV under 15% when standard operating procedures are followed. Reconstituted standards remain stable for approximately 14 days at 2-8 °C, and any unused strips should be returned to the foil pouch with desiccant and sealed to preserve antibody activity for subsequent runs.