Oxiselect 8 iso prostaglandin f2α elisa kit

Urine samples with sufficient volume were analyzed for 8-OHdG and F₂-isoprostane at the New York University High Throughput Biology Laboratory (N = 2,465 samples, N = 618 individuals, and N = 1,287 samples, N = 522 individuals, respectively). 8-OHdG was quantified by competitive ELISA using the OxiSelect Oxidative DNA Damage ELISA (Cell Biolabs, San Diego, CA). Similarly, F₂-isoprostane was measured with a competitive enzyme-linked immunoassay, the OxiSelect 8-iso-Prostaglandin F₂α ELISA Kit (Cell Biolabs). All analyses were conducted in duplicate as directed by the manufacturer. Intra-assay CVs ranged from 4.6% to 11.1% for 8-OHdG and from 6.8% to 9.7% for F₂-isoprostane. Inter-assay CVs ranged from 3.9% to 16.6% for 8-OHdG and from 14.2% to 15.9% for F₂-isoprostane. Measures below the LOD were imputed by the LOD divided by the square root of 2 [50 ].

Secreted 8-iso-PGF2α was measured by competitive enzyme-linked immunoassay using the OxiSelect 8-iso-Prostaglandin F2α ELISA kit (STA-337, Cell Biolabs Inc, San Diego, CA, USA). Levels were measured in 55 μl of cell culture media, in duplicate, according to the manufacturer’s instructions. Samples were processed on three species-balanced 96-well plates. 8-iso-PGF2α was quantified relative to a standard curve using 4- and 5-parameter logistic models implemented in the drc package in R. Final 8-iso-PGF2α release is reported as four measurements either relative to the baseline, condition A, or the hypoxic condition that is A (A-A), B (B-A), C (C-B) and D (D-B). For the three individuals with replicate experiments in each species, mean values from both experiments are reported.

The determination of thiobarbituric acid reactive substances (TBARS) was carried out with the TBARS (Trichloroacetic acid method) Assay Kit (Cayman Chemical, Ann Arbor, MI, USA) using a colorimetric standard and following manufacturer’s recommendations. The measurement of F2-isoprostanes in 24-h urine was performed with the OxiSelect™ 8-iso-prostaglandin F2α ELISA kit (Cell Biolabs, Inc., San Diego, CA, USA). A competitive Elisa kit was used to measure 8-OHdG in 24-h urine (Abcam, Cambridge, MA, USA). Urinary creatinine was used as a control for the 24-h urine collection and to normalize the values of urinary F2-isoprostanes and 8-OHdG, which are presented as ng/mg creatinine.