Common use of ELISA Clause in Contracts

ELISA. Concentrations of IL-6, IL-8, FGF2, thymus- and activation-regulated chemokine (TARC) and VEGF in cell culture supernatants were measured by enzyme- linked immunosorbent assay (ELISA) (all purchased from PeproTech, London) according to the manufacturer’s instructions. The standard curves for human IL- 8, VEGF, IL-6 and FGF2 ranged from 16-2000 pg/ml, 16-1000 pg/ml, 32-2000 pg/ml and 63-4000 pg/ml respectively. The individual assays were reported to show no cross-reactivity. A summary of an ELISA protocol is outlined here: A 96-well plate was incubated overnight with the capture antibody at 0.5 µg/ml in PBS (pH 7.20 in sterile water), and subsequently blocked with PBS containing 1 % BSA (R&D Systems). After washing the plate with PBS buffer containing 0.05 % Tween-20 (R&D Systems), appropriate dilutions of samples and standards were added, incubated for 2 hours at room temperature, washed, and incubated with a detection antibody for 2 hours at room temperature. After washing, streptavidin-HRP conjugate (1:200; R&D Systems) was added. Finally, ABTS substrate (100 μl) (R&D Systems) was added. The reaction was stopped with 2 N H2SO4 (R&D Systems), 50 μl per well. The plate was read on an ELISA plate- reader at an optical density (OD) of 450 nm with wavelength correction set at 650 nm.