Assay Principle

This kit employs double antibody sandwich ELISA technology: Capture Antibody is coated on the microplate to capture IFN-γ from samples and standards. After washing, biotin-labeled Detection Antibody is added and incubated, followed by washing to form a "Capture Antibody-Antigen-Detection Antibody" immune-complex. Subsequently, Streptavidin-Horseradish Peroxidase (SA-HRP) is added and incubated. After incubation and washing, TMB Substrate is added for color development. If the target analyte is present in the sample, a blue color develops. Stop Solution is then added to terminate the reaction. During the assay, unbound components are washed away. The Optical Density (OD) is measured at 450 nm using a microplate reader. The intensity of the color is proportional to the IFN-γ concentration in the sample, and the concentration is calculated by plotting a standard curve.

Precision

Intra-assay and inter-assay Coefficients of Variation (CV) are both <10%.

· Intra-assay Precision: Three known concentration samples were assayed 20 times on one plate. The CV of the concentrations was calculated.

· Inter-assay Precision: Three known concentration samples were assayed in 20 replicates across three different plates. The CV of the concentrations was calculated.

Recovery

Recovery was tested by spiking known concentrations of Bovine IFN-γ into different sample matrices. The recovery range and average recovery are shown below.