Assay Principle

This kit employs double antibody sandwich ELISA technology: Capture Antibody is coated on the microplate to capture TNF-α 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 TNF-α concentration in the sample, and the concentration is calculated by plotting a standard curve.

Typical Data

The following data and curve are for reference only. The experimenter must establish a standard curve based on their own experimental data.

 

Standard Conc. (pg/mL)	1000	500	250	125	62.5	31.25	15.625	0
OD Value	2.707	1.531	0.791	0.422	0.221	0.131	0.088	0.036
Corrected OD Value	2.671	1.495	0.755	0.386	0.185	0.095	0.052	0

 

Sensitivity

The minimum detectable concentration (sensitivity) of Human TNF-α, determined by testing samples, is 7.81pg/mL.

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