This free PSA enzyme linked immunosorbent assay (ELISA) kit uses a technique called quantitative sandwich immunoassay. The microtiter plate provided in this kit has been pre-coated with a monoclonal antibody specific for free PSA. Standards or samples are then added to the microtiter plate wells and free PSA, if present, will bind to the antibody pre-coated on the wells.

In order to quantify the amount of free PSA present in the sample, a preparation of horseradish peroxidase (HRP)-conjugated PSA antibody is added to each well. The conjugated antibody will bind to the free PSA immobilized on the plate after incubation. All unbound components will be removed by subsequent washing procedure. Next, 3, 3', 5, 5' tetramethyl-benzidine(TMB), a substrate for HRP is added to each well. The colorimetric reaction is proportional to the free PSA content in each well. The reaction is terminated by the addition of a sulphuric acid solution and the colour change is measured spectrophotometrically at the wavelength of 450nm ± 2nm.

In order to measure the concentration of free PSA in samples, this Human Free PSA ELISA Kit includes a set of calibration standards (7 standards). The calibration standards should be assayed at the same time as the samples to allow the operator to produce a standard curve of Optical Density (O.D.) versus free PSA concentration (ng/mL). The concentration of free PSA in the samples is then determined by comparing the O.D. of the samples to the standard curve.