The IDK® IDO Activity ELISA is intended for the quantitative determination of L-kynurenine and L-tryptophan in EDTA plasma, serum, and dried blood spots.

For research use only. Not for use in diagnostic procedures.

For Laboratory Professional Use Only.

This is the RUO version of Immundiagnostik part number K 7726.

Test Principle of the IDK® IDO Activity ELISA

The IDK® IDO Activity ELISA is designed for the quantitative determination of L-kynurenine and L-tryptophan. The assay is based on the method of competitive enzyme-linked immunoassays.

The sample preparation includes the addition of a derivatization reagent for the derivatization of kynurenine and tryptophan. Afterward, the derivatized standards, controls, and samples are incubated in the wells of two microtiter plates coated with:
(I) L-kynurenine-derivative (tracer) (red mark),
(II) L-tryptophan-derivative (tracer) (yellow dot).

Also, a polyclonal L-kynurenine antiserum and a polyclonal L-tryptophan antiserum are added, respectively. During the incubation period, the target antigen in the sample competes with the tracer, immobilized on the wall of the microtiter wells, to bind the polyclonal antibodies.

In the second incubation step, a peroxidase-conjugated antibody is added to each microtiter well to detect the polyclonal antibodies. After washing away the unbound components, tetramethylbenzidine (TMB) is added as a peroxidase substrate. Finally, the enzymatic reaction is terminated by an acidic stop solution. The color changes from blue to yellow, and the absorbance is measured in the photometer at 450 nm. The intensity of the yellow color is inversely proportional to the target antigen concentration in the sample. This means high L-kynurenine or L-tryptophan concentration in the sample reduces the concentration of tracer-bound antibodies and lowers the photometric signal.

A dose-response curve of absorbance unit (optical density, OD at 450 nm) vs. standard concentration is generated using the values obtained from the standards. The presence of L-kynurenine and L-tryptophan in the patient samples is determined directly from this curve.