PTH, ELISA

The DiaSource hPTH-ELISA is a solid phase Enzyme Amplified Sensitivity Immunoassay performed on microtiterplates. Calibrators and samples react with the capture polyclonal antibodies (PAb, goat anti 1-34 PTH fragment) coated on microtiter well.

After incubation, the excess of antigen is removed by washing. Then monoclonal antibodies (MAb, mouse anti 44-68 PTH fragment) labelled with horseradish peroxidase (HRP) are added. After an incubation period allowing the formation of a sandwich: coated PAbs – human PTH – MAb – HRP, the microtiterplate is washed to remove unbound enzyme labelled antibody.

Bound enzyme-labelled antibody is measured through a chromogenic reaction. The chromogenic solution (TMB) is added and incubated.

The reaction is stopped with the addition of Stop Solution and the microtiterplate is then read at the appropriate wavelength.

The amount of substrate turnover is determined colourimetrically by measuring the absorbance, which is proportional to the PTH concentration. A calibration curve is plotted and PTH concentration in samples is determined by interpolation from the calibration curve.

The use of the ELISA reader (linearity up to 3 OD units) and a sophisticated data reduction method (polychromatic data reduction) result in a high sensitivity in the low range and in an extended calibration range.