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CA125 and HE4 assay results and menopausal status can be used to estimate the likelihood of epithelial ovarian cancer in women presenting with a pelvic mass with the Risk of Ovarian Malignancy Algorithm (ROMA™). Results should be determined using the HE4 EIA assay (specifications outlined below) in combination with the ARCHITECT CA125II.
HE4 EIA specifications13
- Product number: 404-10
- Results: Within 2.5 hours, 2-step procedure
- Detection limit: ≤15 pM
- Measuring range: 15 to 900 pM
- Sample volume: 25 uL
- Stability: Until expiry at 2-8 °C
- Calibrator range: 0 to 900 pM
- Incubation temperature: Room temperature (20-25 °C)
- Detection: 620 nm or 405 nm
The HE4 EIA is a solid-phase, non-competitive immunoassay based upon the direct sandwich technique using two mouse-monoclonal antibodies, 2H5 and 3D8, directed against two epitopes in the C-WFDC domain of HE4. Calibrators, controls and patient samples are incubated together with biotinylated Anti-HE4 monoclonal antibody (Mab) 2H5 in streptavidin coated microstrips. HE4 present in calibrators or samples is adsorbed to the streptavidin coated microstrips by the biotinylated Anti-HE4 Mab during the incubation. The strips are then washed and incubated with HRP labeled Anti-HE4 Mab 3D8. After washing, buffered Substrate/Chromagen reagent (hydrogen peroxide and 3,3’,5,5’ tetra-methyl-benzedine) is added to each well and the enzyme reaction is allowed to proceed. During the enzyme reaction a blue color will develop if antigen is present. The intensity of the color is proportionate to the amount of HE4 present in the samples. The color intensity is determined in a microplate spectrophotometer at 620 nm (or optionally at 405 nm after addition of Stop Solution). Calibration curves are plotted for each assay by plotting absorbance value versus the concentration for each calibrator. The HE4 concentration in patient samples is then read from the calibration curve.