HDGF ELISA Assay Kit
The HDGF ELISA Assay Kit is For Research Use Only
Size: 1 x 96 wells
Sensitivity: 30 pg/ml
Dynamic Range: 78-5000 pg/ml
Incubation Time: 3.5 hours
Sample Type: Serum, Plasma, Cell Culture Supernatants
Sample Size: 50 µl
Alternative Name: Human Heparin Binding Growth Factor
The HDGF ELISA Assay employs the sandwhich enzyme immunoassay technique for the detection of Human HDGF in serum, plasma and cell culture supernatant samples. An antibody specific HDGF has been pre-coated onto a microtiter plate. Standards or samples are pipetted into the wells and any HDGF present is bound on the plate. After washing away an unbound substances, a Horseradish Peroxidase (HRP) conjugated primary antibody that binds to HDGF is added to each well and incubates. After washing away any unbound antibody-enzyme reagent, a substrate solution (TMB) is added to the wells and color develops in proportion to the amount of total HDGF bound in the initial step. The color development is stopped by the addition of acid and the intensity of the color is measured at a wavelength of 450 nm ±2nm. The concentration of total HDGF in the sample is then determined by comparing the O.D. of samples to the standard curve.
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HDGF is a member of the hepatoma-derived growth factor family. The encoded protein has mitogenic and DNA-binding activity and may play a role in cellular proliferation and differentiation. This gene was thought initially to be located on chromosome X, however, that location has been determined to correspond to a related pseudogene. Alternatively spliced transcript variants encoding distinct isoforms have been described. HDGF is a Heparin-binding protein, with mitogenic activity for fibroblasts. Acts as a transcriptional repressor.
- Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal it.
- Add 50 µl of standards, samples and zero controls into appropriate wells.
- Add 50 µl of Assay buffer into all wells immediately. Mix thoroughly by gently shaking or tapping the plate. Cover the plate and incubate for 2 hour at room temperature.
- Aspirate each well and wash, repeating the process 2 times for a total 3 washes. Wash by filling each well with 1X wash buffer (300 μl) using a squirt bottle, manifold dispenser, or autowasher. Complete removal of liquid at each is essential to good performance. After the last wash, remove any remaining buffer by aspirating, decanting or blotting against clean paper towels.
- Add 100 µl 1X HRP-antibody conjugate into each well. Mix thoroughly by gently shaking or tapping the plate. Cover wells and incubate for 1 hour at room temperature in dark.
- Aspirate each well and wash as step 4.
- Add 100 μl of TMB substrate to each well. Incubate for 10 minutes at RT in dark. Substrate will change from colorless to different strengths of blue.
- Add 50 μl of Stop solution to each well. The color of the solution should change from blue to yellow. Mix thoroughly by gently shaking the plate.