Human EGF Quantikine ELISA Kit Summary

Product Summary

The Quantikine Human EGF Immunoassay kit is a 3.5 or 4.5 hour solid phase ELISA designed to measure EGF levels in cell culture supernates, serum, plasma, and urine. It contains E. coli-derived recombinant human EGF and antibodies raised against the recombinant factor. It has been shown to accurately quantitate recombinant human EGF. Results obtained measuring natural human EGF showed linear curves that were parallel to the standard curves obtained using the kit standards. These results indicate that this kit can be used to determine relative mass values for natural EGF.

Preparation and Storage

Background: EGF

EGF (Epidermal Growth Factor) is expressed as a transmembrane protein that is proteolytically cleaved to generate soluble forms. It signals through EGF R/ErbB1 which heterodimerizes with ErbB2, ErbB3, or ErbB4 and can also associate with PDGF R beta and HGF R/c-MET. During development, EGF regulates thymocyte differentiation, neuroglia production, and adipocyte maturation. In the adult, EGF plays a role in mammary gland lactogenesis, fibroblast mitosis, dissociation of the extracellular matrix, and cell migration.

Assay Procedure

Refer to the product datasheet for the complete assay procedure.

Bring all reagents and samples to room temperature before use. It is recommended that all samples, standards, and controls be assayed in duplicate.

1.     Prepare all reagents, standard dilutions, and samples as directed in the product insert.

2.     Remove excess microplate strips from the plate frame, return them to the foil pouch containing the desiccant pack, and reseal.

3.     Add 50 μL of Assay Diluent to each well.

4.     Add 50 μL of Standard, control, or sample to each well. Cover with a plate sealer, and incubate at room temperature for 2 hours.

5.     Aspirate each well and wash, repeating the process 4 times for a total of 5 washes.

6.     Add 100 μL of Conjugate to each well. Cover with a new plate sealer, and incubate at room temperature for 2 hours.

7.     Aspirate and wash 5 times.

8.     Add 100 μL Substrate Solution to each well.

9.     Add 100 μL of Stop Solution to each well. Read at 450 nm within 30 minutes. Set wavelength correction to 540 nm or 570 nm.