Human PDGF-BB Quantikine ELISA Kit Summary

Product Summary

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

Preparation and Storage

Background: PDGF-BB

The platelet-derived growth factor (PDGF) family of disulfide-linked dimeric proteins consists of four homodimeric proteins, PDGF-AA, PDGF-BB, PDGF-CC and PDGF-DD, and one heterodimeric protein, PDGF-AB. Members of this protein family act mainly on connective tissue. The dimeric isoforms of PDGF are expressed by different cell types and at different times during embryonic development. All PDGF isoforms are synthesized as inactive precursors and are processed to active forms that contain a PDGF/VEGF cysteine knot domain. Only PDGF-C and PDGF-D, also named spinal cord-derived growth factors SCDGF and SCDGF-B, also contain an N-terminal CUB domain. Mature PDGF dimers bind and induce the homo- or hetero-dimerization of two receptor tyrosine kinases (PDGF R alpha and R beta).

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.