I coated an iron oxide nanoparticle as a drug delivery vehicle. Bombesin peptide as a targeting agent and bromelain protein nature anticancer agent are conjugated to nanoparticle. I want to detect the presence and to determine the concentration for both bromelain and bombesin peptide, but I do not know how I should do quantitatively and qualitatively assay. I would much appreciate your comments.
You could possibly enzymatically assay bromelain activity (perhaps with a fluorometric substrate) and the quantify the bombesin peptide with a conjugated antibombesin antibody (HRP conjugate). If you had a fluorometric product from the bromelain assay and a colorometric product from the bombesin peptide assay you might be able to quantify both in the same well of a microtiter plate. Good luck.
You could try ELISA by coating the ELISA plate with primary antibody of each of the protein you are looking for and tag with a detectable secondary..
Aritra suggestion has an added benefit. If you are careful with the selection of labels for the enzyme and peptide and pick two capable of FRET you might also get an idea of the surface density of the two molecules on your nanoparticles by measuring energy transfer. I'm not sure if the probes would be close enough but it would be worth trying.
Depending on the MW of your peptides you have sevra choices: The ordinary HPLC-Reversed phase can be used for peptides separation and determination. But in case of larger MW, you have to use HPLC-gel filtration column. On other hand if available, the HPLC-MALDI well be the best.
- Badges
- Science topic
- Similar topics
- Cell Biology
- Methods