Binding Assays

A binding assay I was working on. I was pretty proud of this image that I took. :-) During my last couple of weeks, I did a lot of binding assays on the BTC3 cells. We wanted to find out how well the heterobivalent ligand that we had created would bind to the cells at different concentrations, from 2.5 nM all the way up to 100 nM. We also measured how well it would bind if it was in competition with one of the monovalent ligands.
First, the heterobivalent ligand was labeled with Cy-5 (a fluorescent dye). We measured its binding (which had a high affinity and bound tightly) at different concentrations, then using new cells, would incubate them with a monovalent ligand for five minutes to take up all the receptors for that ligand, then add the multivalent ligand. We found that while binding was diminished, it was still high and tight, showing that our ligand bound with great specificity and tightness. My data was in the form of imges such as these, which I used to analyze pixel intensity, which correlated to binding of receptors. The flourescent outlines of the cells shows that this ligand bound to the cell's receptors.