HCP Assay

On Friday (October 11th), I had a variety of different tasks at RPI. My main task was to help Doug with an host cell protein (HCP) assay. In this experiment, we plan to analyze the binding of HCP to a peptide array using high-thoroughput screening of fluorescent tags. To do so, the peptide arrays will be incubated with HCP solution, primary antibody, and secondary antibody, separated by thorough washings to remove the excess solution. We will be testing different concentrations of primary antibody. The secondary antibody is fluorescently tagged, so we can screen the slides for intensity, indicating the intensity of HCP bound by each peptide.

Figure by JP Trasatti, Karande Lab, RPI

Previously, Doug printed peptides arrays onto three different slides and coated them with an HCP solution to bind the peptides. My first job was to wash the HCP solution off of the slides. This washing involves pouring off the solution from the slide and adding 10 mL of PBS to the petri dish, then rotating them for 10 minutes to wash off any excess HCP that was not bound by the peptide array. We then made the different concentrations of primary antibody and applied them to the respective slides, making sure all of the solution stayed on the slide and was evenly distributed. After an hour of incubation, we then washed the primary antibody and applied the light-sensitive secondary antibody.

While we were waiting for the primary antibody incubation, we were going to make western transfer buffer, but we did not have enough methanol. Instead, we made 2L of PBS, which is a process I have done before!

I am excited to return to RPI and see the results of this experiment, but I will unfortunately not be able to go this Friday (October 18th) due to Parents' Day.