I have used octet red96 with 8 channel. depending on affinity, it needs very little protein. I mainly worked with either biotinylated peptide or avi tagged protein. I typically load 5 ug/ml of peptide or ~25-50 ug /ml of avi tagged protein (due to poor labeling of avi tag). of course for your analyte, you need to vary protein concentration from less than Kd to 3-5 times more than Kd. Kd of my protein was less than 1 uM. when your Kd increases, it start showing nonspecific binding at higher concentration of analyte.

I like streptavidin sensor due to less non-specific binding. Ni-NTA is not good due to non-specific interaction. I used 18 amino acid long peptide or 45 KDa protein on load and ~60 KDa protein as analyte with good results. others have studied peptide- pdz domain (~100 aa) interaction, and some had used 20 nucleotide DNA with ~20 KDa bZIP. it all depends on increase in length after interaction.

hope its helpful.

Dhiraj

Thank you!