Fundamentals/Therapeutics: F.GI01.11: Live Keynote III
F.GI01.01.03 Membrane Based Affinity Capture to Quantify Antibodies to COVID-19
Merlin Bruening, University of Notre Dame
This live session talk is part of the Fundamentals/Therapeutics Live Keynote III. Prof Merlin Bruening, from the University of Notre Dame, and their group have devised a process to use membrane-based affinity to capture, elute, and quantify the concentration of COVID-19 specific antibodies. This technique could elucidate an easy way to calculate the amounts of antibodies against SARS-CoV2 in serum.
The aim is to remove/elute antibody of interest specific to SARS-CoV2 from the mix of antibodies and proteins available in serum. To make this possible, the team uses nylon membrane and functionalizes them to capture/attract antibodies. The nylon membrane is functionalized with layer-by-layer deposition of polyelectrolytes like poly(acrylic acid) (PAA). Rinsing the membrane at low pH effects in carboxylic acid groups which can be used to then be modified with polyethyleneimine (PEI) polycations. Further after functionalization, a peptide mimotope can be added to the modified nylon which binds to specific protein regions. Preliminary data shows that when using Avastin/serum and eluting them using SDS/DTT, the resulting eluate contains pure Avastin captured and eluted. The next objective was to quantify the Avastin antibody. Avastin antibody quantitation was performed with spot blotting over PAA membrane and observing fluorescence with a secondary fluorescent-labeled antibody.
For calibration of the technique with COVID-19 antibodies, an anti-RBD protein is being functionalized with the membrane which showed a calibrated increase in the spot intensity of SARS-CoV2 monoclonal Antibodies. Data collected from fluorescence capturing of COVID antibodies would be helpful in analyzing if the patient has sufficient amounts of antibodies to have monoclonal antibody therapy. Further developments can yield in an efficient antibody detecting system!
Picture credit: Anal. Chem - Liu et al. Anal. Chem. 2018, 90, 20, 12161-12167