Functionalized Porous Membranes for Antibody Quantification

Modification of porous membranes with polyelectrolyte multilayers and subsequent attachment of mimotope peptides or antigens enables capture of relevant monoclonal antibodies (mAbs) in diluted human serum. This thesis explores the functionalization of nylon membrane and followed by embedding the membrane in spin columns for a 30 min assay. Elution with detergent/reducing agent solution and analysis of intrinsic tryptophan residues in eluted mAbs allows concentration measurement, using calibration curves established using mAb standards. To further improve our antibody quantification method, we modify glass fiber membrane in a 96-well plate and this assay requires only 5 min processing time. We reduce detection of non-specific binding by employing fluorescently labelled secondary antibodies with specificity against human immunoglobulin G (IgG) to bind to immobilized mAb and the measured fluorescence intensity directly correlates with the level of mAb in samples. We demonstrated the capability of our functionalized membranes in spin column and 96-well plate format in quantifying Trastuzumab, Bevacizumab, and SARS-CoV-2 mAb in relevant clinical concentration ranges. Future work would involve incorporating our functionalized membranes in microfluidic devices for user-friendly and convenient point-of-care assays.