ABSTRACT
Control of monoclonal antibody (mAb) concentrations in serum is important for maintaining the safety and efficacy of these lifesaving therapeutics. Point-of-care (POC) quantification of therapeutic mAbs could ensure that patients have effective mAb levels without compromising safety. This work uses mimotope-functionalized microporous alumina affinity membranes in vertical flow assays for detection and quantitation of therapeutic mAbs. Selective capture of bevacizumab from 1000:1 diluted serum or plasma and binding of a fluorescently labelled anti-human IgG secondary antibody enable fluorescence-based analysis of bevacizumab at its therapeutically relevant concentration range of â¼50-300 µg/mL. The assay results in a linear relationship between the fluorescence intensity of the antibody capture spot and the bevacizumab concentration. A simple prototype microfluidic device containing these membranes allows washing, reagent additions and visualization of signal within 15 min using a total of 5 mL of fluid. The prototype devices can monitor physiologically relevant bevacizumab levels in diluted serum, and future refinements might lead to a POC device for therapeutic drug monitoring.
Subject(s)
Antibodies, Monoclonal , Lab-On-A-Chip Devices , Humans , BevacizumabABSTRACT
Abasic site-containing DNA duplex is a versatile structural motif that can be used for the design of purine aptamers and sensors. In this study, several modifications were introduced to the abasic site to explore possible specific binding of free 8-oxoG, a product of DNA base excision repair. The nucleoside opposite the abasic site was replaced by pyrrolo-dC as a reporter group. Binding of 8-oxoG quenched pyrrolo-dC fluorescence by as much as 70%. In contrast, adenine, guanine, thymine, and cytosine showed only minimal fluorescence quenching effect. The best aptamer binds 8-oxoG with a dissociation constant of 5.5±0.8µM. This sensor can be used to accurately measure 8-oxoG concentrations in the presence of guanine.
