Visualization of the membrane engineering concept: evidence for the specific orientation of electroinserted antibodies and selective binding of target analytes

Abstract

Membrane engineering is a generic methodology for increasing the selectivity of a cell biosensor against a target molecule, by electroinserting target-specific receptor-like molecules on the cell surface. Previous studies have elucidated the biochemical aspects of the interaction between various analytes (including viruses) and their homologous membrane-engineered cells. In the present study, purified anti-biotin antibodies from a rabbit antiserum along with in-house prepared biotinylated bovine serum albumin (BSA) were used as a model antibody-antigen pair of molecules for facilitating membrane engineering experiments. It was proven, with the aid of fluorescence microscopy, that (i) membrane-engineered cells incorporated the specific antibodies in the correct orientation and that (ii) the inserted antibodies are selectively interacting with the homologous target molecules. This is the first time the actual working concept of membrane engineering has been visualized, thus providing a final proof of the concept behind this innovative process. In addition, the fluorescence microscopy measurements were highly correlated with bioelectric measurements done with the aid of a bioelectric recognition assay.

An Ultra-Rapid Biosensory Point-of-Care (POC) Assay for Prostate-Specific Antigen (PSA) Detection in Human Serum

“…The fact that the lower anti-PSA antibody concentration (0.25 ng/mL) produced the best results can be explained as follows: in accordance with previous reports [ 18 ], increasing the density of electroinserted antibodies on membrane-engineered cells and/or the concentration of target analytes above an upper limit is not associated with a titrimetric between the analyte (PSA) and the membrane- engineered carrier cells with the anti-PSA antibodies. This is due to the fact that the analyte-electroiserted antibody reaction involves an electromechanical stress at the site of antibody area on the membrane, triggering to membrane status changes such as conductivity and porosity [ 32 ]. Quite frequently, there is a limitation to the modification of cell membrane potential status caused by the increasing density of electroinserted antibodies, with lower densities producing better resolution of response.…”

Detection of Superoxide Alterations Induced by 5-Fluorouracil on HeLa Cells with a Cell-Based Biosensor

“…Receptor molecules can vary from antibodies to enzymes to polysaccharides [50][51][52][53][54]. It has been previously proven [55] that this principle is associated, in a unique way, with certain changes in the cellular electric properties (in particular, cell membrane hyperpolarization) as a consequence of the interaction of the analytes under determination with the electroinserted molecules and therefore changes in the cellular structure.…”

Development of a Portable, Ultra-Rapid and Ultra-Sensitive Cell-Based Biosensor for the Direct Detection of the SARS-CoV-2 S1 Spike Protein Antigen

“…This approach, known as Molecular Identification through Membrane Engineering, is a generic cell-based assay principle for the determination of analytes, including biomolecules, on the basis of the specific and selective interaction of the target analytes with cellular biorecognition elements, the surfaces of which have been modified by the electroinsertion of target-specific antibodies. It has been previously demonstrated that the binding of the target molecules to the electroinserted antibodies resulted in a unique and measurable change in the electric properties of the biorecognition elements, in particular the hyperpolarization of the engineered cell membrane [14][15][16]. These changes can be measured by any appropriate bioelectric/bioelectrochemical sensor.…”