Atomic Force Microscopy-based Immune Recognition Of Biomolecules

The immune recognition of biomolecules should facilitate the disease diagnosis and treatment because many diseases highly correlate with biomolecular functions and structures. Here, atomic force microscopy (AFM) and electrochemistry analyses were employed to construct and characterize the nanostructures and electrochemistry of biosensing surface that was created by a sequential self-assembling of bioactive aminobenzenthiol oligomer (O-ABT), glutareldehyde and anti-transferrin (anti-Tf) antibody on the electrode gold surface. Under AFM, a complete coverage of bioactive O-ABT interface could be achieved by anti-Tf antibody at an optimal concentration. The anti-Tf antibody immobilized on electrode surface of the immunosensor exhibited globular-shape topography with some degree of aggregation. The anti-Tf molecules on the immunosensor exhibited a greater molecular force bound to holo-Tf (iron-containing form of Tf) than that to apo-Tf (iron-absent form of Tf). Consistently, the anti-Tf immunosensor had a greater electrochemical capacity to sensitize apo-Tf than holo-Tf, supporting the molecular force-based finding by AFM. The present study elucidated the nanostructures and molecular recognition bases for the immunosensing capacity of a highly sensitive capacitive immunosensor. And then, molecular architecture was employed to construct and characterize the specific cell surface labeling that was created by a sequential specific binding of outer molecules (OMs) with appropriate structures and functions and targeted molecules (TMs) on the cell surface. The specificity and feasibility of molecular architecture were demonstrated. The molecular architecture was tested in K562 cells and the result reveled that the fibronectin receptors on cell surface could be clearly localized under AFM. Further, AFM was used to characterize whether the endocytosis of ligand-conjugated quantum dots (QDs) resulted in the nanostructure changes, which should highly correlate with the basic mechanism of the receptor-mediated transduction. The cellular nanostructure analysis by AFM revealed that the ligand-conjugated QDs were highly specific and potent to cell receptor. The confocal scanning microscopy further demonstrated that the fluorescent intensity in the cell central parts is larger than the cell edge, which was most likely the result of internalization of ligand-conjugated QDs. The experiment also indicated that theinternalization of ligand-conjugated QDs during endocytosis was near linear with time, which is consistent with a previous reported mechanism. The flow cytometry that associated with the QDs endocytosis well confirmed the microscopic results. This experiment should provide novel insights into the design and application of QDs fluorescent probes in biology and immunology. Importantly, this work demonstrated that, combined with QDs, AFM was a powerful tool to investigate the receptor-mediated transduction and the molecular recognition of cell surface molecules.