Single-Molecule Ex Situ Atomic Force Microscopy Allows Detection of Individual Antibody-Antigen Interactions on a Semiconductor Chip Surface.

Although in situ atomic force microscopy (AFM) allows single-molecule detection of antibody-antigen binding, the practical applications of in situ AFM for disease diagnosis are greatly limited, due to its operational complexity and long operational times, including the execution time for the surface chemical/biological treatments in the equipped glass liquid cell. Herein, a method of graphically superimposed alignment that enables ex situ AFM analysis of an immobilized antibody at the same location on a semiconductor chip surface before and after incubation with its antigen is presented. All of the required chemical/biological treatments are executed feasibly using standard laboratory containers, allowing single-molecule ex situ AFM detection to be conducted with great practicality, flexibility, and versatility.