Recognition Functionality Mapping at a Single Molecular Level by Frequency-Modulation AFM Imaging

Molecular recognition is an essential function in developing advanced materials, including biomaterials. The precise design of molecular recognition functionalities, such as selectivity, affinity, and stimuli-responsibility, is the most important challenge in this research field. There are many studies on the design of molecular recognition elements using macrocyclic molecules, metal-organic complexes, and biomolecules. The designed recognition elements can capture guest molecules based on specific interactions. In addition, developing characterization tools is equally important to understand molecular-recognition functionalities and apply the advanced materials to various fields. Depending on their application, the designed recognition elements are often fixed on material surfaces. We easily imagine that the functionalities of the fixed recognition elements are influenced by their orientation and distribution on the material surfaces. However, a molecular scale understanding of the functionalities of the fixed recognition elements is still challenging due to the lack of analytical methods. Although the functionalities, such as the amount of capturing guest molecules, can be investigated with conventional analytical techniques, understanding the functionalities of each recognition element at a single molecular scale is difficult due to the averaging over numerous elements in conventional methods.<br/>To overcome the limitation of conventional analytical methods, a method based on frequency modulation atomic force microscopy (FM-AFM). The FM-AFM technique is known to have the capability to visualize subnanometer-scale structures of various molecules and materials in liquid. The subnanometer resolution of the FM-AFM technique in liquid is based on the high sensitivity for interaction forces acting on the tip apex (the detectable theoretical limit is four pN). Moreover, conservative and non-conservative (dissipative) forces are measured separately in FM-AFM imaging. The characteristics of the FM-AFM technique suggest establishing a novel analytical method to investigate the recognition functionalities at a single molecular level based on the FM-AFM.<br/>In this study, molecular-scale investigation of recognition events was demonstrated by FM-AFM imaging with a guest molecule-modified tip. Metal-salen complexes were fixed on a Si substrate as a model of molecular recognition elements. Amino-terminated AFM tips were prepared in this study because the metal-salen complexes have reported the capture of amino-containing molecules through the formation of coordination bonds. The FM-AFM experiments were performed with the amino-terminated AFM tips and the metal-salen complex fixed substrate. As a result, the fixed metal-salen complexes were visualized in the height images constructed by the measured signal of conservative forces. In contrast, energy dissipation images showed that dissipative interactions occurred in only a part of the fixed metal-salen complexes visualized in the height images. The results indicate that only a part of the fixed metal-salen complexes can recognize the amino group at the tip apex. The difference in the recognition function between the fixed metal-salen complexes is most likely caused by their orientation and dynamics. Our results demonstrate that the proposed analytical method based on the FM-AFM techniques with guest molecule-modified tips has the potential to investigate the recognition functionalities of the fixed recognition elements at the single molecular level, indicating future contributions to the development of advanced materials with recognition functions.