Physiology InFocus
From Molecules to Organisms: Approaches to Systems and Integrative Physiology
Investigating Cellular Signaling with Atomic Force Microscopy Methods

Enormous progress has been made in the recent past in understanding the cellular signaling mechanisms that allow the human organism to respond to external stimuli and yet maintain a constant, balanced internal state.  By the same token, there have been equally large strides in the application of nanotechnology to many problems in the physical sciences; but the use of nanotechnology to solve problems of biology and medicine has been limited, particularly in those areas that could have direct applicability to the health care of the American population.  Atomic force microscopy (AFM) can follow nanometer-scale surface topography rather well, and can even track dynamic changes as the same molecules are scanned repeatedly while chemical conditions are changed.  Perhaps more importantly, AFM can probe and manipulate single molecules.  However, to date it has proved to be a rather blunt instrument, scanning slowly, somewhat intrusively, and almost entirely lacking in the ability to resolve the chemical identities of molecules in biological samples.  However, recently, there have been significant advances in extending the capabilities of AFM, enabling rapid sensing of localized cellular signaling events, mapping of cell surface proteins, and detecting intermolecular interactions while still providing topological information about cellular structure.  It is now possible to add small molecule chemical detection to AFM measurements by using nanofabrication methods to incorporate nanometer-scale electrochemical sensors into the AFM tips to acquire chemical and topographical information with high temporal and spatial resolution (Presentation 1).  More recently, techniques have been developed for attaching large (bio)molecules such as antibodies or transcription factors to the AFM tip and detecting the interaction between these molecules and binding partners as the AFM tip is moved across the cell surface.  This allows detection and localization of cell surface receptors and ion channels (Presentation 2) or transcription factor and histone binding and unbinding to DNA (Presentation 3).  The Introduction will also describe other AFM approaches that are on the horizon (e.g., combining optical detection with AFM measurements) but have not yet been completely realized in biological systems.