Atomic force microscopy (AFM) operation in liquid environments presents unique challenges that differ significantly from measurements in air or vacuum. The presence of liquid introduces complications such as thermal drift, meniscus forces, and probe damping, which can degrade image resolution and measurement accuracy. However, overcoming these challenges enables critical applications in biological samples and electrochemical systems, where in situ characterization is essential.

Thermal drift is a major concern when operating AFM in liquid. Temperature fluctuations cause expansion or contraction of the microscope components and the sample stage, leading to lateral and vertical drift. In liquid cells, thermal equilibration is slower due to the higher heat capacity of liquids compared to air. This drift can distort images and make long-term measurements unreliable. Solutions include using thermally stable materials for the AFM setup, such as low-expansion alloys, and allowing sufficient time for temperature equilibration before imaging. Active temperature control systems, such as feedback-stabilized heating or cooling stages, can further minimize drift. Additionally, closed-loop scanner systems compensate for positional errors in real time, improving stability.

Meniscus forces arise from the interaction between the AFM probe and the liquid surface. When the probe enters or exits the liquid, a meniscus forms around the cantilever, creating unwanted forces that affect deflection measurements. These forces can lead to artifacts in height measurements and reduce probe sensitivity. To mitigate this, hydrophobic or hydrophilic coatings on the cantilever can be tailored to match the liquid properties, minimizing meniscus formation. Using shorter cantilevers reduces the interaction area, thereby decreasing meniscus effects. Another approach involves operating the probe fully submerged, eliminating air-liquid interfaces entirely.

Probe damping in liquid is caused by the higher viscosity of liquids compared to air, which reduces the quality factor (Q-factor) of the cantilever resonance. A low Q-factor results in slower response times and decreased sensitivity to tip-sample interactions. Solutions include using higher-frequency cantilevers designed for liquid operation, which maintain better oscillation stability. Tuning the drive frequency to match the damped resonance peak improves signal-to-noise ratio. Additionally, specialized feedback algorithms adjust for damping effects in real time, preserving imaging performance.

Biological samples benefit significantly from AFM imaging in liquid, as it maintains physiological conditions and prevents dehydration. For example, live cell imaging allows observation of dynamic processes such as membrane remodeling or cytoskeletal rearrangements. AFM can measure mechanical properties like stiffness and adhesion forces, providing insights into cell health or disease states. However, challenges include sample softness and mobility, which require gentle imaging modes such as tapping mode or fast-scanning techniques to avoid damage. Functionalized tips enable molecular recognition studies, mapping specific receptors or ligands on cell surfaces with nanoscale precision.

In electrochemical systems, AFM in liquid enables real-time observation of processes at electrode-electrolyte interfaces. Applications include studying nucleation and growth of electrodeposited materials, corrosion mechanisms, and solid-electrolyte interphase formation in batteries. The AFM probe must be electrically insulated to avoid interference with electrochemical measurements, except in conductive AFM modes where the tip acts as a nanoelectrode. Challenges include maintaining stable electrochemical conditions while scanning, as local potential gradients can influence tip-sample interactions. Using potentiostatic control synchronizes AFM and electrochemical measurements, ensuring accurate correlation between structural changes and electrochemical activity.

Quantitative mechanical measurements in liquid require careful calibration of the cantilever spring constant and sensitivity, as liquid damping affects both. Thermal tuning methods are commonly used to determine the spring constant in situ. For force spectroscopy, accounting for hydrodynamic drag forces is essential to accurately interpret adhesion or indentation data. Advanced models incorporate viscous drag effects when calculating interaction forces from deflection data.

In summary, AFM operation in liquid demands specialized approaches to address thermal drift, meniscus forces, and probe damping. Solutions involving hardware modifications, probe design, and advanced feedback algorithms enable high-resolution imaging and force measurements. Biological applications leverage AFM’s ability to probe soft samples in native environments, while electrochemical studies benefit from real-time interfacial characterization. By overcoming these challenges, AFM remains a powerful tool for nanoscale investigation in liquid-phase systems.