Andany, S. H. | Search bioRxiv

bioRxiv2025

Separate-scan atomic force microscope for fast infrared scattering-type scanning near-field optical microscope

Pseudo-heterodyne scattering-type scanning near-field optical microscopy (sSNOM) is applied in the mid-infrared region to detect the chemical composition of biomolecules on the nanoscale. However, the application of sSNOM in molecular biology has been limited to static images in air. Recently, bottom illumination sSNOM (BI-sSNOM) was developed for operation in water. Yet, the scan rate of sSNOM remains a bottleneck to record protein structural changes in aqueous solution on the seconds time scale. We designed an optical and mechanical system consisting of a separate scan high-speed atomic force microscope (HS-AFM) coupled to the BI-sSNOM optics. The designed AFM scanner has a mechanical bandwidth of ca 70 kHz along the Z-axis, and ca 6 kHz along the XY-axis, equivalent to the sample scanning HS-AFM. The AFM performance is demonstrated by imaging actin filaments. The optical design is validated by sSNOM experiments on purple membranes and microtubules.

biophysics↗

bioRxiv2025

A hermetically closed sample chamber enables time-lapse nano-characterization of pathogenic microorganisms in-vitro

Pathogenic microorganisms, such as pathogenic mycobacteria, pose a global health burden. Studying these organisms is crucial for gaining detailed knowledge about the pathogens and the diseases they cause. To handle pathogenic organisms, specific biosafety measures appropriate to the virulence of the organism must be fulfilled, most importantly ensuring that all manipulations of pathogenic material are performed within a confined environment. Atomic force microscopy (AFM) is a powerful technique to study biological samples at nanometer-scale resolution, yielding also mechanical properties, all while maintaining physiological conditions. However, standard AFM sample holders do not meet stringent biosafety requirements since they do not constitute a confined system. AFM imaging relies on direct contact between the cantilever and the sample and is sensitive to mechanical interference, rendering conventional containment systems for handling infectious substances inapplicable. Here, we introduce a hermetically sealed AFM sample chamber that meets biosafety demands while satisfying the mechanical and optical constraints of correlated optical microscopy and AFM. We imaged various pathogenic mycobacteria to demonstrate the chambers versatility and effectiveness in containing biohazardous materials. This sample chamber enables high-resolution, time-lapse correlated imaging and biomechanical characterization of pathogenic microorganisms in-vitro. It broadens the scope of research with pathogenic microorganisms under safe and controlled conditions.

bioengineering↗