Quantitative and Fast AFM Co-Localized with STED Microscopy in Living Cell Experiments

Atomic force microscopy (AFM) has become an essential tool for investigating a variety of samples with nanoscale resolution under physiological conditions. Recently, in addition to topographic measurements, information about the interaction and mechanical properties, like adhesion and elasticity, has been extracted. Furthermore, novel developments focus on fast AFM and high-speed AFM to increase the temporal resolution. The integration of AFM with optical microscopy has increased the number of applications, particularly when specificity is required. In the last decade, super-resolution light microscopy has become a popular technique that can be paired with AFM for the imaging of specifically labeled structures with nanoscale resolution. One of many super-resolution microscopy techniques is that of stimulated emission depletion, or STED.

The performance, reliability, and benefits of a combined AFM and STED show that correlative nanoscopy is a powerful tool for biological applications. As a result of the ability to perform measurements simultaneously. Correlative microscopy provides supplementary information and improves data. Super-resolution light microscopy provides specificity while ADM delivers true surface visualization in 3D, as well as nanomechanical information. The possibility to perform full simultaneous measurements was demonstrated in nanomanipulation experiments. Depicted measurements on living human skin fibroblasts are the first examples of an almost unlimited array of applications for the new and innovative correlative technique.

Readers can expect to learn about:

• The limitations of AFM and super-resolution microscopy when used separately;
• How AFM with STED overcomes the resolution mismatch between AFM and conventional (diffraction-limited) light microscopy; and
• The supplemental information and improved data provided by a correlative approach.

_{KEYWORDS: Corrleative Nanoscopy; Microtubules; STED; Super-Resolution}