We propose a new analytical approach combining vibrational spectroscopy and acoustic tomography for the detection and characterization of vesicles inside Streptomyces bacteria. Using atomic force microscopy and infrared spectroscopy (AFM-IR), we detect the presence of triglyceride vesicles. Their sizes in depth are measured with high accuracy using mode synthesizing atomic force microscopy (MS-AFM). We conducted a comparative study of AFM-IR and MS-AFM, and highlighted the advantages of the coupling of these techniques in having a full characterization (chemical, topographical, and volumetric) of a biological sample. With these complementary techniques, a complete access to the vesicle size distribution has been achieved with an accuracy of less than 50 nm. A 3D reconstruction of bacteria showing the in-depth distribution of vesicles is given to underline the great potential of the acoustic method.

Challenges in nanoscale characterization call for non-invasive, yet sensitive subsurface characterization of low-density materials such as polymers. In this work, we present new evidence that mode-synthesizing atomic force microscopy can be used to detect minute changes in low-density materials, such as those engendered in electro-sensitive polymers during electron beam lithography, surpassing all common nanoscale mechanical techniques.

Moreover, we propose 3D reconstruction of the exposed polymer regions using successive high-resolution frames acquired at incremental depths inside the sample. In addition, the results clearly show the influence of increasing dwell time on the depth profile of the nano-sized exposed regions. Hence, the simple approach described here can be used for achieving sensitive nanoscale tomography of soft materials with promising applications in material sciences and biology.