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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.
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