Cell Property Determination from the Acoustic Microscope Generated Voltage Versus Frequency Curves

Abstract

Among the methods for the determination of mechanical properties of living cells acoustic microscopy provides some extraordinary advantages. It is relatively fast, of excellent spatial resolution and of minimal invasiveness. Sound velocity is a measure of the stiffness or Young's modulus of the cell. Attenuation of cytoplasm is a measure of supramolecular interactions. These parameters are of crucial interest for studies of cell motility, volume regulations and to establish the functional role of the various elements of the cytoskeleton. Using a phase and amplitude sensitive modulation of a scanning acoustic microscope (Hillman et al., 1994, J. Alloys Compounds. 211/212:625-627) longitudinal wave speed, attenuation and thickness profile of a biological cell are obtained from the voltage versus frequency or V(f) curves. A series of pictures, for instance in the frequency range 980-1100 MHz with an increment of 20 MHz, allows the experimental generation of V(f) curves for each pixel while keeping the lens-specimen distance unchanged. Both amplitude and phase values of the V(f) curves are used for obtaining the cell properties and the cell thickness profile. The theoretical analysis shows that the thin liquid layer, between the cell and the substrate, has a strong influence on the reflection coefficient and should not be ignored during the analysis. Cell properties, cell profile and the thickness of the thin liquid layer are obtained from the V(f) curves by the simplex inversion algorithm. The main advantages of this new method are that imaging can be done near the focal plane, therefore an optimal signal to noise ratio is achieved, no interference with Rayleigh waves occurs, and the method requires only an approximate estimate of the material properties of the solid substratum where the cells are growing on.

An acoustic microscopy technique reveals hidden morphological defenses inDaphnia

“…Detailed theory is available to relate the elastic (mechanical) properties of the surface to the contrast, and this enables informed interpretation of the acoustic images to be made (17). This modus operandi provides excellent possibilities for probing mechanical properties of biological specimens, e.g., cells and cell compounds (18,19). Crucial advantages of acoustic microscopy are that it is nondestructive and the results are not altered by specific preparation methods, such as embedding or sectioning.…”

Quantifying Micro-mechanical Properties of Soft Biological Tissues with Scanning Acoustic Microscopy

“…There are few reported studies carried out on unfixed soft biological tissue sections and there is currently no ideal method to relate the high resolution acoustic information to the quantitative mechanical properties of the soft tissues. The authors have previously used the V(f) method (recording of voltage output vs. acoustic frequency) [12], developed by Kundu et al [13], which was thought suitable for the study of soft biological tissues because it is based on the interference between different reflections from a layered media. However, the sound waves reflected from the specimen also interfere with the stray echoes inside the lens, and this interference produces a voltage signal which is strongly dependent on acoustic frequency and the distance between the lens and the specimen.…”

Mapping the Micromechanical Properties of Cryo-sectioned Aortic Tissue with Scanning Acoustic Microscopy

“…However, this technique is not suitable for soft biological specimens because it requires a very smooth specimen surface and due to the rapid attenuation of Rayleigh waves in tissue. The V(f) methodology (recording of voltage output vs. signal frequency) was developed by Kundu et al [17,18] and is more suited to soft biological tissues because it allows quantitative measurements to be made from full images of the tissue. Crucially, for rough biological specimens the method accounts for variations in surface topography.…”

“…The acoustic wave speed cannot be solved analytically and an optimization method based on the Simplex algorithm is used to determine the best solution in terms of specimen thickness, attenuation and density. [17] The SAM images were analysed using STAN (Soft Tissue Analysis) software developed at Aarhus University Hospital (Denmark), which is based on underlying DOS software developed by Kundu [19]. Ten line measurements were made which spanned the entire cross-section of an aorta section, from the adventitial to the intimal layer.…”