An alternative quantitative acoustical and electrical method for detection of cell adhesion process in real-time.

Sauerbrey [(1956), Z Phys 55:206-222] showed that the shift in resonance frequency of thickness shear mode (TSM) of a quartz crystal sensor is proportional to the mass, which is deposited on it. However, new powerful electrical circuits were developed that are capable of operating TSM quartz crystal sensors in fluids which enabled this method to be introduced into electrochemical and biological applications. These applications include the detection of virus capsids, bacteria, mammalian cells, the interaction of DNA and RNA with complementary strands, specific recognition of protein ligands by immobilized receptors, and last but not least the study of complete immunosensors. Piezoelectric quartz transducers allow a label-free identification of molecules; they are more than mass sensors since the biosensor response is also influenced by the surface charge of adsorbed proteins, interfacial phenomena, surface roughness and viscoelastic properties of the adhered biomaterial. These new characteristics have recently been used to investigate cell, liposome, and protein adhesion onto surfaces, thus permitting the rapid determination of morphological cell changes as a response to pharmacological substances, and changes in the water content of biopolymers avoiding of time-consuming methods. We validated an alternative quantitative acoustical engineering for cell adhesion process monitored by the TSM. Shear acoustical results (motional resistance) are further correlated to cell counting procedures and are sensitive of adhesion processes in real-time.

Additive effect of RGD coating to functionalized titanium surfaces on human osteoprogenitor cell adhesion and spreading.

Titanium-based biomaterials for endosseous implants have found widespread applications in the orthopedic, maxillofacial, and dental domains. Indeed, the surface characteristics such as their chemical modification control considerably the cellular response and, subsequently, the quality and the quantity of new-formed bone around the implant. In this study, human osteoprogenitor (HOP) cell adhesion on different titanium surfaces functionalized with hydroxyapatite (HA), type I collagen, or Arg-Gly-Asp (RGD)-containing peptides is investigated by the quartz crystal resonators and by confocal laser scanning microscopy (CLSM) for the imaging of focal contact formation. Data obtained by quartz crystal resonator technique revealed that RGD-containing peptides alone increase HOP cell adhesion in early time period of culture. Moreover, association of RGD-containing peptides with either type I collagen or with HA layers induces an additive effect on HOP cell adhesion compared to Ti-Coll or Ti-HA. CLSM shows both the area of focal contact by cell unit and the cytoskeleton network organization to differ according to the surfaces. Interestingly, association of RGD-containing peptides with HA layers induces an additive effect on focal contact formation on HOP cells compared to Ti-HA alone. These data confirm that an RGD peptide effect occurs in the early time of culture, which is beneficial for osteoblast to spreading, differentiation, and survival.

Progress towards in vitro quantitative imaging of human femur using compound quantitative ultrasonic tomography.

The objective of this study is to make cross-sectional ultrasonic quantitative tomography of the diaphysis of long bones. Ultrasonic propagation in bones is affected by the severe mismatch between the acoustic properties of this biological solid and those of the surrounding soft medium, namely, the soft tissues in vivo or water in vitro. Bone imaging is then a nonlinear inverse-scattering problem. In this paper, we showed that in vitro quantitative images of sound velocities in a human femur cross section could be reconstructed by combining ultrasonic reflection tomography (URT), which provides images of the macroscopic structure of the bone, and ultrasonic transmission tomography (UTT), which provides quantitative images of the sound velocity. For the shape, we developed an image-processing tool to extract the external and internal boundaries and cortical thickness measurements. For velocity mapping, we used a wavelet analysis tool adapted to ultrasound, which allowed us to detect precisely the time of flight from the transmitted signals. A brief review of the ultrasonic tomography that we developed using correction algorithms of the wavepaths and compensation procedures are presented. Also shown are the first results of our analyses on models and specimens of long bone using our new iterative quantitative protocol.

Monitoring cell adhesion processes on bioactive polymers with the quartz crystal resonator technique.

The Thickness Shear Mode (TSM) quartz crystal resonator has been extensively used as sensitive sensor in various electrochemical and biological applications. This technique based on the propagation of an ultrasonic shear wave generated by a sinusoidal electric field through a piezoelectric quartz resonator, provides a non-destructive and powerful means to probe changes at solid-solid or solid-liquid interfaces. In this study, TSM was used to characterize cell-polymer interactions developing during the cell adhesion process. TSM sensing was used to monitor the inhibiting properties of bioactive polymers towards fibroblast McCoy adhesion processes. For this purpose, thin films of various bioactive polymers exhibiting either carboxylate or/and sulfonate functional groups were deposited onto the TSM. Measurements of the time variation of the electrical motional resistance in the vicinity of the mechanical sensor resonant frequency were performed as the quartz crystal resonator was either coated with the continuous polymer phase or polymer plus cell suspensions. Cell adhesion processes on these surfaces was investigated by cell counting and the quartz resonator-based technique. Inhibition of fibroblast McCoy adhesion onto thin polymer films of various chemical compositions was analyzed and discussed in the perspective of a possible application of these bioactive polymers to fabricate intraocular lenses able to prevent secondary cataract phenomena.

Chirp-Z analysis for sol-gel transition monitoring.

Gelation is a complex reaction that transforms a liquid medium into a solid one: the gel. In gel state, some gel materials (DMAP) have the singular property to ring in an audible frequency range when a pulse is applied. Before the gelation point, there is no transmission of slow waves observed; after the gelation point, the speed of sound in the gel rapidly increases from 0.1 to 10 m/s. The time evolution of the speed of sound can be measured, in frequency domain, by following the frequency spacing of the resonance peaks from the Synchronous Detection (SD) measurement method. Unfortunately, due to a constant frequency sampling rate, the relative error for low speeds (0.1 m/s) is 100%. In order to maintain a low constant relative error, in the whole speed time evolution range, Chirp-Z Transform (CZT) is used. This operation transforms a time variant signal to a time invariant one using only a time dependant stretching factor (S). In the frequency domain, the CZT enables us to stretch each collected spectrum from time signals. The blind identification of the S factor gives us the complete time evolution law of the speed of sound. Moreover, this method proves that the frequency bandwidth follows the same time law. These results point out that the minimum wavelength stays constant and that it only depends on the gel.

Compressibility of nano inclusions in complex fluids by ultrasound velocity measurements.

We present a high precision ultrasonic velocimeter for a small volume sample (1 cm3) for a path length of 1 cm achieved. The method used is based on the time of flight measurement with an original signal processing technique: the barycenter method. With our system, we have measured the sound velocity with an accuracy of 10(-5). The detection of a difference in velocity between two liquids of about 2 cm/s is achieved. The compressibility of the reference liquid can then be deduced with an accuracy better than 0.2%. Using this custom-made system, we have studied and characterized complex fluids, systems biomimetic of biological membranes, as well as proteins included in nanometric water droplets. Under these experimental conditions, we have reached the value of protein compressibilities with an accuracy better than 10%.