Analysis of the composite response of shear wave resonators to the attachment of mammalian cells

The suitability of the quartz crystal microbalance (QCM) technique for monitoring the attachment and spreading of mammalian cells has recently been established. Different cell species were shown to generate an individual response of the QCM when they make contact with the resonator surface. Little is known, however, about the underlying mechanisms that determine the QCM signal for a particular cell type. Here we describe our results for different experimental approaches designed to probe the particular contributions of various subcellular compartments to the overall QCM signal. Using AC impedance analysis in a frequency range that closely embraces the resonators' fundamental frequency, we have explored the signal contribution of the extracellular matrix, the actin cytoskeleton, the medium that overlays the cell layer, as well as the liquid compartment that is known to exist between the basal plasma membrane and the culture substrate. Results indicate that the QCM technique is only sensitive to those parts of the cellular body that are involved in cell substrate adhesion and are therefore close to the resonator surface. Because of its noninvasive nature, sensitivity, and time resolution, the QCM is a powerful means of quantitatively studying various aspects of cell-substrate interactions.     

The quartz crystal microbalance as a novel means to study cell-substrate interactions In situ

The quartz crystal microbalance (QCM) was first introduced as a mass sensor in gas phase and in vacuum. Since oscillator circuits capable of exciting shear vibrations of quartz resonators under liquid loading have been developed, the QCM became accepted as a new, powerful technique to follow adsorption processes at solid-liquid interfaces in chemical and biological research. Lately, the QCM technique has attracted considerable interest as a novel means to monitor cell-substrate interactions of mammalian cells in vitro. Because the establishment and modulation of cell-substrate contacts is important for many physiological processes, and potent techniques to measure these interactions noninvasively are rare, the present review highlights applications of the QCM technique in this field. The suitability of the QCM device to monitor attachment and spreading of mammalian cells in real time has been well established. The QCM response is dependent on the individual cell type that is examined. In order to identify the sources for these cell-type-specific results of QCM readings, and to understand the information content of the signal, attempts have been made to decompose the overall QCM response into subcellular contributions. The aforementioned subjects, together with a condensed introduction into the QCM technology, are included in this article.     

Monitoring of integrin-mediated adhesion of human ovarian cancer cells to model protein surfaces by quartz crystal resonators: evaluation in the impedance analysis mode

The quartz crystal microbalance (QCM) was used to monitor specific, integrin-mediated adhesion of human ovarian cancer cells to distinct extracellular matrix (ECM) proteins immobilized on gold-coated quartz crystal resonators. The QCM was operated in the impedance analysis mode, where frequency shift as well as bandwidth are accessible on a broad range of overtones. The increase in bandwidth caused by covering the quartz resonator with cells was reproducible and largely independent of overtone order, whereas the frequency shift displayed some variability. Thus the bandwidth proved to be the more robust parameter for sensing cell adhesive events. The bandwidth increased in proportion to the number of seeded cells to the quartz crystal as long as the number was below 150,000 cells/ml. Comparing the resonance parameters on different harmonics, one finds that viscoelastic modeling with homogeneous layer systems cannot reproduce the results: lateral heterogeneity has to be taken into account. The differences in adhesive strength of human ovarian cancer cells towards selected ECM proteins monitored by QCM was in good agreement with data obtained by conventional cell adhesion assays. Strong cell adhesion was observed to the ECM proteins vitronectin (VN) and fibronectin (FN), while only weak attachment occurred on laminin. In order to prove specific, integrin alphavbeta3-mediated cell adhesion to its ligands FN and VN, the cyclic integrin alphavbeta3-directed peptide c(RGDfV) was used as competitor and significantly reversed cell adhesion. Since integrin interaction with ECM proteins is dependent on the presence of bivalent cations, cell detachment was also seen after treatment of cell monolayers with the chelator ethylene-dinitro-tetra-acetic acid (EDTA). The QCM technique is a reliable method to monitor cell adsorption to ECM-pretreated surfaces in real time. It may be an alternative tool for screening specific and selective antagonists of integrin/ECM interaction.     

The quartz crystal microbalance as a continuous monitoring tool for the study of endothelial cell surface attachment and growth

The quartz crystal microbalance (QCM) was used to monitor endothelial cell (EC) adhesion on the gold surface of an oscillating quartz crystal contained in a QCM device. A number of parameters were investigated. First, we observed differential QCM O-ring toxicities for ECs. Second, appropriate conditions for cell culture and QCM cell environment were identified that can eliminate large-scale frequency oscillations in the measurements. These artifacts are not due to added cells but originate in the time-dependent evaporation of water. Having eliminated these artifacts, we then demonstrated that the measured steady-state crystal frequency shift, Delta f, and motional resistance shift, DeltaR, were determined by the number of firmly attached ECs requiring trypsinization from the crystal surface. Last, following steady-state attachment of ECs, the EC growth stimulation by fibroblast growth factor was monitored in a continuous fashion by measuring f and R values over a 72 h. period. We observed the Delta f values to increase in a way that reflected the increase in EC number bound to the QCM surface. Following addition of ECs to the QCM, the time-dependent increase in DeltaR can be interpreted in terms of increase by the ECs of the energy dissipation properties of the solution at the solution-gold surface interface. This effect is due to their rapid surface attachment and the elaboration of their cytoskeletal properties. These results indicate that the QCM technique can be used for the study of EC attachment and growth and suggest its potential for the real time study of per unit surface area cell mass distribution dynamics and viscoelastic properties and the cells' responses to stresses or perturbations brought about using biologically active molecules.     

Specific adhesion of vesicles monitored by scanning force microscopy and quartz crystal microbalance

The specific adhesion of unilamellar vesicles with an average diameter of 100 nm on functionalized surfaces mediated by molecular recognition was investigated in detail. Two complementary techniques, scanning force microscopy (SFM) and quartz crystal microbalance (QCM) were used to study adhesion of liposomes consisting of 1, 2-dipalmitoyl-sn-glycero-3-phosphocholine and varying concentrations of N-((6-biotinoyl)amino)hexanoyl)-1, 2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine (biotin-X-DHPE). Monitoring the adhesion of the receptor-doped vesicles to avidin-coated gold surfaces by QCM (f(0) = 5 MHz) revealed an increased shift in resonance frequency with increasing biotin concentration up to 10 mol% biotin-X-DHPE. To address the question of how the morphology of the liposomes changes upon adhesion and how that contributes to the resonator's frequency response, we performed a detailed analysis of the liposome morphology by SFM. We found that, with increasing biotin-concentration, the height of the liposomes decreases considerably up to the point where vesicle rupture occurs. Thus, we conclude that the unexpected high frequency shifts of the quartz crystal (>500 Hz) can be attributed to a firm attachment of the spread bilayers, in which the number of contacts is responsible for the signal. These findings are compared with one of our recent studies on cell adhesion monitored by QCM.     

Quartz crystal microbalance study of endothelial cell number dependent differences in initial adhesion and steady-state behavior: evidence for cell-cell cooperativity in initial adhesion and spreading

The quartz crystal microbalance (QCM) technique has been applied to the real time monitoring of endothelial cell (EC) adhesion and spreading on the QCM gold surface. We previously showed that the measured QCM Deltaf and DeltaR shifts were due to cells adhering to the gold crystal surface, requiring proteolytic enzyme treatment to be removed from the surface, in order for the Deltaf and DeltaR shifts to return to zero. In the present report, we demonstrate the quantitative dependence and saturation of the measured Deltaf and DeltaR shifts on the number of firmly attached ECs as measured by electronic counting of the cells. We demonstrate through a light microscope simulation experiment that the different Deltaf and DeltaR regions of the QCM temporal response curve correspond to the incident ECs contacting the surface, followed by their adhesion and spreading, which reflect cellular mass distribution and cytoskeletal viscoelasticity changes. Also, we demonstrate that the dose response curve of Deltaf and DeltaR values versus attached EC number is more sensitive and possesses less scatter for the hydrophilically treated surface compared to the native gold surface of the QCM. For both surfaces, a Deltaf and DeltaR versus trypsinized, attached EC number plot 1 h post-seeding exhibits a sigmoid curve shape whereas a similar plot 24 h post-seeding exhibits a hyperbolic curve shape. This number dependence suggests cell-cell cooperativity in the initial cell adhesion and spreading processes. These QCM data and our interpretation are corroborated by differences in cell appearance and spreading behavior we observed for ECs in a light microscope fluorescence simulation experiment of the cell density effect. For a stably attached EC monolayer at 24 h post-addition, steady-state Deltaf and DeltaR values are higher and exhibit saturation behavior for both the hydrophilically treated gold surface as compared to the untreated surface. The steady-state 24 h Deltaf and DeltaR values of stably attached ECs are shifted from the 1 h attached ECs. The 24 h values are characteristic of a more energy-dissipative structure. This is consistent with the time-dependent elaboration of surface contacts in anchorage-dependent ECs via the attachment of intregrins to underlying extracellular matrix. It is also in agreement with the known energy dissipation function of the ECs that cover the interior of blood vessels and are exposed to continuous pulsatile blood flow.     

Dynamic measurement of the surface stress induced by the attachment and growth of cells on Au electrode with a quartz crystal microbalance

The processes of adhesion, spreading and proliferation of human mammary cancer cells MCF-7 on two Au electrodes with different surface roughness (R(f) and R(f)=3.2 or 1.1) were monitored and clearly identified with the quartz crystal microbalance (QCM) technique. Analyses of the QCM responses on the resonant frequency shifts (Deltaf(0)) vs. the motional resistance changes (DeltaR(1)) revealed a significant surface-stress effect in the involved courses, in addition to a viscodensity effect and a relatively small mass effect (especially at the smooth electrode). Experiments of fluorescence microscopy, cyclic voltammetry and electrochemical impedance spectroscopy were conducted to investigate the cell population on the electrode vs. the electrode-surface roughness. Simplified equations are deduced to quantitatively evaluate the surface stress, and a novel QCM method for dynamically measuring the surface stress on an electrode in cell-culture course is thus described. It was found that the smoother surface (R(f)=1.1) gave a higher surface stress during cell attachment and less cell population on it than the rougher surface (R(f)=3.2). In addition, real-time QCM monitoring showed on the same electrode the surface stress induced by hepatic normal cells being notably higher than that caused by hepatic cancer cells at cell-attachment stage, suggesting that the surface-stress measurement can exhibit the difference of adhesion-performance between the healthy and ill-behaved cells.     

Adhesion of eukaryotic cell lines on the gold surface modified with extracellular matrix proteins monitored by the piezoelectric sensor

The piezoelectric sensor (quartz crystal microbalance, QCM) was used to monitor cell adhesion in real time. Two cell lines, rat epithelial cells (WB F344) and lung melanoma cells (B16F10) were used. The cells were adhered and grown on the gold surface of the sensor pre-coated with adsorbed layer of extracellular matrix proteins as vitronectin and laminin. The process of cell attachment and spreading on the gold surface was continuously monitored and displayed by changes of the resonant frequency Deltaf and resistance DeltaR values of the piezoelectric resonators. The initial phase of cell attachment and spreading induced a decrease of frequency and increase of resistance relating viscoelastic properties of the cell monolayer on the sensing surface. The steady-state of both shifts was achieved after a few hours. The presence and state of cells on the surface was confirmed by fluorescent microscopy. The obtained results demonstrate that the piezoelectric sensor is suitable for studies of the cell adhesion processes. Thus obtained cell-based biosensor has potential for identification and screening of biologically active drugs and other biomolecules affecting cellular shape and attachment.     

Quartz crystal microbalance: a useful tool for studying thin polymer films and complex biomolecular systems at the solution-surface interface

The quartz crystal microbalance (QCM) is a simple, cost effective, high-resolution mass sensing technique, based upon the piezoelectric effect. As a methodology, the QCM evolved a solution measurement capability in largely analytical chemistry and electrochemistry applications due to its sensitive solution-surface interface measurement capability. The technique possesses a wide detection range. At the low mass end, it can detect monolayer surface coverage by small molecules or polymer films. At the upper end, it is capable of detecting much larger masses bound to the surface. These can be complex arrays of biopolymers and biomacromolecules, even whole cells. In addition, the QCM can provide information about the energy dissipating properties of the bound surface mass. Another important and unique feature of the technique is the ability to measure mass and energy dissipation properties of films while simultaneously carrying out electrochemistry on solution species or upon film systems bound to the upper electrode on the oscillating quartz crystal surface. These measurements can describe the course of electropolymerization of a film or can reveal ion or solute transport within a film during changes in the film environment or state, including the oxidation state for an electroactive film driven by the underlying surface potential. The past decade has witnessed an explosive growth in the application of the QCM technique to the study of a wide range of molecular systems at the solution-surface interface, in particular, biopolymer and biochemical systems. In this report, we start with a brief historical and technical overview. Then we discuss the application of the QCM technique to measurements involving micellar systems, self-assembling monolayers and their phase transition behavior, molecularly imprinted polymers, chemical sensors, films formed using the layer-by-layer assembly technique, and biopolymer films and point out the utility of the electrochemical capabilities of the technique to characterizing film properties, especially electroactive polymer films. We also describe the wide range of surface chemistries and attachment strategies used by investigators to bring about surface attachment and multi-layer interactions of these thin film systems. Next we review the wide range of recent applications of the technique to: studies of complex biochemical and biomimetic systems, the creation of protein and nucleic acid biosensors, studies of attached living cells and whole cell biosensor applications. Finally, we discuss future technical directions and applications of the QCM technique to areas such as drug discovery.     

Application of parylene-coated quartz crystal microbalance for on-line real-time detection of microbial populations

A novel technique of applying a quartz crystal microbalance (QCM) sensor to the on-line real-time detection of microbial populations is described. The pQCM sensor was fabricated by depositing di-para-xylene (parylene) over the entire surface of a QCM sensor through a chemical vapor deposition (CVD) process. An electrically insulated film of parylene on the QCM sensor enabled the operation of the sensor in the liquid environment, and the resonance frequency of the pQCM sensor set in the medium of a cultivation flask shifted in response to the microbial population. The effects of pH, conductivity, and viscosity of the medium on the frequency shift of the pQCM sensor were investigated. Ignorable responses (less than 1% at 10(3)cells) were obtained during an incubation cycle. The detection limit of the pQCM sensor was identified as 10(2) cells ml(-1) with a frequency shift of around 2 x 10(3)Hz. The cell numbers of Escherichia coli cultivated in both the YEM medium and whole milk were detected. A satisfactory correlation (r(2)=0.95) was obtained between the cell number and the response of the pQCM sensor. Experimental results suggest that the pQCM described here is applicable to the continuous long-term detection of microbial populations during a fermentation process.     

A quartz crystal microbalance cell biosensor: detection of microtubule alterations in living cells at nM nocodazole concentrations

The quartz crystal microbalance (QCM) was used to create a piezoelectric biosensor utilizing living endothelial cells (ECs) as the biological signal transduction element. ECs adhere to the hydrophilically treated gold QCM surface under growth media containing serum. At 24 h following cell addition, calibration curves were constructed relating the steady state Δf and ΔR shift values observed to the numbers of electronically counted cells requiring trypsinization to be removed from the surface. We then utilized this EC QCM biosensor for the detection of the effect of [nocodazole] on the steady state Δf and ΔR shift values. Nocodazole, a known microtubule binding drug, alters the cytoskeletal properties of living cells. At the doses used in these studies (0.11–15 μM), nocodazole, in a dose dependent fashion, causes the depolymerization of microtubules in living cells. This leads a monolayer of well spread ECs to gradually occupy a smaller area, lose cell to cell contact, exhibit actin stress fibers at the cell periphery and acquire a rounded cell shape. We observed the negative Δf shift values and the positive ΔR shift values to increase significantly in magnitude over a 4-h incubation period following nocodazole addition, in a dose dependent fashion, with a transition midpoint of 900 nM. Fluorescence microscopy of the ECs, fixed on the gold QCM surface and stained for actin, demonstrated that the shape and cytoskeleton of ECs were affected by as little as 330 nM nocodazole. These results indicate that the EC QCM biosensor can be used for the study of EC attachment and to detect EC cytoskeletal alterations. We suggest the potential of this cellular biosensor for the real time identification or screening of all classes of biologically active drugs or biological macromolecules that affect cellular attachment, regardless of their molecular mechanism of action.     

Application of peptide probe for evaluating affinity properties of proteins using quartz crystal microbalance

This study proved a possibility of a peptide probe for evaluating affinity properties of proteins. We have designed and synthesized three different peptide probes, H-Ala3-(Gly-Pro5)3-Gly-OH (peptide A), H-Ala3-(Gly-Pro5)-Gly-OH (peptide B) and H-Ala3-Gly-OH (peptide C) for testing their affinities to profilin. Each peptide probe was immobilized on a quartz crystal microbalance (QCM) sensor. The QCM sensor with the peptide A showed a 93 Hz decrease of resonant frequency which indicated profilin bound to the QCM sensor in a single layer. In a successive reaction with actin, the QCM analysis resulted in a 123 Hz decrease of resonant frequency which showed actin bound to the QCM sensor. A fluorescence microscope image of the sensor surface exhibited clear fluorescence after binding a rhodamine labeled actin on the sensor surface. These results supported stepwise reactions of profilin binding to the peptide A and actin binding to profilin. In the three peptide probes, the peptide A showed the highest affinity to profilin, i.e., sequence dependent affinity was confirmed.     

Effects of extremely low-frequency pulsed electromagnetic fields (ELF-PEMFs) on glioblastoma cells (U87)

The impact of extremely low-frequency pulsed electromagnetic fields (ELF-PEMFs) at various frequencies and amplitudes was investigated on cell cycle, apoptosis and viability of the Glioblastoma Multiforme (GBM) cell line (U87), in vitro. The GBM is a malignant brain tumor with high mortality in humans and poorly responsive to the most common type of cancer treatments, such as surgery, chemotherapy and radiation therapy. U87 cells with five experimental groups (I–V) were exposed to various ELF-PEMFs for 2, 4 and 24 h, as follows: (I) no exposure, control; (II) 50 Hz 100 ± 15 G; (III) 100 Hz 100 ± 15 G; (IV) 10 Hz 50 ± 10 G; (V) 50 Hz 50 ± 10 G. The morphology properties, cell viability and gene expression of proteins involved in cell cycle regulation (Cyclin-D1 and P53) and apoptosis (Caspase-3) were investigated. After 24 h, the cell viability and Cyclin-D1 expression increased in Group II (30%, 45%), whereas they decreased in Groups III (29%, 31%) and IV (21%, 34%); P53 and Caspase-3 elevated only in Group III; and no significant difference was observed in Group V, respectively, compared with the control (p < 0.05). The data suggest that the proliferation and apoptosis of human GBM are influenced by exposure to ELF-PEMFs in different time-dependent frequencies and amplitudes. The fact that some of the ELF-PEMFs frequencies and amplitudes favor U87 cells proliferation indicates precaution for the use of medical devices related to the MFs on cancer patients. On the other hand, some other ELF-PEMFs frequencies and intensities arresting U87 cells growth could open the way to develop novel therapeutic approaches.     

Acoustic detection of cell adhesion to a coated quartz crystal microbalance – implications for studying the biocompatibility of polymers

Biocompatibility of polymers is an important parameter for the successful application of polymers in tissue engineering. In this work, quartz crystal microbalance (QCM) devices were used to follow the adhesion of NIH 3T3 fibroblasts to QCM surfaces modified with fibronectin (FN) and poly-D -lysine (PDL). The variations in sensor resonant frequency (Δf) and motional resistance (ΔR), monitored as the sensor signal, revealed that cell adhesion was favored in the PDL-coated QCMs. Fluorescence microscopy images of seeded cells showed more highly spread cells on the PDL substrate, which is consistent with the results of the QCM signals. The sensor signal was shown to be sensitive to extracellular matrix (ECM)-binding motifs. Ethylenediaminetetraacetic acid (EDTA) and soluble Gly-Arg-Gly-Asp-Ser (GRGDS) peptides were used to interfere with cell-ECM binding motifs onto FN-coated QCMs. The acquired acoustic signals successfully showed that in the presence of 30 mM EDTA or 1 mM GRGDS, cell adhesion is almost completely abolished due to the inhibition/blocking of integrin function by these compounds. The results presented here demonstrate the potential of the QCM sensor to study cell adhesion, to monitor the biocompatibility of polymers and materials, and to assess the effect of adhesion modulators. QCM sensors have great potential in tissue engineering applications, as QCM sensors are able to analyze the biocompatibility of surfaces and it has the added advantage of being able to evaluate, in situ and in real time, the effect of specific drugs/treatments on cells.     

Magnetic acoustic resonance immunoassay (MARIA): a multifrequency acoustic approach for the non-labelled detection of biomolecular interactions

A unique sensing platform, comprising an electromagnetic field detector and an acoustic resonator, has been used as a wireless system for remote sensing of biorecognition events. The MARS (Magnetic Acoustic Resonator Sensor) technique has proven useful for detecting the formation of protein multilayers derived from specific binding phenomena. The technique enables multifrequency analysis, without the need of electrodes attached to the sensing element, and also facilitates the in situ surface modification of the substrate for antibody attachment. The MARS sensor was utilized as the platform on which a standard immunoassay was carried out. Two different conditions for the attachment of the first antibody to the quartz surface were tested: (i) Adsorption of the antibody onto the surface of a bare quartz disc; (ii) covalent immobilization of the antibody to a chemically modified quartz surface. Both methods can be successfully utilized for the 'label-less' detection of the biorecognition event between goat IgG and anti-goat IgG by analysis of the multifrequency spectrum. Covalent attachment of the primary antibody results in a more efficient immobilization, with higher surface density, and a consistently enhanced response for the binding of the secondary antibody. This approach will be of interest to life scientists and biochemists that require high performance assay methodologies that do not use chemical labels.     

Integrating the QCM detection with magnetic separation for on-line analysis

We investigate the feasibility of coupling the quartz crystal microbalance (QCM) with magnetic separation for on-line analysis. A flow cell was integrated with QCM and magnetic force for the analysis of magnetic and nonmagnetic samples. The resonant frequency change (Deltaf) of QCM was related to the amount of deposited magnetic nanoparticles. This experiment demonstrates that QCM can be used as an on-line detector for magnetic separation. The QCM also gives a characteristic response of the binding between the streptavidin and biotin labeled on the magnetic nanoparticles. Biotin-labeled magnetic nanoparticles were flowed through a gold electrode of QCM to deposit as a matrix for selective capturing streptavidin. The resonant frequency change of QCM was proportional to the amounts of streptavidin captured by biotin. This technique can provide a simple, economic, and automatic method for on-line detection of biomarkers.     

Development of a direct-binding chloramphenicol sensor based on thiol or sulfide mediated self-assembled antibody monolayers

A batch-type antibody-immobilized quartz crystal microbalance (QCM) system for detecting chloramphenicol (CAP) was developed. To bind an anti-CAP antibody onto the gold electrode surface of piezoelectric crystals, self-assembled monolayers (SAMs) of different thiols or sulfides were formed by a chemisorption procedure. Then, the anti-CAP antibody was covalently linked to the pre-formed monolayers by an activation procedure using 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and N-hydroxysulfosuccinimide. The antibody-immobilized QCM chip thus prepared was installed in a well holder and was measured for sensor response. Compared with the bare QCM chip and the QCM chip only coated with 3-mercaptopropionic acid (MPA), the antibody-immobilized sensor showed greatly enhanced frequency shifts by 10-50-fold after CAP injection. In this case, CAP detection which was indicated by steady-state resonant frequency shift was accomplished within 10 min. When CAP solution was injected into the reaction cell in 50mM concentration, the frequency shifts obtained were, respectively, 530 and 505 Hz in case of thiosalicylic acid and MPA immobilization. Repeated use of the sensor chips up to eight times was possible after 1 min regeneration with 0.1M NaOH. This system demonstrated a potential application of thiol or sulfide mediated SAMs as the pre-coatings of a real-time detection on CAP in solution.     

ITO石英晶体电极上不同内皮细胞浓度的粘弹性研究及药物作用评估

用石英晶体微天平(quartz crystal microbalance,QCM)和活细胞成像技术实时监测人脐静脉内皮细胞(HUVEC)在ITO石英晶体电极上的动态粘附响应过程。在ITO晶体电极上加入不同浓度的HUVEC,测定细胞在QCM上谐振频率以及耗散的实时变化。通过ITO电极与光学显微镜的联用,监测了HUVEC在药物处理前后的动态变化过程。用细胞粘弹性指数(QCM的动态电阻变化与频移变化之比,CVI=ΔR/Δf)表征细胞的粘弹性变化,同时通过活细胞成像技术的联用,实时监测细胞的形态变化。结果表明:细胞浓度为10万个/m L时,细胞在ITO电极上铺展完全且粘弹性最大。抑制剂y-27632和激动剂凝血酶thrombin药物处理细胞前后,在显微镜的实时监测下细胞形态变化不明显,但CVI粘弹性指数变化较大,说明QCM信号比光学信号更为敏感,且在药物筛选方面有有很大的应用前景。     

Novel quartz crystal microbalance based biosensor for detection of oral epithelial cell-microparticle interaction in real-time

Recent applications of quartz crystal resonant sensor technology to monitor cell adhesion and specific ligand interaction processes has triggered the development of a new category of quartz crystal microbalance (QCM) based biosensors. In this study human oral epithelial cells (H376) were cultured on quartz sensors and their response to microspheres investigated in situ using the QCM technique. The results demonstrated that this novel biosensor was able to follow cell-microsphere interactions in real-time and under conditions of flow as would occur in the oral cavity. Unique frequency profiles generated in response to the microspheres were postulated to be due to phases of mass addition and altered cellular rigidity. Supporting microscopic evidence demonstrated that the unique frequency responses obtained to these interactions were in part due to binding between the cell surface and the microspheres. Furthermore, a cellular uptake process, in response to microsphere loading was identified and this, by influencing the rigidity of the cellular cytoskeleton, was also detectable through the frequency responses obtained.     

Molecularly imprinted polymers (MIP) in electroanalysis of proteins

The review summarizes current knowledge on the main approaches used for creation of high affinity polymer analogs of antibodies (known as molecularly imprinted polymers, MIP) applicable for electroanalysis of functionally important proteins such as myoglobin, troponin T, albumin, ferritin, lysozyme, calmodulin. The main types of monomers for MIP preparation as well as methods convenient for analysis of MIP/protein interactions, such as surface plasmon resonance (SPR), nanogravimetry with the use of a quartz crystal resonator (QCM), spectral and electrochemical methods have been considered. Special attention is paid to experimental data on electrochemical registration of myoglobin by means of o-phenylenediaminebased MIP electrodes. It was shown that the imprinting factor calculated as a ratio of the myoglobin signal obtained after myoglobin insertion in MIP to the myoglobin signal obtained after myoglobin insertion in the polymer lacking the molecular template (NIP) is 2–4.