Bioanalytical sensors using the heat-transfer method HTM and related techniques.

This review provides an overview on bio- and chemosensors based on a thermal transducer platform that monitors the thermal interface resistance R th between a solid chip and the supernatant liquid. The R th parameter responds in a surprisingly strong way to molecular-scale changes at the solid-liquid interface, which can be measured thermometrically, using for instance thermocouples in combination with a controllable heat source. In 2012, the effect was first observed during on-chip denaturation experiments on complementary and mismatched DNA duplexes that differ in their melting temperature. Since then, the concept is addressed as heat-transfer method, in short HTM, and numerous applications of the basic sensing principle were identified. Functionalizing the chip with bioreceptors such as molecularly imprinted polymers makes it possible to detect neurotransmitters, inflammation markers, viruses, and environmental pollutants. In combination with aptamer-type receptors, it is also possible to detect proteins at low concentrations. Changing the receptors to surface-imprinted polymers has opened up new possibilities for quantitative bacterial detection and identification in complex matrices. In receptor-free variants, HTM was successfully used to characterize lipid vesicles and eukaryotic cells (yeast strains, cancer cell lines), the latter showing spontaneous detachment under influence of the temperature gradient inherent to HTM. We will also address modifications to the original HTM technique such as M-HTM, inverted HTM, thermal wave transport analysis TWTA, and the hot-wire principle. The article concludes with an assessment of the possibilities and current limitations of the method, together with a technological forecast.

A Review on Bio- and Chemosensors for the Detection of Biogenic Amines in Food Safety Applications: The Status in 2022.

This article provides an overview on the broad topic of biogenic amines (BAs) that are a persistent concern in the context of food quality and safety. They emerge mainly from the decomposition of amino acids in protein-rich food due to enzymes excreted by pathogenic bacteria that infect food under inappropriate storage conditions. While there are food authority regulations on the maximum allowed amounts of, e.g., histamine in fish, sensitive individuals can still suffer from medical conditions triggered by biogenic amines, and mass outbreaks of scombroid poisoning are reported regularly. We review first the classical techniques used for selective BA detection and quantification in analytical laboratories and focus then on sensor-based solutions aiming at on-site BA detection throughout the food chain. There are receptor-free chemosensors for BA detection and a vastly growing range of bio- and biomimetic sensors that employ receptors to enable selective molecular recognition. Regarding the receptors, we address enzymes, antibodies, molecularly imprinted polymers (MIPs), and aptamers as the most recent class of BA receptors. Furthermore, we address the underlying transducer technologies, including optical, electrochemical, mass-sensitive, and thermal-based sensing principles. The review concludes with an assessment on the persistent limitations of BA sensors, a technological forecast, and thoughts on short-term solutions.

An Overview on Recent Advances in Biomimetic Sensors for the Detection of Perfluoroalkyl Substances.

Per- and polyfluoroalkyl substances (PFAS) are a class of materials that have been widely used in the industrial production of a wide range of products. After decades of bioaccumulation in the environment, research has demonstrated that these compounds are toxic and potentially carcinogenic. Therefore, it is essential to map the extent of the problem to be able to remediate it properly in the next few decades. Current state-of-the-art detection platforms, however, are lab based and therefore too expensive and time-consuming for routine screening. Traditional biosensor tests based on, e.g., lateral flow assays may struggle with the low regulatory levels of PFAS (ng/mL), the complexity of environmental matrices and the presence of coexisting chemicals. Therefore, a lot of research effort has been directed towards the development of biomimetic receptors and their implementation into handheld, low-cost sensors. Numerous research groups have developed PFAS sensors based on molecularly imprinted polymers (MIPs), metal-organic frameworks (MOFs) or aptamers. In order to transform these research efforts into tangible devices and implement them into environmental applications, it is necessary to provide an overview of these research efforts. This review aims to provide this overview and critically compare several technologies to each other to provide a recommendation for the direction of future research efforts focused on the development of the next generation of biomimetic PFAS sensors.

Synchronized, Spontaneous, and Oscillatory Detachment of Eukaryotic Cells: A New Tool for Cell Characterization and Identification.

Despite the importance of cell characterization and identification for diagnostic and therapeutic applications, developing fast and label-free methods without (bio)-chemical markers or surface-engineered receptors remains challenging. Here, we exploit the natural cellular response to mild thermal stimuli and propose a label- and receptor-free method for fast and facile cell characterization. Cell suspensions in a dedicated sensor are exposed to a temperature gradient, which stimulates synchronized and spontaneous cell-detachment with sharply defined time-patterns, a phenomenon unknown from literature. These patterns depend on metabolic activity (controlled through temperature, nutrients, and drugs) and provide a library of cell-type-specific indicators, allowing to distinguish several yeast strains as well as cancer cells. Under specific conditions, synchronized glycolytic-type oscillations are observed during detachment of mammalian and yeast-cell ensembles, providing additional cell-specific signatures. These findings suggest potential applications for cell viability analysis and for assessing the collective response of cancer cells to drugs.

The hot-wire concept: Towards a one-element thermal biosensor platform.

In this work, the 3ω hot-wire concept is explored as a prospective biosensing platform with a single sensing element that can detect analytes based on a change in the thermal interface conductance. A uniform receptor layer such as single-stranded DNA is immobilized on a thin aluminium wire, which serves not only as an immobilization platform but also as a heating element and temperature sensor together. The wire is heated periodically with an alternating current (angular frequency ω) and the third harmonic (frequency 3ω) of the voltage across the wire renders the efficiency of heat transfer from the wire to the surrounding medium. The amplitude of the 3ω voltage depends sensitively on the composition and conformation of the biofunctional interface layer. We illustrate this with a model system that includes blank aluminium wires, wires with silanes bound covalently to the native surface oxide, and with single-, respectively double-stranded DNA tethered to the silanes. The difference in heat-transfer due to these coatings is significant and measurable not only in a liquid but also in air. Based on this proof-of-concept, various applications come in sight such as mutation analysis and analyte detection with aptamers or molecularly-imprinted polymers as receptors. Wire materials other than aluminium are possible as well and the concept is suitable for miniaturization and parallelization.

Ionic strength controls long-term cell-surface interactions - A QCM-D study of S. cerevisiae adhesion, retention and detachment.

Understanding microbial adhesion and retention is crucial for controlling many processes, including biofilm formation, antimicrobial therapy as well as cell sorting and cell detection platforms. Cell detachment is inextricably linked to cell adhesion and retention and plays an important part in the mechanisms involved in these processes. Physico-chemical and biological forces play a crucial role in microbial adhesion interactions and altering the medium ionic strength offers a potential means for modulating these interactions. Real-time studies on the effect of ionic strength on microbial adhesion are often limited to short-term bacterial adhesion. Therefore, there is a need, not only for long-term bacterial adhesion studies, but also for similar studies focusing on eukaryotic microbes, such as yeast. Hereby, we monitored, in real-time, S. cerevisiae adhesion on gold and silica as examples of surfaces with different surface charge properties to disclose long-term adhesion, retention and detachment as a function of ionic strength using quartz crystal microbalance with dissipation monitoring. Our results show that short- and long-term cell adhesion levels in terms of mass-loading increase with increasing ionic strength, while cells dispersed in a medium of higher ionic strength experience longer retention and detachment times. The positive correlation between the cell zeta potential and ionic strength suggests that zeta potential plays a role on cell retention and detachment. These trends are similar for measurements on silica and gold, with shorter retention and detachment times for silica due to strong short-range repulsions originating from a high electron-donicity. Furthermore, the results are comparable with measurements in standard yeast culture medium, implying that the overall effect of ionic strength applies for cells in nutrient-rich and nutrient-deficient media.

QCM-D Study of Time-Resolved Cell Adhesion and Detachment: Effect of Surface Free Energy on Eukaryotes and Prokaryotes.

Cell-material interactions are crucial for many biomedical applications, including medical implants, tissue engineering, and biosensors. For implants, while the adhesion of eukaryotic host cells is desirable, bacterial adhesion often leads to infections. Surface free energy (SFE) is an important parameter that controls short- and long-term eukaryotic and prokaryotic cell adhesion. Understanding its effect at a fundamental level is essential for designing materials that minimize bacterial adhesion. Most cell adhesion studies for implants have focused on correlating surface wettability with mammalian cell adhesion and are restricted to short-term time scales. In this work, we used quartz crystal microbalance with dissipation monitoring (QCM-D) and electrical impedance analysis to characterize the adhesion and detachment of S. cerevisiae and E. coli, serving as model eukaryotic and prokaryotic cells within extended time scales. Measurements were performed on surfaces displaying different surface energies (Au, SiO2, and silanized SiO2). Our results demonstrate that tuning the surface free energy of materials is a useful strategy for selectively promoting eukaryotic cell adhesion and preventing bacterial adhesion. Specifically, we show that under flow and steady-state conditions and within time scales up to ∼10 h, a high SFE, especially its polar component, enhances S. cerevisiae adhesion and hinders E. coli adhesion. In the long term, however, both cells tend to detach, but less detachment occurs on surfaces with a high dispersive SFE contribution. The conclusions on S. cerevisiae are also valid for a second eukaryotic cell type, being the human embryonic kidney (HEK) cells on which we performed the same analysis for comparison. Furthermore, each cell adhesion phase is associated with unique cytoskeletal viscoelastic states, which are cell-type-specific and surface free energy-dependent and provide insights into the underlying adhesion mechanisms.

Passive permeability assay of doxorubicin through model cell membranes under cancerous and normal membrane potential conditions.

Doxorubicin is an anti-cancer drug that is important for breast cancer therapy. In this study, the effects of the membrane potential of breast cancer cells (-30 mV) and normal breast epithelial cells (-60 mV) on doxorubicin (DOX) permeability was studied. To achieve this goal, black lipid membranes (BLMs) as a model cell membrane were formed with DPhPC phospholipids in a single aperture of a Teflon sheet by the Montal and Mueller method. The presence of the BLM was characterized by capacitive measurements. The measured specific capacitance of 0.6 µF/cm2 after applying the Montal and Mueller method, confirming the presence of a BLM in the aperture. In addition, the very low current leakage of the BLM (9-24 pA) and ClyA-protein channel insertion in the BLM indicate the compactness, high quality, and thickness of 3-5 nm of the BLM. Afterwards, the permeability of doxorubicin through the BLM was studied at defined cell conditions (37 °C and pH 7.4), as well as cancerous and healthy epithelial-cell membrane potentials (-30 mV and -60 mV, respectively). The results show a slow DOX penetration within the first few hours, which increases rapidly with time. The initial slow penetration can be attributed to an electrostatic interaction between doxorubicin and DPhPC molecules in the model cell membrane. Furthermore, a MTT assay on MCF-10A and MCF-7 under different concentrations of doxorubicin confirmed that the cancerous MCF-7 cell line is more resistant to doxorubicin in comparison with the non-cancerous MCF-10A. Such studies highlight important strategies for designing and tuning the interaction efficacy of novel pharmaceuticals at molecular level.

Interactions of Aqueous Imidazolium-Based Ionic Liquid Mixtures with Solid-Supported Phospholipid Vesicles.

Despite the environmentally friendly reputation of ionic liquids (ILs), their safety has been recently questioned given their potential as cytotoxic agents. The fundamental mechanisms underlying the interactions between ILs and cells are less studied and by far not completely understood. Biomimetic films are here important biophysical model systems to elucidate fundamental aspects and mechanisms relevant for a large range of biological interaction ranging from signaling to drug reception or toxicity. Here we use dissipative quartz crystal microbalance QCM-D to examine the effect of aqueous imidazolium-based ionic liquid mixtures on solid-supported biomimetic membranes. Specifically, we assess in real time the effect of the cation chain length and the anion nature on a supported vesicle layer of the model phospholipid DMPC. Results indicate that interactions are mainly driven by the hydrophobic components of the IL, which significantly distort the layer and promote vesicle rupture. Our analyses evidence the gradual decrease of the main phase transition temperature upon increasing IL concentration, reflecting increased disorder by weakening of lipid chain interactions. The degree of rupture is significant for ILs with long hydrophobic cation chains and large hydrophobic anions whose behavior is reminiscent of that of antimicrobial peptides.

Heat-transfer-method-based cell culture quality assay through cell detection by surface imprinted polymers.

Previous work has indicated that surface imprinted polymers (SIPs) allow for highly specific cell detection through macromolecular cell imprints. The combination of SIPs with a heat-transfer-based read-out technique has led to the development of a selective, label-free, low-cost, and user-friendly cell detection assay. In this study, the breast cancer cell line ZR-75-1 is used to assess the potential of the platform for monitoring the quality of a cell culture in time. For this purpose, we show that the proposed methodology is able to discriminate between the original cell line (adherent growth, ZR-75-1a) and a descendant cell line (suspension growth, ZR-75-1s). Moreover, ZR-75-1a cells were cultured for a prolonged period of time and analyzed using the heat-transfer method (HTM) at regular time intervals. The results of these experiments demonstrate that the thermal resistance (Rth) signal decays after a certain number of cell culture passages. This can likely be attributed to a compromised quality of the cell culture due to cross-contamination with the ZR-75-1s cell line, a finding that was confirmed by classical STR DNA profiling. The cells do not express the same functional groups on their membrane, resulting in a weaker bond between cell and imprint, enabling cell removal by mechanical friction, provided by flushing the measuring chamber with buffer solution. These findings were further confirmed by HTM and illustrate that the biomimetic sensor platform can be used as an assay for monitoring the quality of cell cultures in time.

Increase in concentration of soluble HLA-G in high-quality embryos after intracytoplasmic sperm injection.

Non-invasive methods are normally preferred to conventional invasive methods when selecting suitable embryos to improve pregnancy rates after assisted reproduction techniques. One of the most recognized non-invasive methods is to examine the supernatants of embryo culture media. Soluble human leukocyte antigen, class I, G (sHLA-G) antigen is a non-classical class I molecule that has been widely considered as a marker of pregnancy failure or implantation success. In the current study of some Iranian patients, we examined the concentration of sHLA-G at different time points after intracytoplasmic sperm injection and compared the rates to the morphology and quality of the selected embryos. We showed that the concentration of sHLA-G increases over time in high-quality embryos. We conclude that there is a positive relationship between morphology, quality, and sHLA-G concentration. We suggest that this relationship can be used to increase the chance of a successful pregnancy.