To achieve better accuracy in their investigations, biologists have recently been using three-dimensional models as intermediates between two-dimensional cell culture and the in vivo study of tissues. Some of these models are based on spheroids, which are cellular aggregates retaining many characteristics of in vivo behaviors while being still easy to use and implement in labs. To study such objects, multiple observation techniques can be used according to the objective of the study, including those using electromagnetic waves as nondestructive, noninvasive, and label-free analysis. Low-frequency (<1 MHz) ones are currently under investigation as electrochemical impedance spectroscopy. However, unlike microwaves (from 300 MHz to 60 GHz), they cannot penetrate the cell. Furthermore, most of the devices dedicated to microwave dielectric characterization are only focusing on cellular scales lower than spheroids (single cell, cell mats, or suspensions) or on tissues and organs. In this article, a microwave spectroscopy device using a coplanar waveguide adapted to the study of spheroids is presented with the dielectric characterization of fixed spheroids, with capacitance and conductance measurements from 0.5 to 20 GHz, aiming at filling the scale gap in the state of the art.

Pathogenic helminth infections are responsible for severe health problems and economic losses worldwide. Timely and accurate diagnosis of helminth infections is critical for adopting suitable strategies for pathogen control. Here, we review recent advances in nucleic acid-based diagnostic methods, including polymerase chain reaction, quantitative qPCR, loop-mediated isothermal amplification and recombinase polymerase amplification, and discuss their advantages and disadvantages for diagnosing helminth infections. In addition, we highlight recent advances in biosensors for the detection of nucleic acid biomarkers that can potentially be used for the diagnosis of helminth infection.

We report results of the studies relating to the fabrication of nanostructured zirconia (nZrO2) based immunosensor for cardiac troponin I biomarker (acute myocardial infarction) detection. One step, low temperature hydrothermal process was used for the synthesis of nZrO2 (∼5 nm). This nZrO2 was functionalized with 3-aminopropyl triethoxy silane (APTES) and thereafter it was electrophoretically deposited on to indium tin oxide (ITO) coated glass electrode. EDC/NHS surface chemistry was used for covalent immobilization of monoclonal anti-troponin-I (anti-cTnI) antibodies onto APTES/nZrO2/ITO electrode. The structural, morphological and functional characterization of the synthesized nanoparticles and the fabricated immunoelectrode were conducted via X-ray diffraction (XRD), transmission electron microscopy (TEM), Fourier transform infrared spectroscopy (FTIR), and electrochemical techniques. The results of electrochemical response studies of BSA/anti-cTnI/APTES/nZrO2/ITO immunoelectrode reveal that, this platform can be used for efficient detection of cardiac troponin I (cTnI) biomarker with a wide linear detection range (0.1–100 ng/mL) and sensitivity [3.9 µA mL/(ng cm2)].

Bovine mastitis is the most costly disease for dairy farmers, hence, control measures to prevent it are crucial for dairy farm sustainability. Staphylococcus aureus is considered a major mastitis pathogen because of its impact on milk quality and low cure rates. Prevention of S. aureus mastitis includes segregation of infected animals, whilst treatment of such animals should be performed for a longer time to improve cure rates. This makes identification of S. aureus infected quarters and animals of significant importance. The experiments reported in this research paper aimed to develop and validate a sensitive method for magnetic detection of S. aureus and of the Staphylococcus genus in raw milk samples. Mastitic milk samples were collected aseptically from 47 cows with subclinical mastitis, from 12 Portuguese dairy farms. Forty nine quarter milk samples were selected based on bacteriological results. All samples were submitted to PCR analysis. In parallel, these milk samples were mixed with a solution combining specific antibodies and magnetic nanoparticles, to be analysed using a lab-on-a-chip magnetoresistive cytometer, with microfluidic sample handling. The antibodies used in this work were a rabbit polyclonal IgG anti-S. aureus ScpA protein and a mouse monoclonal IgM anti-S. aureus ATCC 29740. This paper describes the methodology used for magnetic detection of bacteria, including analysis of false positive/negative results. This immunological recognition was able to detect bacterial presence above 100 cfu/ml, independently of antibody and targeted bacteria used in this work. Comparison with PCR results showed sensitivities of 57·1 and 79·3%, specificity values of 75 and 50%, and PPV values of 40 and 95·8% for magnetic identification of Staphylococci species with an anti-S. aureus antibody and an anti-Staphylococcus spp. antibody, respectively. Some constraints are described as well as the method's limitations in bacterial quantification. Sensitivities and specificities require to be improved, nevertheless, the methodology described may form the basis for a means of identifying S. aureus infected cows at the point of care.

Genetically encoded Förster resonance energy transfer (FRET)-based biosensors have been used to report relative concentrations of ions and small molecules, as well as changes in protein conformation, posttranslational modifications, and protein–protein interactions. Changes in FRET are typically quantified through ratiometric analysis of fluorescence intensities. Here we describe methods to evaluate ratiometric imaging data acquired through confocal microscopy of a FRET-based inorganic phosphate biosensor in different cells and subcellular compartments of Arabidopsis thaliana. Linear regression was applied to donor, acceptor, and FRET-derived acceptor fluorescence intensities obtained from images of multiple plants to estimate FRET ratios and associated location-specific spectral correction factors with high precision. FRET/donor ratios provided a combination of high dynamic range and precision for this biosensor when applied to the cytosol of both root and leaf cells, but lower precision when this ratiometric method was applied to chloroplasts. We attribute this effect to quenching of donor fluorescence because high precision was achieved with FRET/acceptor ratios and thus is the preferred ratiometric method for this organelle. A ligand-insensitive biosensor was also used to distinguish nonspecific changes in FRET ratios. These studies provide a useful guide for conducting quantitative ratiometric studies in live plants that is applicable to any FRET-based biosensor.

Carbon nanotubes (CNT) have proven to be materials with great potential for the construction of biosensors. Development of fast, simple, and low cost biosensors to follow reactions in bioprocesses, or to detect food contaminants such as toxins, chemical compounds, and microorganisms, is presently an important research topic. This report includes microscopy and spectroscopy to characterize raw and chemically modified multiwall carbon nanotubes (MWCNTs) synthesized by chemical vapor deposition with the intention of using them as the active transducer in bioprocessing sensors. MWCNT were simultaneously purified and functionalized by an acid mixture involving HNO3–H2SO4 and amyloglucosidase attached onto the chemically modified MWCNT surface. A 49.0% decrease in its enzymatic activity was observed. Raw, purified, and enzyme-modified MWCNTs were analyzed by scanning and transmission electron microscopy and Raman and X-ray photoelectron spectroscopy. These studies confirmed purification and functionalization of the CNTs. Finally, cyclic voltammetry electrochemistry was used for electrical characterization of CNTs, which showed promising results that can be useful for construction of electrochemical biosensors applied to biological areas.

This work focuses on studying the effect of flow velocity on microcantilever-based biosensor by numerical simulation. The microcantilever sensors used in detecting biomolecules have attractive advantages like cost efficiency, real-time and ability of fabricating in array. Both rectangular and triangular shapes of a general model of microcantilever beam are considered. Several important physical phenomena are obtained. Comparing with the first order Langmuir theory, we have calculated the effect on the reactive rate, produced concentration, the distribution of concentration and deflection in the z axis by solving these physical coupled problem involving flow field, concentration field and chemical reaction on the reaction surface. It is found numerically that the transportation of analyte, reactive rate, the distribution of concentration and deflection in the z axis are all effected by changing the flow velocity. The result has shown that flow velocity is an important factor for this biosensor.

We demonstrate a new sensitive biosensor for detection of vital fungal spores of Aspergillus niger. The biosensor is based on silicon microfabricated cantilever arrays operated in dynamic mode. The change in resonance frequency of the sensor is a function of mass binding to the cantilever surface. For specific A. niger spore immobilization on the cantilever, each cantilever was individually coated with anti-Aspergillus niger polyclonal antibodies. We demonstrate the detection of single A. niger spores and their subsequent growth on the functionalized cantilever surface by online measurements of resonance frequency shifts. The new biosensor operating in humid air allows quantitative and qualitative detection of A. niger spores as well as detection of vital, functional spores in situ within ∼4 h. The detection limit of the sensor is 103 CFU mL−1. Mass sensitivity of the cantilever sensor is ∼53 pg Hz−1.

An immunoassay to quantify αS1-casein (αS1-CN) in milk using an optical biosensor, based on surface plasmon resonance (SPR) measurement, has been developed. The assay consists of a two-step sandwich strategy, with two anti-αS1-CN antibodies directed against each extremity of the molecule. This strategy permits only intact αS1-CN to be quantified and not its degradation products. The calibration curve was obtained using a reference milk powder with a known αS1-CN concentration. Analysis time per sample was less than ten minutes. The antibody-coated surface could be used for more than 150 determinations. Detection limit was established at 0·87 μg/ml and the intra- and inter-assay variation coefficients were 2·86 and 5·31%, respectively. The method was applied to raw milk to quantify intact αS1-CN, with no pre-treatment of the sample. An initial analysis of 48 milk samples permitted αS1-CN concentrations ranging from 8·8 to 12·06 mg/ml to be obtained.