A molecularly imprinted nanofilm‐based quartz crystal microbalance sensor for the real‐time detection of pirimicarb

This study aimed to prepare a novel quartz crystal microbalance (QCM) sensor for the detection of pirimicarb. Pirimicarb‐imprinted poly (ethylene glycol dimethacrylate‐N‐metacryloyl‐(l )‐tryptophan methyl ester) [p (EGDMA‐MATrp)] nanofilm (MIP) on the gold surface of a QCM chip was synthesized using the molecular imprinting technique. A nonimprinted p (EGDMA‐MATrp) nanofilm (NIP) was also synthesized using the same experimental technique. The MIP and NIP nanofilms were characterized via Fourier transform infrared spectroscopy attenuated total reflectance spectroscopy, contact angle, atomic force microscopy, and an ellipsometer. A competitive adsorption experiment on the sensor was performed to display the selectivity of the nanofilm. An analysis of the QCM sensor showed that the MIP nanofilm exhibited high sensitivity and selectivity for pirimicarb determination. A liquid chromatography‐tandem mass spectrometry method was prepared and validated to determine the accuracy and precision of the QCM sensor. The accuracy and precision of both methods were determined by a comparison of six replicates at three different concentrations to tomato samples extracted by using a Quick, Easy, Cheap, Effective, Rugged and Safe (QuEChERS) method. The limit of detection of the QCM sensor was found to be 0.028 nM. In conclusion, the QCM sensor showed good accuracy, with recovery percentages between 91 and 94%. Also, the pirimicarb‐imprinted QCM sensor exhibited a fast response time, reusability, high selectivity and sensitivity, and a low limit of detection. Therefore, it offers a serious alternative to the traditional analytical methods for pesticide detection in both natural sources and aqueous solutions.     

A thin quartz cell suitable for vacuum ultraviolet absorption and circular dichroism measurements

The design of a thin quartz cell suitable for absorption and circular dichroism measurements in the vacuum ultraviolet is described. Important features of the cell are (1) that it can be disassembled for cleaning and reproducibly reassembled with path lengths up to 0.3 mm, and (2) that strain in the windows from the compressed sample can be relieved by a sample overflow port. The latter feature allows the cell to be used for circular dichroism as well as absorption measurements.     

3D modeling and characterization of the human CD115 monoclonal antibody H27K15 epitope and design of a chimeric CD115 target

The humanized monoclonal antibody H27K15 specifically targets human CD115, a type III tyrosine kinase receptor involved in multiple cancers and inflammatory diseases. Binding of H27K15 to hCD115 expressing cells inhibits the functional effect of colony-stimulating factor-1 (CSF-1), in a non-competitive manner. Both homology modeling and docking programs were used here to model the human CD115 extracellular domains, the H27K15 variable region and their interaction. The resulting predicted H27K15 epitope includes mainly the D1 domain in the N-terminal extracellular region of CD115 and some residues of the D2 domain. Sequence alignment with the non-binding murine CD115, enzyme-linked immunosorbent assay, nuclear magnetic resonance spectroscopy and affinity measurements by quartz crystal microbalance revealed critical residues of this epitope that are essential for H27K15 binding. A combination of computational simulations and biochemical experiments led to the design of a chimeric CD115 carrying the human epitope of H27K15 in a murine CD115 backbone that is able to bind both H27K15 as well as the murine ligands CSF-1 and IL-34. These results provide new possibilities to minutely study the functional effects of H27K15 in a transgenic mouse that would express this chimeric molecule.     

Crystal structure of the C-terminal domain of Mu phage central spike and functions of bound calcium ion

Bacteriophage Mu, which has a contractile tail, is one of the most famous genus of Myoviridae. It has a wide host range and is thought to contribute to horizontal gene transfer. The Myoviridae infection process is initiated by adhesion to the host surface. The phage then penetrates the host cell membrane using its tail to inject its genetic material into the host. In this penetration process, Myoviridae phages are proposed to puncture the membrane of the host cell using a central spike located beneath its baseplate. The central spike of the Mu phage is thought to be composed of gene 45 product (gp45), which has a significant sequence homology with the central spike of P2 phage (gpV). We determined the crystal structure of shortened Mu gp45Δ1-91 (Arg92–Gln197) at 1.5 Å resolution and showed that Mu gp45 is a needlelike structure that punctures the membrane. The apex of Mu gp45 and that of P2 gpV contained iron, chloride, and calcium ions. Although the C-terminal domain of Mu gp45 was sufficient for binding to the E. coli membrane, a mutant D188A, in which the Asp amino acid residue that coordinates the calcium ion was replaced by Ala, did not exhibit a propensity to bind to the membrane. Therefore, we concluded that calcium ion played an important role in interaction with the host cell membrane.     

A phylogeographic study of the stoneplant Conophytum (Aizoaceae; Ruschioideae; Ruschieae) in the Bushmanland Inselberg Region (South Africa) suggests anemochory

The Bushmanland Inselberg Region (BIR) of South Africa provides an ideal system to study population interactions, as these inselbergs function as islands of Succulent Karoo surrounded by Nama Karoo vegetation. The population genetics of four Conophytum taxa endemic to the quartz-associated habitats of inselbergs in the BIR were investigated using amplified fragment length polymorphisms (AFLP), namely C. marginatum subsp. haramoepense, C. marginatum subsp. marginatum, C. maughanii, and C. ratum. Conophytum marginatum colonizes the quartz outcrops on the summits of the inselbergs, while C. maughanii and C. ratum occupy quartz patches at the summit and base of the inselbergs. A total of 24 populations were sampled to assess genetic differentiation between populations of each species, specifically between summit and base populations of C. ratum, eastern and western populations of C. maughanii and populations of the two subspecies of C. marginatum. Moderate levels of genetic differentiation were recovered between the summit and base populations of C. ratum, with an indication of some genetic connectivity between the populations. Slight differentiation between the eastern and western populations of C. maughanii was recovered, however, this was not reflected in the PCoA and STRUCTURE results. In C. marginatum, no significant genetic differentiation was recovered between populations of the subspecies. These results may reflect evidence of different dispersal mechanisms in the species, with the genetic connectivity between populations of C. ratum possibly indicating dispersal through hygrochastic capsules, while genetic connectivity between populations of C. maughanii and C. marginatum may, for the first time, suggest long-distance dispersal, i.e. anemochory. This study provides the first insights into population interactions across the BIR and highlights the importance of conservation in the region, particularly of the Gamsberg, in light of the recent mining activities.     

EDTA-induced Membrane Fluidization and Destabilization： Biophysical Studies on Artificial Lipid Membranes

The molecular mechanism of ethylenediaminetetraacetic acid （EDTA）-induced membrane destabilization has been studied using a combination of four biophysical techniques on artificial lipid membranes. Data from Langmuir film balance and epifluorescence microscopy revealed the fluidization and expansion effect of EDTA on phase behavior of monolayers of either 1,2-dipalmitoyl-sn-glycero-3-phosphocholine （DPPC） or mixtures of DPPC and metal-chelating lipids, such as N^a,N^a-Bis[carboxymethyl]-N^ε [（dioctadecylamino）succinyl]-L-lysine or 1,2-dioleoyl-sn-glycero-3-[N-（5-amino- 1 -carboxypentyl iminodiacetic acid） succinyl]. A plausible explanation could be drawn from the electrostatic interaction between negatively charged groups of EDTA and the positively charged choline head group of DPPC. Intercalation of EDTA into the lipid membrane induced membrane curvature as elucidated by atomic force microscopy. Growth in size and shape of the membrane protrusion was found to be time-dependent upon exposure to EDTA. Further loss of material from the lipid membrane surface was monitored in real time using a quartz crystal microbalance. This indicates membrane restabilization by exclusion of the protrusions from the surface. Loss of lipid components facilitates membrane instability, leading to membrane permeabilization and lysis.     

Water availability shapes edaphic and lithic cyanobacterial communities in the Atacama Desert

In the Atacama Desert, cyanobacteria grow on various substrates such as soils (edaphic) and quartz or granitoid stones (lithic). Both edaphic and lithic cyanobacterial communities have been described but no comparison between both communities of the same locality has yet been undertaken. In the present study, we compared both cyanobacterial communities along a precipitation gradient ranging from the arid National Park Pan de Azúcar (PA), which resembles a large fog oasis in the Atacama Desert extending to the semiarid Santa Gracia Natural Reserve (SG) further south, as well as along a precipitation gradient within PA. Various microscopic techniques, as well as culturing and partial 16S rRNA sequencing, were applied to identify 21 cyanobacterial species; the diversity was found to decline as precipitation levels decreased. Additionally, under increasing xeric stress, lithic community species composition showed higher divergence from the surrounding edaphic community, resulting in indigenous hypolithic and chasmoendolithic cyanobacterial communities. We conclude that rain and fog water, respectively, cause contrasting trends regarding cyanobacterial species richness in the edaphic and lithic microhabitats.     

Application of a quartz crystal microbalance to evaluate biodegradability of starch by Bacillus subtilis

Biodegradation of solution-cast starch films by Bacillus subtilis was monitored using a quartz crystal microbalance (QCM). A starch film was formed on the crystal by solution casting and exposed to the Bacillus subtilis culture in a bioreactor. The high sensitivity of the QCM could monitor small weight changes of the starch films on the crystal in the initial stages of biodegradation by secreted exo-enzymes of the bacterium. The feasibility of this approach as a means of quantification and characterisation of biodegradability of different polymeric materials by selected organisms is discussed.     

Measurement of fluid viscosity at microliter volumes using quartz impedance analysis

The purpose of this work was to measure viscosity of fluids at low microliter volumes by means of quartz crystal impedance analysis. To achieve this, a novel setup was designed that allowed for measurement of viscosity at volumes of 8 to 10 μL. The technique was based on the principle of electromechanical coupling of piezoelectric quartz crystals. The arrangement was simple with measurement times ranging from 2 to 3 minutes. The crystal setup assembly did not impose any unwanted initial stress on the unloaded quartz crystal. Quartz crystals of 5- and 10-MHz fundamental frequency were calibrated with glycerol-water mixtures of known density and viscosity prior to viscosity measurements. True frequency shifts, for the purpose of this work, were determined followed by viscosity measurement of aqueous solutions of sucrose, urea, PEG-400, glucose, and ethylene glycol at 25°C±0.5°C. The measured viscosities were found to be reproducible and consistent with the values reported in the literature. Minor inconsistencies in the measured resistance and frequency shifts did not affect the results significantly, and were found to be experimental in origin rather than due to electrode surface roughness. Besides, as expected for a viscoelastic fluid, PEG 8000 solutions, the calculated viscosities were found to be less than the reported values due to frequency dependence of storage and loss modulus components of complex viscosity. From the results, it can be concluded that the present setup can provide accurate assessment of viscosity of Newtonian fluids and also shows potential for analyzing non-Newtonian fluids at low microliter volumes.     

Sandwich microgravimetric immunoassay: sensitive and specific detection of low molecular weight analytes using piezoelectric quartz crystal

A multi-channel sandwich microgravimetric immunoassay (sMIA), using the quartz crystal microbalance (QCM) principle, has been developed to quantify low molecular weight substances in standard solutions. An antigen is sandwiched between two antigen-specific antibodies: the first antibody is coated on the quartz crystal surface and the second antibody is used for the detection of analyte. The concentration of low molecular weight antigen (insulin was used in this study, M _r6000 Da) was correlated with the shift of resonant frequency of QCM system before and after second antibody binding to insulin. The developed assay is highly specific showing low cross-reactivity, and is sensitive to approx. 1 ng insulin ml^–1 with a linear response for insulin from 10 g ml^–1 to 10 ng ml^–1 in standard solutions. The technique may also be applied for the detection of other small biomolecules.