Combinatorial effect of calcium chloride and polyethylene glycol on efficient isolation of small extracellular vesicles.

Small extracellular vesicles (sEVs) are cargo-carrying cellular nano-vesicles that have been explored for developing organic drug delivery modalities (DVM), an alternative to synthetic liposomes. However, scaled-up production of sEVs is a notable challenge in bringing sEV-based DVMs from the bench to the clinic. Ultracentrifugation is the most accepted isolation approach, but the cumbersome logistical issues and aftereffects of intense 'g' force hinder their applicability. In this study, we developed a new amenable isolation strategy for sEVs using a combinatorial treatment of calcium chloride and polyethylene glycol (PEG). An equivalent volume of cell culture medium from growing lung cancer A549 and H1299 cells was incubated overnight at 4 °C with different formulations (0.1 M CaCl2, 8% PEG, 12% PEG, 0.1 M CaCl2 + 8% PEG, and 0.1 M CaCl2 + 12% PEG) and centrifuged at 4000g to purify the precipitated sEVs as a pellet. Next, the extra CaCl2 was chelated out and the buffer was exchanged with PBS. The sEV number and protein content were assessed using the NTA (nanoparticle tracking analysis) and the BCA assay, respectively. Finally, transmission electron microscopy (TEM) was used to visualize the sEVs. The data from the present study demonstrated that the combination of 8% PEG and 0.1 M CaCl2 produced comparable numbers of sEVs with the ultracentrifugation technique. The sEV characteristics and structural integrity also remained intact, as evident from the TEM images and western blot assay. Thus, here we report an efficient technique for sEV isolation that can be easily scaled up.

Cannabidiol-loaded microparticles embedded in a porous hydrogel matrix for biomedical applications.

In this study, poly (lactic-co-glycolic acid) (PLGA) microparticles loaded with cannabidiol (CBD) were synthesized (PLGA@CBD microparticles) and embedded up to 10 wt% in a chondroitin sulfate/polyvinyl alcohol hydrogel matrix. In vitro chemical, physical, and biological assays were carried out to validate the potential use of the modified hydrogels as biomaterials. The microparticles had spherical morphology and a narrow range of size distribution. CBD encapsulation efficiency was around 52%, loading was approximately 50%. Microparticle addition to the hydrogels caused minor changes in their morphology, FTIR and thermal analyses confirmed these changes. Swelling degree and total porosity were reduced in the presence of microparticles, but similar hydrophilic and degradation in phosphate buffer solution behaviors were observed by all hydrogels. Rupture force and maximum strain at rupture were higher in the modified hydrogels, whereas modulus of elasticity was similar across all materials. Viability of primary human dental pulp cells up to 21 days was generally not influenced by the addition of PLGA@CBD microparticles. The control hydrogel showed no antimicrobial activity against Staphylococcus aureus, whereas hydrogels with 5% and 10% PLGA@CBD microparticles showed inhibition zones. In conclusion, the PLGA@CBD microparticles were fabricated and successfully embedded in a hydrogel matrix. Despite the hydrophobic nature of CBD, the physicochemical and morphological properties were generally similar for the hydrogels with and without the CBD-loaded microparticles. The data reported in this study suggested that this original biomaterial loaded with CBD oil has characteristics that could enable it to be used as a scaffold for tissue/cellular regeneration.

The use of bioluminescent enzyme bioassay for the analysis of human saliva: Advantages and disadvantages.

The purpose of the work was to find optimal conditions for bioluminescent enzymatic analysis of saliva (based on the use of NADH:FMN oxidoreductase + luciferase) and then to determine the biological effect of using bioluminescence assay of saliva to study the physiological state of the body under normal and pathological conditions. The saliva of snowboarders and students were studied in the "rest-training" model. The saliva of patients diagnosed with acute pharyngitis was examined in the "sick-healthy" model. Bioluminescence assay was performed with a lyophilized and immobilized bi-enzyme system using cuvette, plate, and portable luminometers. The concentrations of secretory immunoglobulin A (sIgA) and cortisol were determined by enzyme immunoassay, and the total protein content was measured by spectrophotometric method. The activity of the bioluminescent system enzymes increased as the amount and volume of saliva in the sample was decreased. The cuvette and plate luminometers were sensitive to changes in the luminescence intensity in saliva assay. Luminescence intensity correlated with the concentrations of sIgA and cortisol. The integrated bioluminescent index for saliva was reduced in the "rest-training" model and increased in the "sick-healthy" model. Thus, the non-invasive bioluminescent saliva analysis may be a promising tool for assessing the health of the population.

Distance-based paper analytical devices integrated with molecular imprinted polymers for Escherichia coli quantification.

The development of distance-based paper analytical devices (dPADs) integrated with molecularly imprinted polymers (MIPs) to monitor Escherichia coli (E. coli) levels in food samples is presented. The fluidic workflow on the device is controlled using a designed hydrophilic bridge valve. Dopamine serves as a monomer for the formation of the E. coli-selective MIP layer on the dPADs. The detection principle relies on the inhibition of the E. coli toward copper (II) (Cu2+)-triggered oxidation of o-phenylenediamine (OPD) on the paper substrate. Quantitative detection is simply determined through visual observation of the residual yellow color of the OPD in the detection zone, which is proportional to E. coli concentration. The sensing exhibits a linear range from 25.0 to 1200.0 CFU mL-1 (R2 = 0.9992) and a detection limit (LOD) of 25.0 CFU mL-1 for E. coli detection. Additionally, the technique is highly selective with no interference even from the molecules that have shown to react with OPD to form oxidized OPD. The developed device demonstrates accuracy and precision for E. coli quantification in food samples with recovery percentages between 98.3 and 104.7% and the highest relative standard deviation (RSD) of 4.55%. T-test validation shows no significant difference in E. coli concentration measured between our method and a commercial assay. The proposed dPAD sensor has the potential for selective and affordable E. coli determination  in food samples without requiring sample preparation. Furthermore, this strategy can be extended to monitor other molecules for which MIP can be developed and integrated into paper-microfluidic platform.

Effect-based monitoring of two rivers under urban and agricultural influence reveals a range of biological activities in sediment and water extracts.

Chemical contaminants, such as pesticides, pharmaceuticals and industrial compounds are ubiquitous in surface water and sediment in areas subject to human activity. While targeted chemical analysis is typically used for water and sediment quality monitoring, there is growing interest in applying effect-based methods with in vitro bioassays to capture the effects of all active contaminants in a sample. The current study evaluated the biological effects in surface water and sediment from two contrasting catchments in Aotearoa New Zealand, the highly urbanised Whau River catchment in Tamaki Makaurau (Auckland) and the urban and mixed agricultural Koreti (New River) Estuary catchment. Two complementary passive sampling devices, Chemcatcher for polar chemicals and polyethylene (PED) for non-polar chemicals, were applied to capture a wide range of contaminants in water, while composite sediment samples were collected at each sampling site. Bioassays indicative of induction of xenobiotic metabolism, receptor-mediated effects, genotoxicity, cytotoxicity and apical effects were applied to the water and sediment extracts. Most sediment extracts induced moderate to strong estrogenic and aryl hydrocarbon (AhR) activity, along with moderate toxicity to bacteria. The water extracts showed similar patterns to the sediment extracts, but with lower activity. Generally, the polar Chemcatcher extracts showed greater estrogenic activity, photosynthesis inhibition and algal growth inhibition than the non-polar PED extracts, though the PED extracts showed greater AhR activity. The observed effects in the water extracts were compared to available ecological effect-based trigger values (EBT) to evaluate the potential risk. For the polar extracts, most sites in both catchments exceeded the EBT for estrogenicity, with many sites exceeding the EBTs for AhR activity and photosynthesis inhibition. Of the wide range of endpoints considered, estrogenic activity, AhR activity and herbicidal activity appear to be the primary risk drivers in both the Whau and Koreti Estuary catchments.

A Rationally Designed CRISPR/Cas12a Assay Using a Multimodal Reporter for Various Readouts.

The CRISPR/Cas systems offer a programmable platform for nucleic acid detection, and CRISPR/Cas-based diagnostics (CRISPR-Dx) have demonstrated the ability to target nucleic acids with greater accuracy and flexibility. However, due to the configuration of the reporter and the underlying labeling mechanism, almost all reported CRISPR-Dx rely on a single-option readout, resulting in limitations in end-point result readouts. This is also associated with high reagent consumption and delays in diagnostic reports due to protocol differences. Herein, we report for the first time a rationally designed Cas12a-based multimodal universal reporter (CAMURE) with improved sensitivity that harnesses a dual-mode reporting system, facilitating options in end-point readouts. Through systematic configurations and optimizations, our novel universal reporter achieved a 10-fold sensitivity enhancement compared to the DETECTR reporter. Our unique and versatile reporter could be paired with various readouts, conveying the same diagnostic results. We applied our novel reporter for the detection of staphylococcal enterotoxin A due to its high implication in staphylococcal food poisoning. Integrated with loop-mediated isothermal amplification, our multimodal reporter achieved 10 CFU/mL sensitivity and excellent specificity using a real-time fluorimeter, in-tube fluorescence, and lateral flow strip readouts. We also propose, using artificially contaminated milk samples, a fast (2-5 min) Triton X-100 DNA extraction approach with a comparable yield to the commercial extraction kit. Our CAMURE could be leveraged to detect all gene-encoding SEs by simply reprogramming the guide RNA and could also be applied to the detection of other infections and disease biomarkers.

Stability of SARS-CoV-2 and persistence of viral nucleic acids on common foods and widely used packaging material surfaces.

SARS-CoV-2 is still spreading globally. Studies have reported the stability of SARS-CoV-2 in aerosols and on surfaces under different conditions. However, studies on the stability of SARS-CoV-2 and viral nucleic acids on common food and packaging material surfaces are insufficient. The study evaluated the stability of SARS-CoV-2 using TCID50 assays and the persistence of SARS-CoV-2 nucleic acids using droplet digital polymerase chain reaction on various food and packaging material surfaces. Viral nucleic acids were stable on food and material surfaces under different conditions. The viability of SARS-CoV-2 varied among different surfaces. SARS-CoV-2 was inactivated on most food and packaging material surfaces within 1 day at room temperature but was more stable at lower temperatures. Viruses survived for at least 1 week on pork and plastic at 4°C, while no viable viruses were detected on hairtail, orange, or carton after 3 days. There were viable viruses and a slight titer decrease after 8 weeks on pork and plastic, but titers decreased rapidly on hairtail and carton at -20°C. These results highlight the need for targeted preventive and disinfection measures based on different types of foods, packaging materials, and environmental conditions, particularly in the cold-chain food trade, to combat the ongoing pandemic.

Fabrication of an electrospun polycaprolactone substrate for colorimetric bioassays.

Colorimetric assays rely on detecting colour changes to measure the concentration of target molecules. Paper substrates are commonly used for the detection of biomarkers due to their availability, porous structure, and capillarity. However, the morphological and mechanical properties of paper, such as fibre diameter, pore size, and tensile strength, cannot be easily tuned to meet the specific requirements of colorimetric sensors, including liquid capacity and reagent immobilisation. As an alternative to paper materials, biodegradable polymeric membranes made of electrospun polycaprolactone (PCL) fibres can provide various tunable properties related to fibre diameter and pore size.We aimed to obtain a glucose sensor substrate for colorimetric sensing using electrospinning with PCL. A feeding solution was created by mixing PCL/chloroform and 3,3',5',5'-tetramethylbenzidine (TMB)/ethanol solutions. This solution was electrospun to fabricate a porous membrane composed of microfibres consist of PCL and TMB. The central area of the membrane was made hydrophilic through air plasma treatment, and it was subsequently functionalized with a solution containing glucose oxidase, horseradish peroxidase, and trehalose.The sensing areas were evaluated by measuring colour changes in glucose solutions of varying concentrations. The oxidation reactions of glucose and TMB in sensor substrates were recorded and analysed to establish the correlation between different glucose concentrations and colour changes. For comparison, conventional paper substrates prepared with same parameters were evaluated alongside the electrospun PCL substrates. As a result, better immobilization of reagents and higher sensitivity of glucose were achieved with PCL substrates, indicating their potential usage as a new sensing substrate for bioassays.

In Silico Prediction and Biophysical Validation of Novel 14-3-3σ Homodimer Stabilizers.

14-3-3σ plays an important role in controlling tumor metabolic reprogramming and cancer cell growth. However, its function is often compromised in many cancers due to its downregulation. Previous studies found that homodimerization of 14-3-3σ is critical for its activity. However, to date, it is not known if stabilization of 14-3-3σ homodimers can improve its activity or prevent its degradation. In our previous work, we have showed that GCP-Lys-OMe is a potential 14-3-3σ homodimer stabilizer. However, its stabilizing effect was not experimentally validated. Therefore, in this study, we have attempted to predict few potential peptides that can stabilize the dimeric form of 14-3-3σ using similar in silico techniques as described previously for GCP-Lys-OMe. Subsequent [1H]-CPMG NMR experiments confirmed the binding of the peptides (peptides 3, 5, 9, and 16) on 14-3-3σ, with peptide 3 showing the strongest binding. Competitive [1H]-CPMG assays further revealed that while peptide 3 does not compete with a 14-3-3σ binding peptide (ExoS) for the protein's amphipathic groove, it was found to improve ExoS binding on 14-3-3σ. When 14-3-3σ was subjected to dynamic light scattering experiments, the 14-3-3σ homodimer was found to undergo dissociation into monomers prior to aggregation. Intriguingly, the presence of peptide 3 increased 14-3-3σ stability against aggregation. Overall, our findings suggest that (1) docking accompanied by MD simulations can be used to identify potential homodimer stabilizing compounds of 14-3-3σ and (2) peptide 3 can slow down 14-3-3σ aggregation (presumably by preventing its dissociation into monomers), as well as improving the binding of 14-3-3σ to ExoS protein.

Relationship between Dental Caries Experience and the Levels of Streptococcus mutans and Lactobacillus in Saliva of Pregnant Women.

Pregnancy-related changes may increase the risk of dental caries. The Cariogram software program was created to understand this risk better. This study aims to correlate dental caries experience with Streptococcus mutans and lactobacillus levels in saliva using real-time PCR. This case-control analytical study conducted in Erbil City, Iraq, between 2021 and 2023 used a Cariogram to assess tooth decay risk and a real-time PCR assay to detect oral bacteria. Kurdish chewing gum with oil extract was used to stimulate saliva production. The chance of preventing tooth decay was 50.57% in pregnant women and 60.26% in non-pregnant women, with a statistically significant difference. The correlation between caries risk categories and Streptococcus mutans and Lactobacillus levels in saliva was significant but weakly positive, with strengths of 0.295 and 0.213, respectively. Furthermore, the proportion of pregnant women with Lactobacillus class 2 or Lactobacillus class 3 was significantly higher than that of non-pregnant women (7% and 10% versus 2% and 1%, respectively) with a p of 0.001. The study also found that 82% of pregnant women had a very low or zero amount of Streptococcus mutans compared to 96% of non-pregnant women (p = 0.011). The study concluded that Streptococcus mutans and Lactobacillus (SM and LBs) could be accurately detected through qPCR, and their counts have a significant positive correlation with caries risk categories. These bacteria are considered important causal agents of dental caries, especially in pregnant women.

Considerations about Cytotoxicity of Resin-Based Composite Dental Materials: A Systematic Review.

The dental material industry is rapidly developing resin-based composites (RBCs), which find widespread use in a variety of clinical settings. As such, their biocompatibility has gained increasing interest. This literature review presents a summary of research into the cytotoxicity of methacrylate-based composites published from 2017 to 2023. Subject to analysis were 14 in vitro studies on human and murine cell lines. Cytotoxicity in the included studies was measured via MTT assay, LDH assay, and WST-1 assay. The QUIN Risk of Bias Tool was performed to validate the included studies. Included studies (based entirely on the results of in vitro studies) provide evidence of dose- and time-dependent cytotoxicity of dental resin-based composites. Oxidative stress and the depletion of cellular glutathione (GSH) were suggested as reasons for cytotoxicity. Induction of apoptosis by RBCs was indicated. While composites remain the golden standard of dental restorative materials, their potential cytotoxicity cannot be ignored due to direct long-term exposure. Further in vitro investigations and clinical trials are required to understand the molecular mechanism of cytotoxicity and produce novel materials with improved safety profiles.

Curcumin Attenuates Periodontal Injury via Inhibiting Ferroptosis of Ligature-Induced Periodontitis in Mice.

Periodontitis is a chronic infectious disease characterized by the destruction of connective tissue and alveolar bone that eventually leads to tooth loss. Ferroptosis is an iron-dependent regulated cell death and is involved in ligature-induced periodontitis in vivo. Studies have demonstrated that curcumin has a potential therapeutic effect on periodontitis, but the mechanism is still unclear. The purpose of this study was to investigate the protective effects of curcumin on alleviating ferroptosis in periodontitis. Ligature-induced periodontal-diseased mice were used to detect the protective effect of curcumin. The level of superoxide dismutase (SOD), malondialdehyde (MDA) and total glutathione (GSH) in gingiva and alveolar bone were assayed. Furthermore, the mRNA expression levels of acsl4, slc7a11, gpx4 and tfr1 were measured using qPCR and the protein expression of ACSL4, SLC7A11, GPX4 and TfR1 were investigated by Western blot and immunocytochemistry (IHC). Curcumin reduced the level of MDA and increased the level of GSH. Additionally, curcumin was proven to significantly increase the expression levels of SLC7A11 and GPX4 and inhibit the expression of ACSL4 and TfR1. In conclusion, curcumin plays a protective role by inhibiting ferroptosis in ligature-induced periodontal-diseased mice.

Fabrication and In Vitro Biological Assay of Thermo-Mechanically Tuned Chitosan Reinforced Polyurethane Composites.

The progressive trend of utilizing bioactive materials constitutes diverse materials exhibiting biocompatibility. The innovative aspect of this research is the tuning of the thermo-mechanical behavior of polyurethane (PU) composites with improved biocompatibility for vibrant applications. Polycaprolactone (CAPA) Mn = 2000 g-mol-1 was used as a macrodiol, along with toluene diisocyanate (TDI) and hexamethylene diisocyanate (HMDI), to develop prepolymer chains, which were terminated with 1,4 butane diol (BD). The matrix was reinforced with various concentrations of chitosan (1-5 wt %). Two series of PU composites (PUT/PUH) based on aromatic and aliphatic diisocyanate were prepared by varying the hard segment (HS) ratio from 5 to 30 (wt %). The Fourier-transformed infrared (FTIR) spectroscopy showed the absence of an NCO peak at 1730 cm-1 in order to confirm polymer chain termination. Thermal gravimetric analysis (TGA) showed optimum weight loss up to 500 °C. Dynamic mechanical analysis (DMA) showed the complex modulus (E*) ≥ 200 MPa. The scanning electron microscope (SEM) proved the ordered structure and uniform distribution of chain extender in PU. The hemolytic activities were recorded up to 15.8 ± 1.5% for the PUH series. The optimum values for the inhibition of biofilm formation were recorded as 46.3 ± 1.8% against E. coli and S. aureus (%), which was supported by phase contrast microscopy.

Treating mechanical washing wastewater with iron-in-oil characteristics by changing the fate of iron.

The mechanical washing wastewater contained a large amount of oil, and the iron wrapped in the oil was slowly released into water. This caused the effluent quality to fluctuate, causing common polymeric aluminum chloride (PAC) to ineffectively remove the water-in-oil. The method uses Ca2+ to demulsify and ClOx- to destroy the water-in-oil structure, which releases Fe from the oil droplets. The active oxygen produced by NaClOx further converts Fe2+ into Fe3+ and then combines with NaOH to form Fe(OH)3-flocs core, which improves the flocculation efficiency of PAC. The optimal ratio was approximately 400 µL of NaClOx, 200 µL of 1 mol L-1 CaO, and 12 mL of 12.8 g L-1 PAC. The oil removal rate reached 99.88% and the residue density was 178.42 mg L-1. The maximum Fe and chemical oxygen demand (COD) removal rates were close to 49.2 and 99.89%, respectively. In field applications, wastewater should be acidified first, and acidification oxidation is more effective than direct oxidation. In short, a novel way for treating mechanically washed wastewater with iron-in-oil characteristics by changing the environmental fate of iron is provided.

Rational Design of Conducting Polymer-Derived Tubular Carbon Nanoreactors for Enhanced Enzyme-like Catalysis and Total Antioxidant Capacity Bioassay Application.

The design of void-confined tubular nanostructures has aroused significant interest for catalytic applications because of their distinct microenvironment to modulate the reaction kinetics. Herein, we propose a facile wrapping-pyrolysis strategy to confine Fe0 nanoparticles (Fe NPs) inside N-doped carbon nanotubes (Fe@NC NTs) derived from Fe2O3@polypyrrole (PPy) core-sheath nanofibers (NFs). The resultant Fe@NC NTs can act as efficient enzyme mimics and exhibit a significantly higher peroxidase (POD)-like catalytic activity than unconfined Fe NPs and bare NC NTs. Kinetic experiments demonstrate that the optimized void structure benefits the affinity with the POD substrates and achieves excellent catalytic efficiency. The mechanism study reveals that the generation of •OH from H2O2 endows Fe@NC NTs with excellent POD-like performance. Furthermore, we develop a total antioxidant capacity (TAC) sensing platform on account of this efficient POD-like system, expanding their applications in the field of food safety and human healthcare.

Temperature-Responsive Nanocarrier-Regulated Alternative Release of "Cargos" for a Multiplex Photoelectrochemical Bioassay of Antibiotic-Resistant Genes.

A smart temperature stimuli-driven multiplex photoelectrochemical (PEC) assay was constructed for antibiotic resistance genes (ARGs) detection, where the stimuli-responsive gatekeeping by regulating the alternative release of "cargo" allowed for the simultaneous detection of multiple tetracycline resistance gene, using tetA (TDNA1) and tetC (TDNA2) as the model. Dual temperature-responsive nanoassemblies were embedded in the PEC bioassay as signal DNA tages: one thermoresponsive polymer (poly(N-isopropylacrylamide), PNIPAM)-capped mesoporous silica nanoparticles (MSN) with loading the "cargo" of HgO nanoparticles as signal DNA1 tags (SDNA1-PNIPAM@MSN@HgONPs) and the other antimony tartrate (SbT)-anchored silica nanospheres as signal DNA2 tags (SDNA2-SbT@SiO2NSs). At 20 °C, below the lower critical solution temperature (LCST) of PNIPAM, the "gatekeeper" PNIPAM in SDNA1-PNIPAM@MSN@HgONPs was in an ON state, igniting Hg2+ release through the pore of SiO2. While at above LCST (40 °C), it was in an OFF state. Likewise, the thermo-dependent dissociation of SbT endowed the grafted SDNA2 tags switching from the OFF (at 20 °C) to ON state (at 40 °C), igniting SbO+ release. The released Hg2+ and SbO+ triggered the amplified photocurrents due to the structure evolution of the photoactive layer into HgS/ZnS or Sb2S3/ZnS heterostructure, thus achieving sensitive detection of multiple ARGs: tetA, tetC, tetG, tetM, tetO, tetZ, tetX, and tetW. Combined with heat map analysis, rapid screening of the ARGs profiles in 12 samples could be realized. This bioassay is simple and accessible for multiple genes analysis with the detection limit down to 0.50 nM. And it was successfully applied for measuring tetracycline ARGs in real sludge samples.

Potent Time-Dependent Ice Recrystallization Inhibition Activity of Cellulose Nanocrystals in Sucrose Solutions.

Exploring novel materials with ice recrystallization inhibition (IRI) activity in several fields often starts with a quantitative analysis of ice crystal size change by a splat assay or sandwich assay on a short time scale from 0.5 to 1 h. This study found that this time scale was insufficient to evaluate the IRI activity of cellulose nanocrystals (CNCs) in a model ice cream system-25.0% sucrose solution. No IRI activity was observed in CNCs incubated with ice crystals on a short time scale of 0.5-2.0 h. However, over longer time scales, the growth of ice crystals was entirely inhibited by 1.0% CNCs (between 2 and 24 h) and 0.5% CNCs (between 24 and 72 h) with corresponding final crystal sizes of 25 and 40 µm, respectively. Additionally, ice shaping was observed on a long exposure time, but not on a short exposure time. The findings presented here can be explained by a time-dependent surface coverage of CNCs on ice crystals. The data here indicate the importance of choosing a suitable exposure time for evaluating the IRI activity of new materials and prompt a better understanding of IRI mechanisms involving CNCs.

Fluorescent bioassay for SARS-CoV-2 detection using polypyrene-g-poly(ε-caprolactone) prepared by simultaneous photoinduced step-growth and ring-opening polymerizations.

The construction of a rapid and easy immunofluorescence bioassay for SARS-CoV-2 detection is described. We report for the first time a novel one-pot synthetic approach for simultaneous photoinduced step-growth polymerization of pyrene (Py) and ring-opening polymerization of ε-caprolactone (PCL) to produce a graft fluorescent copolymer PPy-g-PCL that was conjugated to SARS-CoV-2-specific antibodies using EDC/NHS chemistry. The synthesis steps and conjugation products were fully characterized using standard spectral analysis. Next, the PPy-g-PCL was used for the construction of a dot-blot assay which was calibrated for applications to human nasopharyngeal samples. The analytical features of the proposed sensor showed a detection range of 6.03-8.7 LOG viral copy mL-1 (Ct Scores: 8-25), the limit of detection (LOD), and quantification (LOQ) of 1.84 and 6.16 LOG viral copy mL-1, respectively. The repeatability and reproducibility of the platform had a coefficient of variation (CV) ranging between 1.2 and 5.9%. The fluorescence-based dot-blot assay was tested with human samples. Significant differences were observed between the fluorescence intensity of the negative and positive samples, with an overall correct response of 93.33%. The assay demonstrated a high correlation with RT-PCR data. This strategy opens new insights into simplified synthesis procedures of the reporter molecules and their high potential sensing and diagnosis applications.

Combination of Water Quality Parameters and Bioassays for the Assessment of Two Rivers, Southern Brazil.

In the present study, water physicochemical and microbiological parameters, as well as bioassays using Allium cepa L. seeds and the fish species Astyanax jacuhiensis were used to assess the water quality of two rivers - Ilha River and Paranhana River -, located in southern Brazil. Water samples were collected at the source and mouth of the rivers and then, laboratory experiments were performed. The results evidenced high levels of aluminum and iron in water samples collected at the four sampling sites. The micronucleus (MN) test in fish showed significant difference in the frequencies of nuclear abnormalities (NA) in the mouth of the Paranhana River in comparison to control group in one sampling period, whereas the A. cepa test evidenced significant spatial differences in cytotoxicity between the source and mouth of both rivers. Therefore, these data evidence the poor water quality of the rivers studied as well as the potential toxicity to the aquatic organisms.

Reconstitution of cell migration at a glance.

Single cells migrate in a myriad of physiological contexts, such as tissue patrolling by immune cells, and during neurogenesis and tissue remodeling, as well as in metastasis, the spread of cancer cells. To understand the basic principles of single-cell migration, a reductionist approach can be taken. This aims to control and deconstruct the complexity of different cellular microenvironments into simpler elementary constrains that can be recombined together. This approach is the cell microenvironment equivalent of in vitro reconstituted systems that combine elementary molecular players to understand cellular functions. In this Cell Science at a Glance article and accompanying poster, we present selected experimental setups that mimic different events that cells undergo during migration in vivo These include polydimethylsiloxane (PDMS) devices to deform whole cells or organelles, micro patterning, nano-fabricated structures like grooves, and compartmentalized collagen chambers with chemical gradients. We also outline the main contribution of each technique to the understanding of different aspects of single-cell migration.