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Materials (Basel, Switzerland) Jul 2016The biodegradable cellular capsule, being prepared from simple vaporization of liquid marbles, is an ideal vehicle for the potential application of drug encapsulation...
The biodegradable cellular capsule, being prepared from simple vaporization of liquid marbles, is an ideal vehicle for the potential application of drug encapsulation and release. This paper reports the fabrication of cellular capsules via facile vaporization of Pickering emulsion marbles in an ambient atmosphere. Stable Pickering emulsion (water in oil) was prepared while utilizing dichloromethane (containing poly(l-lactic acid)) and partially hydrophobic silica particles as oil phase and stabilizing agents respectively. Then, the Pickering emulsion marbles were formed by dropping emulsion into a petri dish containing silica particles with a syringe followed by rolling. The cellular capsules were finally obtained after the complete vaporization of both oil and water phases. The technique of scanning electron microscope (SEM) was employed to research the microstructure and surface morphology of the prepared capsules and the results showed the cellular structure as expected. An in vitro drug release test was implemented which showed a sustained release property of the prepared cellular capsules. In addition, the use of biodegradable poly(l-lactic acid) and the biocompatible silica particles also made the fabricated cellular capsules of great potential in the application of sustained drug release.
PubMed: 28773693
DOI: 10.3390/ma9070572 -
Membranes Jun 2022Crosslinked carboxymethyl rice starch (CLCMRS), prepared via dual modifications of native rice starch (NRS) with chloroacetic acid and sodium trimetaphosphate, was...
Crosslinked carboxymethyl rice starch (CLCMRS), prepared via dual modifications of native rice starch (NRS) with chloroacetic acid and sodium trimetaphosphate, was employed to facilitate the disintegration of hydroxypropylmethylcellulose (HPMC) orodispersible films (ODFs), with or without the addition of glycerol. Fabricated by using the solvent casting method, the composite films, with the HPMC--LCMRS ratios of 9:1, 7:1, 5:1 and 4:1, were then subjected to physicochemical and mechanical evaluations, including weight, thickness, moisture content and moisture absorption, swelling index, transparency, folding endurance, scanning electron microscopy, Fourier transform infrared spectroscopy, tensile strength, elongation at break, and Young’s modulus, as well as the determination of disintegration time by using the Petri dish method (PDM) and slide frame and bead method (SFM). The results showed that HPMC-CLCMRS composite films exhibited good film integrity, uniformity, and transparency with up to 20% CLCMRS incorporation (4:1 ratio). Non-plasticized composite films showed no significant changes in the average weight, thickness, density, folding endurance (96−122), tensile strength (2.01−2.13 MPa) and Young’s modulus (10.28−11.59 MPa) compared to HPMC film (135, 2.24 MPa, 10.67 MPa, respectively). On the other hand, the moisture content and moisture absorption were slightly higher, whereas the elongation at break (EAB; 4.31−5.09%) and the transparency (4.73−6.18) were slightly lowered from that of the HPMC film (6.03% and 7.03%, respectively). With the addition of glycerol as a plasticizer, the average weight and film thickness increased, and the density decreased. The folding endurance was improved (to >300), while the transparency remained in the acceptable range. Although the tensile strength of most composite films decreased (0.66−1.75 MPa), they all exhibited improved flexibility (EAB 7.27−11.07%) while retaining structural integrity. The disintegration times of most composite films (PDM 109−331, SFM 70−214 s) were lower than those of HPMC film (PDM 345, SFM 229 s). In conclusion, the incorporation of CLCMRS significantly improved the disintegration time of the composite films whereas it did not affect or only slightly affected the physicochemical and mechanical characteristics of the films. The 5:1 and 4:1 HPMC:CLCMRS composite films, in particular, showed promising potential application as a film base for the manufacturing of orodispersible film dosage forms.
PubMed: 35736301
DOI: 10.3390/membranes12060594 -
Scientific Reports Dec 2022In nature, bacteria prevailingly reside in the form of biofilms. These elaborately organized surface-bound assemblages of bacterial cells show numerous features of...
In nature, bacteria prevailingly reside in the form of biofilms. These elaborately organized surface-bound assemblages of bacterial cells show numerous features of multicellular organization. We recently showed that biofilm growth is a true developmental process, which resembles developmental processes in multicellular eukaryotes. To study the biofilm growth in a fashion of eukaryotic ontogeny, it is essential to define dynamics and critical transitional phases of this process. The first step in this endeavor is to record the gross morphological changes of biofilm ontogeny under standardized conditions. This visual information is instrumental in guiding the sampling strategy for the later omics analyses of biofilm ontogeny. However, none of the currently available visualizations methods is specifically tailored for recording gross morphology across the whole biofilm development. To address this void, here we present an affordable Arduino-based approach for time-lapse visualization of complete biofilm ontogeny using bright field stereomicroscopy with episcopic illumination. The major challenge in recording biofilm development on the air-solid interphase is water condensation, which compromises filming directly through the lid of a Petri dish. To overcome these trade-offs, we developed an Arduino microcontroller setup which synchronizes a robotic arm, responsible for opening and closing the Petri dish lid, with the activity of a stereomicroscope-mounted camera and lighting conditions. We placed this setup into a microbiological incubator that maintains temperature and humidity during the biofilm growth. As a proof-of-principle, we recorded biofilm development of five Bacillus subtilis strains that show different morphological and developmental dynamics.
Topics: Time-Lapse Imaging; Microscopy; Bacteria
PubMed: 36476631
DOI: 10.1038/s41598-022-24431-y -
Plant Disease May 2023Strawberry (Fragaria × ananassa Duch.) is a widely cultivated economic crop in China. In April 2022, an unusual wilt disease was observed on strawberry plants (6...
Strawberry (Fragaria × ananassa Duch.) is a widely cultivated economic crop in China. In April 2022, an unusual wilt disease was observed on strawberry plants (6 months' old) in Chenzui town, Wuqing district, Tianjin, China (117°1'E, 39°17'N). The incidence across the greenhouses (≈0.34 ha) was approximately 50 to 75%. The first wilt symptoms were observed on the outer leaves, then the whole seedlings wilted and died. The rhizome of the diseased seedlings changed color and became necrotic and rotted. Symptomatic roots were surface disinfected with 75% ethanol for 30 s, washed with sterile distilled water for three times, and then cut into 3 mm2 pieces (four pieces per seedling) and placed on petri dish with potato dextrose agar (PDA) containing 50 mg/L of streptomycin sulfate and incubated at 26℃ in the dark. After 6 days' incubation, hyphal tips of the growing colonies were transferred onto PDA. Eighty-four isolates belong to five fungal species were obtained from 20 diseased root samples based on their morphological characteristics. To confirm their pathogenicity, ten two-month-old healthy seedlings of strawberry (cv Red Face) planted in sterilized nutrient soil, were inoculated by pouring 50 mL of conidial suspension (107 conidia/mL) (Cai et al. 2021). Another ten seedlings poured with sterile distilled water were used as controls. Each treatment was repeated three times in a greenhouse at 25 to 28℃ and 75% relative humidity under a 12-h photoperiod. After 15 days,only seedlings inoculated with Plectosphaerella (an original percentage of 35.71%) exhibited similar symptoms to those of diseased seedlings originally observed in the field. Seedlings had no symptoms in the control and other fungi inoculation treatments. To fulfill the Koch's postulates, Plectosphaerella isolates were reisolated from each inoculated, symptomatic seedling with a percentage of 100%, but were not recovered from any of the control seedlings. The experiments were repeated twice with similar results. The results indicated that the genus Plectosphaerella was the pathogen causing strawberry wilt. Colonies of the genus Plectosphaerella isolates on PDA were white to cream, and then gradually became salmon pink, with few aerial hyphae and slimy surfaces. Colonies produced numerous hyphal coils with conidiophores. Conidia were 4.56 to 10.07 μm × 1.11 to 4.54 μm (avg. 7.10 × 2.56 μm, n=100), septate or aseptate, ellipsoidal, hyaline and smooth. Such morphological characteristics were identical to those of Plectosphaerella spp. (Palm et al. 1995). For species identification, the ITS region and D1/D2 domain of the 28S rRNA gene of representative isolates (CM2, CM3, CM4, CM5 and CM6) were amplified and sequenced with the primer pair ITS1/ITS4 and NL1/NL4, respectively (White et al. 1990; O'Donnell and Gray 1993). By BLASTn analysis, the obtained sequences of ITS amplicon (ON629742, ON629743, ON629744, ON629745, and ON629746) and D1/D2 domain amplicon (OQ519896, OQ519897, OQ519898, OQ519899, and OQ519900) showed 99.14% to 99.81% identity to sequences of P. cucumerina (MW320463.1 and HQ239025.1) in the NCBI database. A multilocus phylogenetic tree performed by the UPGMA analysis showed that the representative isolates were assigned to the group of P. cucumerina. To our knowledge, this is the first report of P. cucumerina causing strawberry wilt worldwide. This disease may induce serious economic losses in strawberry production, thus effective management strategies should be taken.
PubMed: 37157096
DOI: 10.1094/PDIS-03-23-0544-PDN -
Materials (Basel, Switzerland) Aug 2019The wound-healing assay is commonly and widely used for investigating collective cell migration under various physical and chemical stimuli. Substrate-coating materials...
The wound-healing assay is commonly and widely used for investigating collective cell migration under various physical and chemical stimuli. Substrate-coating materials are shown to affect the wound-healing process in a cell-type dependent manner. However, experiment-to-experiment variations make it difficult to compare results from different assays. In this paper, a modified barrier wound-healing assay was reported for studying the wound-healing process on different substrates in one single petri dish. In short, half of a dish was covered with the tape, and coating materials, poly-l-lysine and gelatin, were applied to the surface. After peeling off the tape, half of the surface was coated with the desired material. Then a customized barrier was placed inside the dish to create the wound. The results indicated that surface coating did not affect cell proliferation/viability, and the wound-healing rate increased in coated surfaces compared to uncoated ones. The present study provides a platform for further understanding the mechanisms of substrate coating-dependent wound-healing processes.
PubMed: 31470524
DOI: 10.3390/ma12172775 -
SLAS Technology Apr 2023The spot assay of the budding yeast Saccharomyces cerevisiae is an experimental method that is used to evaluate the effect of genotypes, medium conditions, and...
The spot assay of the budding yeast Saccharomyces cerevisiae is an experimental method that is used to evaluate the effect of genotypes, medium conditions, and environmental stresses on cell growth and survival. Automation of the spot assay experiments from preparing a dilution series to spotting to observing spots continuously has been implemented based on large laboratory automation devices and robots, especially for high-throughput functional screening assays. However, there has yet to be an affordable solution for the automated spot assays suited to researchers in average laboratories and with high customizability for end-users. To make reproducible spot assay experiments widely available, we have automated the plate-based yeast spot assay of budding yeast using Opentrons OT-2 (OT-2), an affordable liquid-handling robot, and a flatbed scanner. We prepared a 3D-printed mount for the Petri dish to allow for precise placement of the Petri dish inside the OT-2. To account for the uneven height of the agar plates, which were made by human hands, we devised a method to adjust the z-position of the pipette tips based on the weight of each agar plate. During the incubation of the agar plates, a flatbed scanner was used to automatically take images of the agar plates over time, allowing researchers to quantify and compare the cell density within the spots at optimal time points a posteriori. Furthermore, the accuracy of the newly developed automated spot assay was verified by performing spot assays with human experimenters and the OT-2 and quantifying the yeast-grown area of the spots. This study will contribute to the introduction of automated spot assays and the automated acquisition of growth processes in conventional laboratories that are not adapted for high-throughput laboratory automation.
Topics: Humans; Saccharomyces cerevisiae; Agar; Robotics; Automation; Genotype
PubMed: 36503082
DOI: 10.1016/j.slast.2022.12.001 -
Protein Science : a Publication of the... Apr 2016From humble beginnings of a contaminated petri dish, β-lactam antibiotics have distinguished themselves among some of the most powerful drugs in human history. The... (Review)
Review
From humble beginnings of a contaminated petri dish, β-lactam antibiotics have distinguished themselves among some of the most powerful drugs in human history. The devastating effects of antibiotic resistance have nevertheless led to an "arms race" with disquieting prospects. The emergence of multidrug resistant bacteria threatens an ever-dwindling antibiotic arsenal, calling for new discovery, rediscovery, and innovation in β-lactam research. Here the current state of β-lactam antibiotics from a structural perspective was reviewed.
Topics: Anti-Bacterial Agents; Humans; Structure-Activity Relationship; beta-Lactam Resistance; beta-Lactamase Inhibitors
PubMed: 26813250
DOI: 10.1002/pro.2889 -
Sensors (Basel, Switzerland) Oct 2023Colony-Forming Unit (CFU) counting is a complex problem without a universal solution in biomedical and food safety domains. A multitude of sophisticated heuristics and...
Colony-Forming Unit (CFU) counting is a complex problem without a universal solution in biomedical and food safety domains. A multitude of sophisticated heuristics and segmentation-driven approaches have been proposed by researchers. However, U-Net remains the most frequently cited and used deep learning method in these domains. The latter approach provides a segmentation output map and requires an additional counting procedure to calculate unique segmented regions and detect microbial colonies. However, due to pixel-based targets, it tends to generate irrelevant artifacts or errant pixels, leading to inaccurate and mixed post-processing results. In response to these challenges, this paper proposes a novel hybrid counting approach, incorporating a multi-loss U-Net reformulation and a post-processing Petri dish localization algorithm. Firstly, a unique innovation lies in the multi-loss U-Net reformulation. An additional loss term is introduced in the bottleneck U-Net layer, focusing on the delivery of an auxiliary signal that indicates where to look for distinct CFUs. Secondly, the novel localization algorithm automatically incorporates an agar plate and its bezel into the CFU counting techniques. Finally, the proposition is further enhanced by the integration of a fully automated solution, which comprises a specially designed uniform Petri dish illumination system and a counting web application. The latter application directly receives images from the camera, processes them, and sends the segmentation results to the user. This feature provides an opportunity to correct the CFU counts, offering a feedback loop that contributes to the continued development of the deep learning model. Through extensive experimentation, the authors of this paper have found that all probed multi-loss U-Net architectures incorporated into the proposed hybrid approach consistently outperformed their single-loss counterparts, as well as other comparable models such as self-normalized density maps and YOLOv6, by at least 1% to 3% in mean absolute and symmetric mean absolute percentage errors. Further significant improvements were also reported through the means of the novel localization algorithm. This reaffirms the effectiveness of the proposed hybrid solution in addressing contemporary challenges of precise in vitro CFU counting.
PubMed: 37837169
DOI: 10.3390/s23198337 -
Asian Pacific Journal of Allergy and... Dec 2023Cytokine-induced killer (CIK) cells are ex-vivo expanded T cells which present a phenotype of both T and Natural Killer cell properties.
BACKGROUND
Cytokine-induced killer (CIK) cells are ex-vivo expanded T cells which present a phenotype of both T and Natural Killer cell properties.
OBJECTIVE
To compare the proliferation and functional properties of human CIK cells cultured in three cell culture plasticwares.
METHODS
The number and viability of CIK cells were monitored. The expression of surface markers (CD3 and CD56), TH1 cytokines (IFN-γ and TNF-α), and cytolytic granules (granzyme B and perforin) were determined by flow cytometry.
RESULTS
The number of CIK cells cultured in a static bag was highest compared to those in a petri dish and gas-permeable flask. However, CIK cells cultured in all plasticwares similarity expressed surface marker, TH1 cytokines, and cytolytic granules.
CONCLUSIONS
Considering safety, efficacy, and cost, a static bag is the best plasticware for culturing CIK cells.
Topics: Humans; Cytokine-Induced Killer Cells; Cells, Cultured; Interferon-gamma; Cytokines; Cell Culture Techniques
PubMed: 33274953
DOI: 10.12932/AP-140720-0913 -
Scientific Reports Jul 2020Plant growth promoting rhizobacteria (PGPR) are a functionally diverse group of microbes having immense potential as biostimulants and stress alleviators. Their...
Plant growth promoting rhizobacteria (PGPR) are a functionally diverse group of microbes having immense potential as biostimulants and stress alleviators. Their exploitation in agro-ecosystems as an eco-friendly and cost-effective alternative to traditional chemical inputs may positively affect agricultural productivity and environmental sustainability. The present study describes selected rhizobacteria, from a range of origins, having plant growth promoting potential under controlled conditions. A total of 98 isolates (ectophytic or endophytic) from various crop and uncultivated plants were screened, out of which four endophytes (n, L, K and Y) from Phalaris arundinacea, Solanum dulcamara, Scorzoneroides autumnalis, and Glycine max, respectively, were selected in vitro for their vegetative growth stimulating effects on Arabidopsis thaliana Col-0 seedlings with regard to leaf surface area and shoot fresh weight. A 16S rRNA gene sequencing analysis of the strains indicated that these isolates belong to the genera Pseudomonas, Bacillus, Mucilaginibacter and Rhizobium. Strains were then further tested for their effects on abiotic stress alleviation under both Petri-plate and pot conditions. Results from Petri-dish assay indicated strains L, K and Y alleviated salt stress in Arabidopsis seedlings, while strains K and Y conferred increases in fresh weight and leaf area under osmotic stress. Results from subsequent in vivo trials indicated all the isolates, especially strains L, K and Y, distinctly increased A. thaliana growth under both normal and high salinity conditions, as compared to control plants. The activity of antioxidant enzymes (ascorbate peroxidase, catalase and peroxidase), proline content and total antioxidative capacity also differed in the inoculated A. thaliana plants. Furthermore, a study on spatial distribution of the four strains, using either conventional Petri-plate counts or GFP-tagged bacteria, indicated that all four strains were able to colonize the endosphere of A. thaliana root tissue. Thus, the study revealed that the four selected rhizobacteria are good candidates to be explored as plant growth stimulators, which also possess salt stress mitigating property, partially by regulating osmolytes and antioxidant enzymes. Moreover, the study is the first report of Scorzoneroides autumnalis (fall dandelion) and Solanum dulcamara (bittersweet) associated endophytes with PGP effects.
Topics: Arabidopsis; Arabidopsis Proteins; Bacteria; Endophytes; Gene Expression Regulation, Plant; Osmotic Pressure; Phylogeny; Plant Leaves; Plant Shoots; RNA, Ribosomal, 16S; Salt Stress; Soil Microbiology
PubMed: 32728116
DOI: 10.1038/s41598-020-69713-5