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Biomolecular Detection and... Dec 2018Detection of genomic alterations in diseases can be achieved with current molecular technologies. However, the molecules extracted from formalin-fixed, paraffin-embedded...
OBJECTIVES
Detection of genomic alterations in diseases can be achieved with current molecular technologies. However, the molecules extracted from formalin-fixed, paraffin-embedded (FFPE) bio-samples are often limited possibly due to DNA fragmentation and crosslinking caused by the sample fixation and processing. The study objective was to design a droplet digital PCR (ddPCR) assay to assess the quality and quantity of DNA derived from various DNA extraction conditions on FFPE samples.
METHODS
We used 10 μm-thick sections from 5 FFPE oral tumoral blocks, each consisting of 10-15 sections. The protocol variables tested included: 1) tissue staining; 2) duration and 3) temperature of post-digestion heat treatment; and 4) DNA extraction method. DNA quantity was assessed using the NanoDrop 2000 (Thermo Fisher Scientific, USA), the Qubit fluorometer (Thermo Fisher Scientific, USA), and a ddPCR-based assay. DNA quality was assessed using a ddPCR assay for the degree of fragmentation and the effectiveness of removing crosslinks with varying guanine-cytosine (GC)-content.
RESULTS
Deparaffinization with xylene helped to increase the DNA yield. Tissue staining (methyl green staining, pH 6) prior to microdissection, comparing to no staining, caused additional DNA fragmentation. Compared to column-based method, DNA extracted with phenol chloroform and ethanol precipitation increased the degree of fragmentation and lowered the yield of amplifiable DNA. The cross-linking derived from GC-contents may not be the only factor impacting on the DNA quality.
CONCLUSIONS
Samples undergoing different pre-treatment conditions prior to extraction can impact the yield of amplifiable DNA. Our ddPCR assay can be used to assess for both DNA quantity and quality.
PubMed: 30560062
DOI: 10.1016/j.bdq.2018.10.001 -
Life (Basel, Switzerland) Jan 2022Numerous cyanobacteria have the potential to reduce metallic ions to form pure metal nanoparticles in a green biosynthesis process.
BACKGROUND
Numerous cyanobacteria have the potential to reduce metallic ions to form pure metal nanoparticles in a green biosynthesis process.
AIM
To investigate the production capacity of silver nanoparticles by the cyanobacterium sp. and to examine the effect of five different phytohormones, indole acetic acid, kinetin; gibberellic acid; abscisic acid; and methyl jasmonate, on this capacity.
METHODS
The cyanobacterial strain was grown for 60 days and the harvested cyanobacterium biomass was incubated with 0.1 mM of AgNO. Percentage conversion of Ag to Ag was calculated to indicate the AgNPs' production capacity. Different concentrations of the five phytohormones were added to cultures and the AgNP production was monitored throughout different time intervals.
RESULTS
sp. biosynthesized spherical AgNPs (diameter range 70 to 140 nm, average diameter 84.37 nm). The addition of indole acetic acid and kinetin provoked the maximum conversion (87.29% and 55.16%, respectively) of Ag to Ag, exceeding or slightly below that of the control (56%). Gibberellic and abscisic acids failed to elevate the Ag to Ag conversion rate (45.23% and 47.95%, respectively) above that of the control. Methyl jasmonate increased the Ag to Ag conversion rate to 90.29%, although nearly all the cyanobacterial cultures died at the end.
CONCLUSION
Phytohormones could be used to induce or inhibit the green production of AgNPs with the cyanobacterium sp. This novel manipulation technique may have several applications in agriculture or biomedicine.
PubMed: 35207426
DOI: 10.3390/life12020139 -
Animals : An Open Access Journal From... Mar 2022In this study, we present first data concerning the morphological observations of the orbital region, eye tunics, upper and lower eyelids, superficial gland of the third...
In this study, we present first data concerning the morphological observations of the orbital region, eye tunics, upper and lower eyelids, superficial gland of the third eyelid with the third eyelid, and lacrimal gland in captive adult male Asiatic black bear. The following research methods were used in the work: the eyeball morphometry, the orbital region description, macroscopic description, morphometric and histological analysis of the eye tunics and selected the accessory organs of the eye (Fontana-Masson, hematoxylin & eosin (H&E), Methyl-green-pyronin Y (MGP Y), Movat pentachrome, and picro-Mallory trichrome) as well as histochemical examination (PAS, AB pH 1.0, AB pH 2.5, AB pH 2.5/PAS and HDI). The eyeball of the Asiatic black bear was a spherical shape, while the periorbita was funnel/conical-shaped and the eye socket was of the open type. The cornea was absent of the Bowman's membrane similar to all domestic dogs and some wild dogs. There were palisades of Vogt in the corneal limbus epithelium similar to the Canidae. Degenerative choroidal tapetum lucidum similar to ranch mink (Mustelidae) has been found. The pupil was big and round in shape. The ciliary muscle, dilatator and sphincter muscle were well developed, similar to the pinnipeds. The lens was biconvex round, similar to the Canidae. The retina was composed similarly to the diurnal terrestrial carnivores. In both eyelids were observed very well-developed tarsal glands, ciliary glands and sebaceous glands. The orbital zone in the eyelids was characterized by lymphoid follicles, diffuse lymphocytes and specialized high endothelial venules. In the anterior palpebral margin of the upper eyelid, soft and short eyelashes were observed, while in the lower eyelids they were absent. The third eyelid was T-shaped and composed of the hyaline tissue, and it contained CALT, similar to that in Canidae. The superficial gland of the third eyelid was a multilobar alveolar branched complex with seromucous nature, while the lacrimal gland was also a multilobar acinar branched complex gland, but producing a mucous-serous secretion. The results of our research indicate that the features of the anatomy of the eye and orbital region in Asiatic black bear are also typical of the Ursidae family. Moreover, a detailed analysis of the morphological eye region may be useful in comparative studies and veterinary diagnostics in this bear species.
PubMed: 35405790
DOI: 10.3390/ani12070801 -
RSC Advances May 2021-Methyl-2-pyrrolidone is a toxic dipolar aprotic solvent widely used in the synthesis of polyurethane dispersions (PUD). Since legislation strongly restricts this...
-Methyl-2-pyrrolidone is a toxic dipolar aprotic solvent widely used in the synthesis of polyurethane dispersions (PUD). Since legislation strongly restricts this substance, green alternatives are essential. Dihydrolevoglucosenone and gamma valerolactone demonstrate comparable performance to that of NMP as cosolvent in the synthesis and the film forming process of PUD.
PubMed: 35478612
DOI: 10.1039/d1ra03157k -
Scientific Reports Oct 2020Morphometry and histology are essential approaches for investigation and diagnosis of musculo-skeletal disorders. Despite the advent of revolutionary methods of image...
Morphometry and histology are essential approaches for investigation and diagnosis of musculo-skeletal disorders. Despite the advent of revolutionary methods of image analysis and high resolution three-dimensional imaging technology, basic conventional light microscopy still provides an incisive overview of the structure and tissue dynamics of the musculoskeletal system. This is crucial to both preclinical and clinical research, since several clinically relevant processes, such as bone repair, osteoarthritis, and metabolic bone diseases, display distinct, if not pathognomonic, histological features. Due to the particular characteristics of the skeletal tissues (i.e., the existence of mineralized extracellular matrices), a large number of staining methods applicable to either decalcified or undecalcified tissues are available. However, it is usually the case that several staining methods need to be sequentially applied in order to achieve the different endpoints required to fully assess skeletal tissue structure and dynamics, and to allow morphometric quantification. We describe herein a novel staining method, the RGB trichrome, amenable for application to decalcified, paraffin embedded human musculoskeletal tissues. The acronym RGB corresponds to the three primary dyes used: picrosirius Red, fast Green, and alcian Blue. Although these individual pigments are commonly used either isolated, in binary combinations, or as part of more complex polychrome staining methods, when merged in the RGB trichrome staining produce high-quality/high-contrast images, permitting not only clear identification of different tissues (i.e., the different types of cartilage, bone and fibrous connective tissue), but also discrimination between calcified and uncalcified bone and cartilage, as well as an unexpected diversity of shades of color, while displaying singular properties among polychrome staining methods, such as the unveiling of the bone osteocyte dendritic/canalicular network. Hence, we propose the RGB trichrome as simple but highly-reliable tool for the preclinical and clinical study of the musculoskeletal system.
Topics: Azo Compounds; Biopsy; Bone Neoplasms; Bone and Bones; Cartilage; Eosine Yellowish-(YS); Humans; Image Processing, Computer-Assisted; Methyl Green; Muscle, Skeletal; Staining and Labeling
PubMed: 33028938
DOI: 10.1038/s41598-020-74031-x -
ACS Sensors Jul 2018The ability to measure the levels of diagnostically relevant proteins, such as antibodies, directly at the point of care could significantly impact healthcare. Thus...
The ability to measure the levels of diagnostically relevant proteins, such as antibodies, directly at the point of care could significantly impact healthcare. Thus motivated, we explore here the E-DNA "scaffold" sensing platform, a rapid, convenient, single-step means to this end. These sensors comprise a rigid nucleic acid "scaffold" attached via a flexible linker to an electrode and modified on its distal end with a redox reporter and a protein binding "recognition element". The binding of a targeted protein reduces the efficiency with which the redox reporter approaches the electrode, resulting in an easily measured signal change when the sensor is interrogated voltammetrically. Previously we have demonstrated scaffold sensors employing a range of low molecular weight haptens and linear peptides as their recognition elements. Expanding on this here we have characterized sensors employing much larger recognition elements (up to and including full length proteins) in order to (1) define the range of recognition elements suitable for use in the platform; (2) better characterize the platform's signaling mechanism to aid its design and optimization; and (3) demonstrate the analytical performance of sensors employing full-length proteins as recognition elements. In doing so we have enlarged the range of molecular targets amenable to this rapid and convenient sensing platform.
Topics: Biosensing Techniques; DNA; Electrochemical Techniques; Equipment Design; Escherichia coli Proteins; Green Fluorescent Proteins; HIV Core Protein p24; Histidine Kinase; Limit of Detection; Methyl-Accepting Chemotaxis Proteins; Molecular Weight; Nitrilotriacetic Acid; Proteins
PubMed: 29877078
DOI: 10.1021/acssensors.8b00311 -
Materials (Basel, Switzerland) Dec 2022The goal of this study is to assess 's potential for producing carboxymethylcellulose adsorption membranes with the ability to adsorb methyl green from wastewaters by...
The goal of this study is to assess 's potential for producing carboxymethylcellulose adsorption membranes with the ability to adsorb methyl green from wastewaters by the revalorization of its cellulosic fraction. The cellulose from this feedstock was extracted by an alkaline process and TAPPI standard technique T 203 cm-99 and afterwards they were carboxymethylated. The obtained carboxymethylcelluloses were deeply characterized, being observed that the carboxymethylcellulose produced from the alkaline cellulose presented the higher solubility due to its lower crystallinity degree (53.31 vs. 59.4%) and its higher substitution degree (0.85 vs. 0.74). This carboxymethylcellulose was cross-linked with citric acid in an aqueous treatment in order to form an adsorption membrane. The citric acid provided rigidity to the membrane and although it was hydrophilic it was not soluble in water. By evaluating the potential of the produced membrane for the removal of pollutant dyes from wastewater, it was observed that the adsorption membrane prepared from the carboxymethylcellulose's produced from the was able to remove 99% of the dye, methyl green, present in the wastewater. Thus, this work demonstrates the potential of the for the production of a novel and cost-effective carboxymethylcellulose adsorption membrane with high capacity to treat wastewaters.
PubMed: 36556565
DOI: 10.3390/ma15248760 -
Molecules (Basel, Switzerland) May 2019In this work, the antioxidant properties of methanolic extract of were assessed through the free radical scavenging method, ferric reducing antioxidant power and oxygen...
In this work, the antioxidant properties of methanolic extract of were assessed through the free radical scavenging method, ferric reducing antioxidant power and oxygen radical absorbance capacity. The phenolic acids content in the extract was quantified by high-performance liquid chromatography (HPLC) and the total phenol content by the Folin-Ciocalteu method. The extract was used as an antioxidant in biodiesel from canola oil composed mostly by fatty acid methyl esters identified and quantified by gas chromatography-mass spectrophotometry (GC-MS). The performance of the extract as an antioxidant was assessed by the oxidative stability index (OSI) with a Rancimat equipment at 100, 110, 120 and 130 °C. Additionally, the change of the peroxide value (PV) and the higher heating value under conditions of oxidative stress at 100 °C and air injection were measured. The antioxidant capacity of the extract reached 50,000 TAEC (micromole of Trolox antioxidant equivalent capacity per gram). The biodiesel was constituted by more than 70% of unsaturated fatty acid methyl esters (FAME), mainly methyl oleate. The time needed to reach a PV of 100 meqO/kg was almost four times longer with an antioxidant concentration of 250 mg/L than the blank. The biodiesel showed an OSI time of 1.25 h at 110 °C, while it increased to 8.8, 15.89 and 32.27 h with the antioxidant at concentrations of 250, 500 and 1000 mg/L, respectively. The methanolic extract proved to have an antioxidant capacity and it is a green antioxidant in biodiesel to increase its oxidative stability. According to the results obtained, the methanolic extract is an alternative to the commercial synthetic antioxidants used in biodiesel nowadays.
Topics: Antioxidants; Biofuels; Chromatography, High Pressure Liquid; Gas Chromatography-Mass Spectrometry; Larrea; Methanol; Oxidation-Reduction; Oxygen; Phenols; Plant Extracts; Temperature
PubMed: 31072049
DOI: 10.3390/molecules24091786 -
Scientific Reports May 2016A new octahedral ZnO/ZnFe2O4 heterostructure has been fabricated through a facile surfactant-free solvothermal method followed by thermal treatment. It exhibits a...
A new octahedral ZnO/ZnFe2O4 heterostructure has been fabricated through a facile surfactant-free solvothermal method followed by thermal treatment. It exhibits a record-high adsorption capacity (up to 4983.0 mg·g(-1)) of malachite green (MG), which is a potentially harmful dye in prevalence and should be removed from wastewater and other aqueous solutions before discharging into the environment. The octahedral ZnO/ZnFe2O4 heterostructure also demonstrates strong selective adsorption towards MG from two kinds of mixed solutions: MG/methyl orange (MO) and MG/rhodamine B (RhB) mixtures, indicating its promise in water treatment.
PubMed: 27142194
DOI: 10.1038/srep25074 -
Applied and Environmental Microbiology Oct 1981Seventeen commonly used dyes and 16 of their metabolites or derivatives were tested in the Salmonella-mammalian microsome mutagenicity test. Mutagens active with and... (Comparative Study)
Comparative Study
Seventeen commonly used dyes and 16 of their metabolites or derivatives were tested in the Salmonella-mammalian microsome mutagenicity test. Mutagens active with and without added Aroclor-induced rat liver microsome preparations (S9) were 3-aminopyrene, lithol red, methylene blue (USP), methyl yellow, neutral red, and phenol red. Those mutagenic only with S9 activation were 4-aminopyrazolone, 2,4-dimethylaniline, N,N-dimethyl-p-phenylenediamine, methyl red, and 4-phenyl-azo-1-naphthylamine. Orange II was mutagenic only without added S9. Nonmutagenic azo dyes were allura red, amaranth, ponceau R, ponceau SX, sunset yellow, and tartrazine. Miscellaneous dyes not mutagenic were methyl green, methyl violet 2B, and nigrosin. Metabolites of the azo dyes that were not mutagenic were 1-amino-2-naphthol hydrochloride, aniline, anthranilic acid, cresidine salt, pyrazolone T,R-amino salt (1-amino-2-naphthol-3,6-disulfonic disodium salt), R-salt, Schaeffer's salt (2-naphthol-6-sulfonic acid, sodium salt), sodium naphthionate, sulfanilamide, and sulfanilic acid. 4-Amino-1-naphthalenesulfonic acid sodium salt was also not mutagenic. Fusobacterium sp. 2 could reductively cleave methyl yellow to N,N-dimethyl-p-phenylenediamine which was then activated to a mutagen.
Topics: Coloring Agents; Microsomes, Liver; Mutagenicity Tests; Mutagens; Salmonella; Salmonella typhimurium
PubMed: 7039509
DOI: 10.1128/aem.42.4.641-648.1981