Assessment of dimensional stability of novel VPES impression material at different time intervals with standard disinfectants.

BACKGROUND: Vinyl polyether silicone (VPES) is a novel impression biomaterial made of a combination of vinyl polysiloxane (VPS) and polyether (PE). Thus, it is significant to assess its properties and behaviour under varied disinfectant test conditions. This study aimed to assess the dimensional stability of novel VPES impression material after immersion in standard disinfectants for different time intervals. METHODS: Elastomeric impression material used -medium body regular set (Monophase) [Exa'lence GC America]. A total of 84 Specimens were fabricated using stainless steel die and ring (ADA specification 19). These samples were distributed into a control group (n=12) and a test group (n=72). The test group was divided into 3 groups, based on the type of disinfectant used - Group-A- 2% Glutaraldehyde, Group-B- 0. 5% Sodium hypochlorite and Group-C- 2% Chlorhexidine each test group was further divided into 2 subgroups (n=12/subgroup) based on time intervals for which each sample was immersed in the disinfectants - subgroup-1- 10 mins and Subgroup 2- 30 mins. After the impression material was set, it was removed from the ring and then it was washed in water for 15 seconds. Control group measurements were made immediately on a stereomicroscope and other samples were immersed in the three disinfection solutions for 10 mins and 30 mins to check the dimensional stability by measuring the distance between the lines generated by the stainless steel die on the samples using a stereomicroscope at x40 magnification. RESULTS: The distance measured in the control group was 4397.2078 µm and 4396.1571 µm; for the test group Group-A- 2% Glutaraldehyde was 4396.4075 µm and 4394.5992 µm; Group-B- 0. 5% Sodium hypochlorite was 4394.5453 µm and 4389.4711 µm Group-C- 2% Chlorhexidine was 4395.2953 µm and 4387.1703 µm respectively for 10 mins and 30 mins. Percentage dimensional change was in the range of 0.02 - 0.25 for all the groups for 10 mins and 30 mins. CONCLUSIONS: 2 % Glutaraldehyde is the most suitable disinfectant for VPES elastomeric impression material in terms of dimensional stability and shows minimum dimensional changes as compared to that of 2% Chlorhexidine and 0.5% Sodium hypochlorite.

Assessment of the efficiency and stability of enzymatic membrane reaction utilizing lipase covalently immobilized on a functionalized hybrid membrane.

Enzyme immobilization in membrane bioreactors has been considered as a practical approach to enhance the stability, reusability, and efficiency of enzymes. In this particular study, a new type of hybrid membrane reactor was created through the phase inversion method, utilizing hybrid of graphene oxide nanosheets (GON) and polyether sulfone (PES) in order to covalently immobilize the Candida rugosa lipase (CRL). The surface of hybrid membrane was initially modified by (3-Aminopropyl) triethoxysilane (APTES), before the use of glutaraldehyde (GLU), as a linker, through the imine bonds. The resulted enzymatic hybrid membrane reactors (EHMRs) were then thoroughly analyzed by using field-emission scanning electron microscopy (FE-SEM), contact angle goniometry, surface free energy analysis, X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectroscopy, attenuated total reflection (ATR), and energy-dispersive X-ray (EDX) spectroscopy. The study also looked into the impact of factors such as initial CRL concentration, storage conditions, and immobilization time on the EHMR's performance and activity, which were subsequently optimized. The results demonstrated that the CRLs covalently immobilized on the EHMRs displayed enhanced pH and thermal stability compared to those physically immobilized or free. These covalently immobilized CRLs could maintain over 60% of their activity even after 6 reaction cycles spanning 50 days. EHMRs are valuable biocatalysts in developing various industrial, environmental, and analytical processes.

Fast and cost-effective preparation of plant cells for scanning electron microscopy (SEM) analysis.

The analysis of plant cell structure provides valuable information about its morphological, physiological, and biochemical characteristics. Nowadays, scanning electron microscope (SEM) is widely used to provide high-resolution images at the surface of biological samples. However, biological specimens require preparation, including dehydration and coating with conductive materials for imaging by SEM. There are several techniques for providing images with maximum maintenance of cell structure and minimum cellular damage, but each requires the use of expensive and hazardous materials, which can be damaging to the cell in many cases. Therefore, the provision of new and effective preparation methods based on maintaining cell structure for imaging can be very practical. In the present study, a fast and cost-effective protocol was first performed for chemical fixation and preparation of the plant cells for imaging by SEM. Taxus baccata and Zhumeria majdae cells were chemically fixed using glutaraldehyde and then successfully dried with different percentages of ethanol including 70, 80, 90, and 100%. In addition, SEM was performed for imaging the cell surface in different micro-scales. This protocol can be used by plant cell biologists and biotechnologists who are interested in studying structural and biochemical responses of treated or stressed plant cells by SEM.

Quantitative Assessment of Cerebral Basement Membranes Using Electron Microscopy.

In this chapter we describe in detail the tissue processing techniques we employ for the study of cerebral tissue by transmission electron microscopy (TEM). In particular, we explain a technique that enables quantification of changes in cerebral basement membranes at the ultrastructural level. This is significant, as age related pathological conditions affecting the brain are often accompanied by ultrastructural changes in the cerebral vasculature.Briefly, experimental mice are fixed by perfusion and their brains removed. Brains are then vibratomed into 100 µm slices with regions of interest microdissected and processed for TEM following a protocol optimized for the preservation of cerebral tissue. Changes in the thickness of cerebral basement membranes are then quantified using novel software. Some prior knowledge of general TEM specimen preparation and sectioning will be useful when performing this protocol.

Aggregation-induced emissive nanoparticles for fluorescence signaling in a low cost paper-based immunoassay.

Low cost paper based immunoassays are receiving interest due to their fast performance and small amounts of biomolecules needed for developing an immunoassay complex. In this work aggregation-induced emissive (AIE) nanoparticles, obtained from a diastereoisomeric mixture of 1,2-di-(4-hydroxyphenyl)-1,2-diphenylethene (TPEDH) in a one-step top-down method, are characterized through Dynamic Light Scattering (DLS), Scanning Electron Microscopy (SEM), and Zeta potential. By measuring the Zeta potential before and after labeling the nanoparticles with antibodies we demonstrate that the colloidal system is stable in a wide pH-range. The AIE-active nanoparticles are deposited on chitosan and glutaraldehyde modified paper pads overcoming the common aggregation-caused quenching (ACQ) effect. Analyte concentrations from 1000ng and below are applied in a model immunocomplex using Goat anti-Rabbit IgG and Rabbit IgG. In the range of 7.81ng-250ng, linear trends with a high R(2) are observed, which leads to a strong increase of the blue fluorescence from the TPEDH nanoparticles.

Gelatin porous scaffolds fabricated using a modified gas foaming technique: characterisation and cytotoxicity assessment.

The current study presents an effective and simple strategy to obtain stable porous scaffolds from gelatin via a gas foaming method. The technique exploits the intrinsic foaming ability of gelatin in the presence of CO2 to obtain a porous structure stabilised with glutaraldehyde. The produced scaffolds were characterised using physical and mechanical characterisation methods. The results showed that gas foaming may allow the tailoring of the 3-dimensional structure of the scaffolds with an interconnected porous structure. To assess the effectiveness of the preparation method in mitigating the potential cytotoxicity risk of using glutaraldehyde as a crosslinker, direct and in-direct cytotoxicity assays were performed at different concentrations of glutaraldehyde. The results indicate the potential of the gas foaming method, in the preparation of viable tissue engineering scaffolds.

The use of glutaraldehyde-fixed chicken red blood cells as counting beads for performing affordable single-platform CD4(+) T-lymphocyte count in HIV-1-infected patients.

CD4(+) T-lymphocyte count is an important marker in management of HIV-1-infected patients. The standard single-platform (SP) flow cytometric (FCM) CD4(+) testing that uses the known reference microbeads is expensive; more affordable alternatives are therefore needed. We evaluated the use of glutaraldehyde-fixed chicken red blood cells (CRBCs) as counting beads as an alternative for enumerating CD4(+) T-lymphocyte counts in 87 HIV-1-infected patients. Linear regression analyses revealed an excellent correlation of the SP FCM using CRBCs with the standard SP bead-based FCM method (percentages, r(2) > 0.99; absolute counts, r(2) > 0.98) over the entire range including the clinically relevant range. Mean percent bias for the CRBC method was +0.35% [limits of agreement (LOA): -1.86% to +2.57%]. For absolute CD4(+) T-lymphocytes, the mean biases was -47.76 cells per microliter (LOA: -191.34 to +98.81 cells/microL) with much lower bias for CD4 T-lymphocyte counts <200 cells per microliter (LOA: -31.92 to +22.95 cells/microL). The use of CRBCs is comparable with the use of commercial microbeads. This has resulted in major cost savings to resource-limited countries where the health care system is under increasing pressure to operate cost effectively. This can greatly facilitate and ensure the success of the ongoing antiretroviral therapy program in these countries.

Transcardial perfusion versus immersion fixation for assessment of peripheral nerve regeneration.

Accurate assessment of peripheral nerve regeneration requires fixation techniques that preserve tissue in a natural state with minimal artifact. While transcardial perfusion fixation is accepted as the gold standard for tissue fixation, the less cumbersome approach of immersion fixation has been criticized for introducing artifacts in brain tissue. We investigated whether immersion fixation increased artifact compared to perfusion fixation in the rat sciatic nerve. Eighteen Lewis rats were randomized into three groups: glutaraldehyde immersion fixation; glutaraldehyde transcardial perfusion; and paraformaldehyde transcardial perfusion. All animals underwent sciatic nerve transection and repair followed by tissue harvest and fixation at three weeks. Qualitative assessment of neural architecture and histological features was followed by quantitative analysis of nerve regeneration parameters. Outcome measures included quantitative histomorphometry, analysis of axon/myelin ratios, assessment of fiber distributions, and ultrastructural analysis. No qualitative or quantitative differences were observed with immersion fixation when compared to the transcardial perfusion fixation methods. Immersion fixation is a valid method for assessment of peripheral nerve regeneration in a rat model.

Purification and viral inactivation of hemoglobin from human placenta blood.

A new procedure was developed to obtain high-quality polymerized human hemoglobin by modifying purified hemoglobin with PLP and polymerized with GDA. Comparing polymerized hemoglobin products obtained from different methods, the product from the new procedure has similar physical, chemical, and biological properties in the molecular distribution, methemoglobin concentration, oxygen carrier capacity, P(50) and spectral analysis. Furthermore, the new procedure of modification after polymerization can save PLP greatly, and significantly reduce the cost. So the procedure of modification after polymerization is a better way in research and production.

A high throughput chemiluminescence method for determination of chemical oxygen demand in waters.

A novel and high throughput chemiluminescence (CL) method for determination of chemical oxygen demand (COD) in water sample was originally developed based on potassium permanganate-glutaraldehyde CL system. With this method, dissolved organic matter in water samples was digested by excess acid potassium permanganate, the reacted mixture solutions containing surplus KMnO(4) were added in wells of a 96-well plate, followed by injection of glutaraldehyde in the wells, and CL was then produced along with the reaction of the added glutaraldehyde with the surplus KMnO(4) and detected by a photomultiplier tube (PMT). The difference (DeltaI) between the CL intensity for distilled water and that for sample water was proportional to the COD value of water sample. The calibration graph was linear in the range of 0.16-19.24 mg L(-1) with a detection limit of 0.1 mg L(-1). A complete analysis could be performed in 40 min including digestion and detection, giving a very high throughput of 3 x 96 samples in about 60 min. Compared with the conventional methods, this method is simple and sensitive and consumes very limited and cheap reagents. Owing to its rapid, automatic, high throughput and low cost characteristics, the presented CL method has been applied successfully to the determination of COD in real water samples (n=32) with satisfactory results.

Gamma irradiated micro system for long-term parenteral contraception: An alternative to synthetic polymers.

An injectable system of levonorgestrel (LNG) was developed using biodegradable polymer of natural origin. The parenteral system was optimized for particle size and higher drug loading. The microparticulate system was characterised by scanning electron microscopy, encapsulation efficiency, moisture content, IR, DSC, XRD, residual solvent content, sterility testing, test of abnormal toxicity and test for pyrogens. The microparticles were sterilised by gamma irradiation (2.5Mrad). The system was injected intramuscularly in rabbits and the blood levels of LNG were determined using radioimmunoassay technique. An optimized drug to polymer ratio of 0.3-1.0 (w/w ratio) gave improved drug loading of about 52%. In vivo studies in rabbits showed that the drug was released in a sustained manner for a period of 1 month. The AUC(0-t) was found to be 9363.6+/-2340pg/mLday(-1) with MRT calculated to be about 16 days and Kel of 0.01day(-1). LNG levels were maintained between 200 and 400pg/mL. In vivo release exhibited an initial burst effect which was not observed in the in vitro dissolution. This promising "Progestin-only" long-term contraceptive with improved user compliance is an alternative to the synthetic expensive polymeric carriers.

Decay model for biocide treatment of unballasted vessels: application for the Laurentian Great Lakes.

A biocide decay model was developed to assess the potential efficacy and environmental impacts associated with using glutaraldehyde to treat unballasted overseas vessels trading on the Laurentian Great Lakes. The results of Monte Carlo simulations indicate that effective glutaraldehyde concentrations can be maintained for the duration of a vessel's oceanic transit (approximately 9-12 days): During this transit, glutaraldehyde concentrations were predicted to decrease by approximately 10% from initial treatment levels (e.g., 500 mgL(-1)). In terms of environmental impacts, mean glutaraldehyde concentrations released at Duluth-Superior Harbor, MN were predicted to be 100-fold lower than initial treatment concentrations, and ranged from 3.2 mgL(-1) (2 SD: 2.74) in April to 0.7 mgL(-1) (2 SD: 1.28) in August. Sensitivity analyses indicated that the re-ballasting dilution factor was the major variable governing final glutaraldehyde concentrations; however, lake surface temperatures became increasingly important during the warmer summer months.

Quality assessment of marginal sealing using 7 dentin adhesive systems.

OBJECTIVE: The goal of this investigation was to assess in vitro the quality of marginal sealing of composite-dentin adhesive systems and human dentin. METHOD AND MATERIALS: Forty intact human premolars and third molars were extracted from subjects of both sexes and of different ages. After the enamel layer was removed, a Class V cavity was formed on the buccal surface, and a wedge cavity was formed on the lingual surface. These were restored with resin composite materials and their corresponding dentin adhesive systems. The quality of marginal sealing was evaluated by assessing the linear penetration of silver nitrate dye. RESULTS: The best marginal sealing between composite materials and the cavity walls, in both wedge erosions and Class V cavities, was provided by Scotchbond Multi-Purpose/Valux and Syntac/Helioprogres systems. Dye penetration was slightly greater with the XR-Bond/Herculite, Gluma/Pekafill, and Superlux Universal Bond 2/Superlux Solar systems. The greatest microleakage was observed in Tripton/Opalux and Denthesive/Charisma specimens. CONCLUSION: The use of an adhesive system and the corresponding resin composite does not eliminate microleakage completely when the cavity margins are in dentin.

Assessment of glutaraldehyde crosslinking efficiency with an amine-specific fluorescent probe.

BACKGROUND: Crosslinking of heart valves with glutaraldehyde involves the binding of amine groups. We have developed a technique that provides an inverse measure of the degree of tissue fixation by quantifying the amount of unbound amines. METHODS: Whole aortic valves were exposed to 0.5% glutaraldehyde solution for 0, 1, 15, and 60 minutes, 6 hours, and 1 and 7 days. Frozen sections were exposed to carboxyfluorescein succinimidyl ester, a fluorescent amine-reactive probe. Images were acquired from each section and processed to separate pixels representing tissue from those representing background. An average fluorescent intensity for each image was calculated and related to the number of unbound amines by comparing with standards. RESULTS: The amount of uncrosslinked amines was observed to decrease exponentially with fixation time and achieved a plateau at 1 day of fixation. A significant difference in the amount of unbound amines also exists between valve leaflets fixed while connected to the root and those excised from the root before fixation. CONCLUSIONS: This amine measurement technique, being sensitive to spatially varying differences in chemical fixation, should be useful in evaluating the efficacy of new fixation protocols.

Valved conduit in the descending thoracic aorta in juvenile sheep: a useful, cost-effective model for accelerated calcification study in systemic circulation.

To evaluate the efficacy of any new anticalcificant in bioprostheses, a cost-effective and easy circulatory model is proposed. Calcification of 0.625% glutaraldehyde-treated porcine aortic valved conduits implanted in the descending thoracic aorta in 11 juvenile sheep for 5 months was compared with that of leaflets of glutaraldehyde-treated porcine aortic valve implanted subcutaneously in 3-week-old male Wistar rats for the same period. Cusps of valved conduits (Ca, 205.41 +/- 16.24 mg g(-1)) in sheep and aortic valve leaflets in rats (Ca, 235.21 +/- 45.25 mg g(-1)) (P = 0.0299) were severely calcified. Morphological characteristics of calcification of all explants were virtually identical. This model provides a model for testing calcification that lies between the subcutaneous weanling rat model and orthotopic whole valve replacement on the left side of the heart. It is less costly and easier to perform than the latter, but does provide exposure to the bloodstream under pressure, which the rat model does not.

The effect of glutaraldehyde adulteration of urine specimens on syva EMIT II drugs-of-abuse assays.

The effect of glutaraldehyde (the active component of "UrinAid") on Syva EMIT II drugs-of-abuse screening assays was studied. It was found that, dependent on the assay involved, concentrations of between 0.75 and 2.00% (v/v) of glutaraldehyde in urine could give rise to false-negative screening results. A simple method for identifying urine specimens that have been adulterated with glutaraldehyde, based on final absorbance rate readings (dA/min), is proposed.

An albumin-coated polyester arterial graft: in vivo assessment of biocompatibility and healing characteristics.

The albumin-coated vascular graft (ACG) and its uncoated polyester substrate, the Vascular II (V-II), were evaluated in terms of biocompatibility and biofunctionality using two in vivo animal studies. Biocompatibility and immunoreactivity were assessed by implanting intraperitoneally in the rat small segments of the ACG and the V-II graft and harvesting them with their surrounding tissue 3d, 1, 2 and 4 weeks later. Cytofluorometric determination of total T cells (CD3), the ratio of CD4/CD8 subsets and the percentage of IL-2 receptor-positive T cells in the peripheral blood has revealed that no significant difference in any of the T cell populations was found between the ACG and the V-II graft. The cellular reactivity of the ACG in terms of acid phosphatase activity at the implant side was significantly greater at 3 d but not at longer periods. Biofunctionality was evaluated by implanting both grafts as a thoracoabdominal vascular bypass in dogs for 11 different periods ranging from 4 h to 6 months. The rate of albumin resorption was such that traces were still present at 1 month, but no longer observable at 2 months. Tissue incorporation into the graft wall was earlier for the V-II (2 weeks) than for the ACG (4 weeks), which showed complete encapsulation, tissue incorporation and endothelialization after 2 months in vivo. Only small differences were observed between both grafts in terms of platelet and fibrin uptake on the luminal surface. The prostacyclin/thromboxane A2 ratio increased to a level higher that 1.0 aorta within 1 month for the V-II and 4 months for the ACG. In conclusion, the Bard ACG has demonstrated excellent biocompatibility in terms of blood T cell behaviour and acid phosphatase activity at the implant site. Finally, its healing response is equivalent to that of the uncoated Dacron prosthesis once the albumin coating has been resorbed.

A technique for three-dimensional microleakage assessment using tooth sections.

Microleakage is commonly assessed using restored teeth, sectioned through the midline of the restoration. It is impossible to determine if this is representative of the leakage throughout the whole tooth. This pilot study examined the feasibility of calculating areas and volumes of leakage using serial sections of restored teeth which had been subjected to dye immersion and image analysis. Using the sections, perspex models of the teeth were constructed to present the pattern of the dye leakage into dentine. This appears to be a viable technique for the three-dimensional assessment of microleakage.