Cross-polarized Light Microscopy Research Articles

Fabrication and stability of dual Pickering double emulsions stabilized with food-grade particles

The aim of this study was to create stable, dual Pickering-stabilized double emulsions (DEs) where both the internal and external droplets are stabilized by food grade particles. Water-in-oil (W/O) emulsions were stabilized by using cocoa butter-high oleic sunflower oil based oleogels (CBolg) and then W/O emulsion was used as the dispersed phase in water-in-oil-in-water (W/O/W) emulsions, latter stabilized by whey protein microgels (WPM). The capability of the CBolg (10 v/v% cocoa butter) to form W/O and subsequently a W/O/W emulsion was characterized using cross-polarized light microscopy, cryogenic scanning electron microscopy (cryo-SEM), confocal light microscopy (CLSM) and static light scattering. In addition, the characterization of the WPM's internal structure was evaluated by small angle X-ray scattering (SAXS). Results revealed dual Pickering stabilized W/O/W emulsions with a droplet size of 10 μm could be formed using whey protein microgel (WPM, 3–5 v/v%) as the Pickering stabilizer. Dynamic light scattering (DLS) analysis revealed WPM particles size averaging approximately 100 nm, with internal structures consisting of partially stretched protein strands with smaller compact subunits along the strand (partially folded proteins) was confirmed by SAXS analysis. The dual Pickering-stabilized DE droplets were stable over a month by formation a protective layer of WPM around the oil droplets, as observed in CLSM images, unlike the systems where the external phase was stabilized by non-microgelled whey protein, latter destabilized within hours. Such dual Pickering-stabilized DEs may offer new templates for designing healthy low-fat foods and for co-delivery of nutrients and active in multiple compartments.

Unsupervised topological analysis of polarized light microscopy: application to quantitative birefringence imaging.

The determination of birefringence (magnitude and axis orientation) of optical materials is of significant interest in various fields. In the case of composite samples, this task becomes complicated and time-consuming; therefore, a partially automated procedure for reconstructing birefringence spatial distribution becomes valuable. Herein, we propose a procedure to reconstruct the spatial distributions of the retardance and optical axis orientation in a geological thin section from sparse quantitative birefringence measurements, using automatic boundary detection on cross-polarized light microscopy images. We examine two particular areas on the selected geological thin section: one that presents a uniaxial crystal with a circular cross-section of its refractive index ellipsoid and the other with grains of varying orientations. The measurement gives the orientation of the grain's optical axis both in and out of the plane of the thin section, which explains the qualitative observations with the cross-polarized light microscope. Future work will connect the measured orientation of the rock thin section with its 3D geological orientation in the field.

Amyloid Fibrils Produced by Streptococcus sanguinis Contribute to Biofilm Formation and Immune Evasion.

Bacterial surface proteins assembled into amyloids contribute to biofilm formation and host immune evasion. Streptococcus sanguinis, a pioneer colonizer of teeth commonly involved in cardiovascular infections, expresses about thirty-three proteins anchored to the cell wall by sortase A. Here, we characterized the production of amyloid in S. sanguinis strains differing in biofilm and immune evasion phenotypes and investigated the role of sortase A in amyloidogenesis. Amyloid was identified in biofilms formed by nine strains, using Congo red (CR) staining and cross-polarized light microscopy. Additionally, EGCG, an amyloid inhibitor, impaired biofilm maturation in a strain-specific fashion. The amounts of amyloid-like components quantified in culture fluids of nine strains using thioflavin T and fluorimetry negatively correlated with bacterial binding to complement-activating proteins (SAP, C1q), C3b deposition and rates of opsonophagocytosis in PMNs, implying amyloid production in immune evasion. The deletion of the sortase A gene (srtA) in strain SK36 compromised amyloid production and sucrose-independent biofilm maturation. The srtA mutant further showed increased susceptibility to C3b deposition and altered interactions with PMNs as well as reduced persistence in human blood. These findings highlight the contribution of amyloids to biofilm formation and host immune evasion in S. sanguinis strains, further indicating the participation of sortase A substrates in amyloidogenesis.

Material Evidence of Sediments Recovered from Ancient Amphorae Found at the Potaissa Roman Fortress

Methods for material investigation are powerful tools that allow specialists to elucidate important aspects regarding ancient artifacts such as the Roman amphorae deposits discovered at Potaissa Fortress in Turda, Romania. Archeological debate states that the deposit contained olive oil and wine amphorae, but no material evidence has been presented until now. The current research is focused on the most representative large amphora fragments found in the Potaissa deposit, with a significant amount of sediment on their walls, to give archeologists the material proof to elucidate their debate. Sediment was collected from each fragment and subjected to complex analysis. XRD investigation combined with cross-polarized light microscopy demonstrated mineral particles such as quartz, clay (muscovite and traces of biotite), and calcite. Quartz and calcite particles have a rounded shape and diameters in a range of 20-200 µm, and clay particles have a lamellar shape and dimensions from 1 to 20 µm, a fact confirmed by SEM microscopy. Sample 2 presented a large amount of amorphous phase followed by Samples 1 and 3, with a low amount of organic phase. FTIR investigation confirms organic phase presence owing to strong absorption bands regarding C-H, C=O, and O-H chemical bonds related to aliphatic compounds in Sample 2, and to some decayed wine residue in Samples 1 and 3. EDS elemental analysis was used for organic particle identification in the amphora sediments and to obtain a correlation with their microstructure. GC-MS investigation showed volatile compounds related to wine residue for Samples 1 and 3 and decomposed fats for Sample 2. Tartaric and malic acid were identified by HPLC in Samples 1 and 3, which are wine biomarkers. The correlation of all experimental results concludes with no doubt that Amphora 2 contained olive oil and Amphorae 1 and 3 contained wine in ancient times.

Fluorescent labeling with 5-DTAF reduces collagen fiber uncrimping in loaded tendons

A fluorescent dye commonly used to image tissues under load (5-DTAF) has previously been shown to stiffen tendons. This study hypothesized that 5-DTAF staining stiffens tendons through reduced fiber sliding, altering the rate at which crimped collagen fibers straighten under load. This was tested by using reflected cross-polarized light microscopy to measure fiber crimp period of cervine extensor digitorum longus tendon specimens under axial load. Specimens were treated with either phosphate buffered saline (negative control), genipin (positive control), or 5-DTAF. In saline treated specimens, crimp period (relative to unstretched) increased at approximately 2.5 times the applied axial strain, indicating substantial fiber sliding. In both 5-DTAF and genipin treated specimens, this ratio was reduced to 1:1, indicating no fiber sliding. These results add further evidence that care should be taken when using 5-DTAF to stain tissue for studying microscale deformations in tissues.

Dynamic dentin: A quantitative microscopic assessment of age and spatial changes to matrix architecture, peritubular dentin, and collagens types I and III

To investigate age and site-related changes to human dentin collagen, sound human teeth collected from donors aged 13–29 (young) and 50–74 (aged) years (n = 9/group) were cut to shallow and deep sites. Dentin collagen orientation and fibril bundling was investigated using the Picrosirius Red (PSR) stain observed under cross-polarized light microscopy (Pol), and collagen distribution was investigated using Confocal Laser Scanning Microscopy (CLSM). Collagen types III to I distribution in peritubular dentin (PTD) was revealed using Herovici stain and brightfield microscopy. Image analysis software and linear mixed modelling quantified outcomes. In situ dentin collagen was observed using Xenon Plasma Focussed Ion Beam Scanning Electron Microscopy (Xe PFIB-SEM). The PSR-Pol analysis revealed less coherently aligned and more bundled collagen fibrils in aged dentin (P = 0.005). Deep inner dentin collagen in both groups were less coherently aligned with reduced bundling. Regardless of age, CLSM showed collagen distribution remained stable; and more collagen type III was detectable in PTD located in inner dentin (Young: P = 0.006; Aged: P = 0.008). Observations following Xe PFIB-SEM cross-sectioning showed apatite-like deposits surrounding large intratubular collagen fibers, and evidence of smaller intertubular dentin collagen fibrils in situ. In conclusion, aging changes collagen network architecture, but not distribution or content.

Reflected cross-polarized light microscopy as a method for measuring collagen fiber crimp in musculoskeletal tissues

Many musculoskeletal tissues are composed primarily of type I collagen, which takes on a periodic crimp morphology that allows large tensile strains in the tissue. The spatial period of collagen fiber crimp may be used to infer internal strains in a tissue and is typically measured using transmitted cross-polarized light imaging of thin slices. However, slicing may induce specimen distortion and precludes mechanical loading of the specimen during imaging. We hypothesized that reflected cross-polarized light imaging of thick tissue explants would yield crimp period measurements comparable to those obtained from transmitted light imaging of thin slices. We further hypothesized that these measurements would be sensitive to applied uniaxial strain in the fiber direction. These hypotheses were tested by imaging both intervertebral disc outer annulus fibrosus and medial collateral ligament tissue specimens. We found that both transmitted and reflected light yielded similar crimp period measurements for intervertebral disc tissue, with an overall average of 43.5 ± 11.5 μm. Reflected light yielded a significantly higher crimp period with lower variance than transmission through thin specimens (54.1 ± 10.6 μm versus 50.4 ± 16.0 μm) in the ligament. Upon application of axial tension, crimp periods in both fibers increased at a rate of approximately three times the applied strain (with 3.17% applied strain yielding a 9.64 ± 4.4% increase in crimp period in the disc and an 11.7 ± 3.7% increase in the ligament), indicating significant fibril sliding. In support of our hypotheses, these findings suggest that reflected cross-polarized light is a suitable method for measuring collagen fiber crimp in musculoskeletal tissues, both statically and under tension.

Rheology Control Using Nonionic Cosurfactants and pH Titration in an Amino Acid-Derived Surfactant Composition.

Sulfate-based formulations can be easily thickened by adding salt or amphoteric cosurfactants. However, sulfate-free and amino acid-based surfactants cannot. We explored an alternative thickening mechanism by studying the thickening effect of adding nonionic cosurfactants to a mixture of an amino acid-based surfactant, sodium lauroyl sarcosinate (SLSar), and a zwitterionic cosurfactant, cocamidopropyl hydroxysultaine (CAHS) at a 6:9 weight ratio. To characterize the formulations, we combined traditional rheometry with a state-of-the-art mesoscopic analysis of micelle dynamics obtained via diffusing wave spectroscopy. In addition, the formulations were characterized by cross-polarized light microscopy and dynamic light scattering. The cosurfactants studied included fatty alcohols, alkanediols, a fatty acid, and fatty alcohol ethoxylates (CnE3 and CnE6). Adding the nonionic cosurfactants increased the zero-shear viscosity up to 350 times the viscosity of the no-additive system at neutral pH. When pH titration was incorporated as a second thickening mechanism, the viscosity maximum was lower than the no-additive mixture. Furthermore, the pH of the viscosity maximum was shifted to higher pH for all systems except for CnE6, which shifted the maximum to lower pH. The nonionic amphiphiles also broadened the viscosity maximum, particularly in the C10OH system. Consequently, the C10OH system had a more favorable profile for development as a practical thickening system for an amino acid-based cleanser. Analysis according to the Zou and Larson micelle dynamics model revealed that the broadening effect was associated with substantially longer breakage times for the C10OH system (4-208 ms) compared to the no-additive system (4-38 ms).

Detecting a spreading non-indigenous species using multiple methodologies

Johansson ML, Lavigne SY, Ramcharan CW, Heath DD, MacIsaac HJ. Detecting a spreading non-indigenous species using multiple methodologies. Lake Reserv Manage. 36:432–443. Non-indigenous species (NIS) are often introduced to novel environments at very low population abundance. Detecting the presence of such an NIS can be very challenging, particularly as it spreads from the initial establishment site. This provides an opportunity to test detection limits using different approaches. This study tested the detection capability of 3 methods as zebra mussels (Dreissena polymorpha) spread from south to north through Lake Winnipeg, Manitoba, Canada. Zebra mussel veliger larvae were detected using cross-polarized light microscopy (CPLM), flow cytometry and microscopy (FlowCam), and conventional polymerase chain reaction (cPCR) analysis of environmental DNA (eDNA) on the same samples. Abundance generally declined from south to north in the lake but was lowest at Calder’s Dock (central). Although abundances could be quite low (i.e., <1 veliger/m3, Calder’s Dock) CPLM prevalence—the percentage of samples with at least one veliger—was high throughout the lake (99–100% of samples). Prevalence was lower for cPCR and FlowCam but was statistically associated with veliger abundance. Using standardized 3 mL subsamples (0.06–0.18 m3 of lake water sampled), all 3 methods had a high probability of veliger detection if large numbers of samples were processed. FlowCam was the most expensive method to process these 3 mL subsamples, while cPCR was least expensive and fastest. eDNA combined with intensive sampling is the most practical method for wide-scale monitoring programs for early detection. However, all 3 methods are complementary and could be deployed sequentially, with rapid initial sample processing using PCR, confirmation and density estimation with FlowCam, and detailed veliger counts using CPLM.

Evaluation of Surface Characteristics and Cytotoxicity of Dental Composites

The purpose of this study was to evaluate the surface and in vitro cytotoxicity on human dysplastic oral keratinocytes (DOK) of four commercial resins-based dental composites commonly used in prosthodontics dental therapies: two indirect composites for crown and bridges—SR Adoro (IvoclarVivadent GmbH) and Solidex (Shofu Dental GmbH); and two dual-curing luting resin cements—RelyxUnicem (3M ESPE Dental Products) and Variolink Esthetic DC (IvoclarVivadentGmbH). A complex assessment of surface characteristics of the four materials was conducted before and after the exposure to artificial saliva through various analyses, such as Scanning Electron Microscopy, Atomic Force Microscopy and Cross Polarized Light Microscopy (PLM). The results showed that DOK viability was not severely affected by exposure to any of these materials; however, Variolink expressed higher values but still above the toxicity level of the rest of the composites. The analysis of the surface structure between initial and artificial saliva exposed specimens returned a compact aspect in both categories and although Variolink and Relyx were subjected to increased roughness after saliva exposure, no damage of the internal compactness was recorded, demonstrating a fair behavior of the luting cements in contact with the saliva.

Effect of voids on thermomechanical cracking in lead-free Sn3Ag0.5Cu interconnections of power modules

In this article, the effect of voids on thermomechanically-induced failure in the lead-free solder interconnections of power amplifier (PA) modules is investigated. The interconnection of interest is between the module's flange and substrate (baseplate) initially having a large contact area, and whose cracking has strong detrimental effects on the RF performance and reliability of the PA. PA modules were attached onto baseplates with lead-free SAC305 solder and put to a thermal cycling test (TCT) in the 15 °C to 95 °C range. X-ray imaging was used to characterize the number of voids in interconnections after reflow soldering. The cross-sections of the pristine reflow-soldered and tested interconnections were inspected with cross-polarized light microscopy to reveal the shapes of the voids and crack paths. It was noted that the voids forming in interconnections during the final stages of the reflow process take an elliptical shape, leaning towards the module's outer edge, due to differences in the thermal expansions of the module and baseplate. During the TCT, high thermomechanical stresses caused localized recrystallization of the as-soldered SAC matrix in the vicinity of the elliptical voids. Finally, creep related intergranular cracks formed in these recrystallized areas. Anand's viscoplastic model was used to model the void formation and the effect of the formed voids on crack evolution in solder. Simulations of the high creep energy of the elliptical, tilted voids and their correlation with the recrystallization behavior of the SAC305 interconnections were consistent with the observed creep related failures. Confirmed by simulations, these voids are detrimental to the reliability of solder interconnection because the creep strain is strongly localized to the sharp curvatures of the elliptic void.

Interactions of Perfluorohexyloctane With Polyethylene and Polypropylene Pharmaceutical Packaging Materials

Semifluorinated alkanes (SFAs) are aprotic solvents, which may be used as drug solvents for topical ocular applications, for instance, in dry eye syndrome. Their physical properties suggest that they might be prone to interaction with plastic materials, such as, polyethylene (PE) and polypropylene (PP), which are commonly used as packaging materials for pharmaceutical products. In this study, we investigate interactions of PE and PP with a liquid SFA perfluorohexyloctane (PFHO) using differential scanning calorimetry (DSC), thermogravimetric analysis (TGA) and cross-polarized light microscopy. Binary phase diagrams of PFHO–PE and PFHO–PP systems demonstrating interactions of PFHO with the polymeric materials were constructed based on DSC data. According to this data, PFHO tends to lower the melting temperatures of PE and PP. The equilibrium values of solubilities of the polymers in PFHO and PFHO in the polymers were obtained by extrapolation of melting enthalpy data. Absorption of PFHO by PE and PP materials at ambient conditions after 4 weeks of equilibration was also studied by TGA. From the presented results, it may be concluded that thorough studies of interactions of PE or PP with SFAs are required when these materials are used as packaging components in SFA-based formulations.

Underpotential‐Assisted Electrodeposition of Highly Crystalline and Smooth Thin Film of Bismuth

AbstractIn this work, the electrodeposition of smooth bismuth thin films was investigated. Bismuth is known for its peculiar magnetic, thermal and electrical properties but the deposition of a uniform and flat film, which are features required for its application in electronic devices, is not trivial. We investigated the morphology of Bi film electrodeposited at increasing overpotential on a monocrystalline silver electrode. We found that the presence of an underpotential deposition (UPD) layer, previously deposited on the surface, drives the overpotential deposition to a smoother growth. The samples were investigated by means of different techniques: atomic force microscopy (AFM) and scanning electron microscopy combined with energy dispersive X‐ray spectroscopy (SEM‐EDS) to study the morphology, X‐ray photoemission spectroscopy (XPS) to assess the composition and X‐ray diffraction spectroscopy (XRD) to check the crystallinity. We also found an unexpected form birefringence behaviour, which has been preliminary investigated with cross polarized light microscopy (CPL).

Vancomycin ocular delivery systems based on glycerol monooleate reversed hexagonal and reversed cubic liquid crystalline phases

Lyotropic bulk reversed hexagonal and reversed cubic liquid crystalline phases (hexagonal and cubic phases) composed of glycerol monooleate (GM) were used to design the vancomycin hydrochloride’s (VHCl) delivery systems aiming to maintain VHCl’s therapeutic concentration during 24 h in the eye, locally (as an insert) and/or intravitreally (as a bulk phase injection).Bulk VHCl’s hexagonal and cubic phases were successfully prepared by melted homogenization and solvent evaporation method, and then an insert was prepared.The structural characteristics of liquid crystalline phases were studied using cross polarized light microscopy and small angle X-ray scattering technique. The presence of VHCl (1–9.5% w/w VHCl solution) did not exhibit any change in the liquid crystalline phase’s structure to another liquid crystalline phase, and showed little effect on the lattice parameter of the existing liquid crystalline phase structure.In order to relate the liquid crystalline phase structure to VHCl’s release rate locally into the eye, in-vitro release test of an implant has been done using a simulated tear fluid. VHCl’s release in the simulated tear fluid from the cubic phase obeyed Higuchi kinetics, with linear VHCl’s release versus the square root of time. The hexagonal phase released VHCl in simulated tear fluid significantly slower than the cubic phase. In order to relate the liquid phase structure to VHCl’s diffusion intravitreally, in vitro release test by the Sirius‘ Subcutaneous Injection Site Simulator (Scissor) has been performed. Comparing the release properties by a Scissor, the VHCl’s cubic phase demonstrated slower permeation through extra-cellular matrix than the VHCl solution.To evaluate the efficacy of the system investigated, the release properties of VHCl’s cubic phase were compared with literature data indicating that the cubic phase could be a potential matrix system in the delivery of VHCl intravitreally during 24 h after intravitreal injection. The release data in the simulated tear fluid indicated that the cubic phase should be further optimized to achieve a therapeutic VHCl concentration locally in the eye during 24 h.

Ultrafiltration membranes from polymerization of self-assembled Pluronic block copolymer mesophases

Industrially, ultrafiltration (UF) membranes are produced through non-solvent induced phase separation (NIPS), but due to environmental hazards inherent to the NIPS process as well as the low surface porosity and fouling resistance of membranes produced via this method, an alternative route to UF membranes is desirable. This work presents the self-assembly of Pluronic block copolymers in the presence of water and a monomeric phase as a new technique for the preparation of UF membranes without the need for organic solvent or post-modification. Different compositions of block copolymer, water, and monomer were polymerized to obtain both hexagonal and lamellar mesostructures, as indicated by small angle X-ray scattering (SAXS) and cross-polarized light microscopy. As-synthesized membranes were found to have pore sizes in the range of 3–4 nm with a molecular weight cutoff of 1500 g/mol and displayed both excellent fouling resistance and high permeance of water, vastly outperforming a conventional NIPS UF membrane. Further, in contrast to NIPS, the proposed method provides flexibility in terms of both the final membrane chemistry and pore size. As such, it is a versatile approach that can be easily tailored to produce membranes for a wide range of applications including wastewater treatment and food processing.

Solid State Characterization of Ciprofloxacin Liposome Nanocrystals.

Liposomes have been widely researched as drug delivery systems; however, the solid state form of drug inside the liposome, whether it is in solution or in a solid state, is often not studied. The solid state properties of the drug inside the liposomes are important, as they dictate the drug release behavior when the liposomes come into contact with physiological fluid. Recently, a new approach of making liposomal ciprofloxacin nanocrystals was proposed by the use of an additional freeze-thawing step in the liposomal preparation method. This paper aims to determine the solid state properties of ciprofloxacin inside the liposomes after this additional freeze-thawing cycle using cryo-TEM, small-angle X-ray scattering (SAXS), and cross-polarized light microscopy (CPLM). Ciprofloxacin precipitated in the ciprofloxacin hydrate crystal form with a unit cell dimension of 16.7 Å. The nanocrystals also showed a phase transition at 93 °C, which represents dehydration of the hydrate crystals to the anhydrate form of ciprofloxacin, verified by temperature-dependent SAXS measurements. Furthermore, the dependence of the solid state form of the nanocrystals on pH was investigated in situ, and it was shown that the liposomal ciprofloxacin nanocrystals retained their crystalline form at pH 6-10. Understanding the solid state attributes of nanocrystals inside liposomes provides improved understanding of drug dissolution and release as well as opening avenues to new applications where the nanosized crystals can provide a dissolution benefit.

The three-dimensional arrangement of the mineralized collagen fibers in elephant ivory and its relation to mechanical and optical properties

Elephant tusks are composed of dentin or ivory, a hierarchical and composite biological material made of mineralized collagen fibers (MCF). The specific arrangement of the MCF is believed to be responsible for the optical and mechanical properties of the tusks. Especially the MCF organization likely contributes to the formation of the bright and dark checkerboard pattern observed on polished sections of tusks (Schreger pattern). Yet, the precise structural origin of this optical motif is still controversial. We hereby address this issue using complementary analytical methods (small and wide angle X-ray scattering, cross-polarized light microscopy and scanning electron microscopy) on elephant ivory samples and show that MCF orientation in ivory varies from the outer to the inner part of the tusk. An external cohesive layer of MCF with fiber direction perpendicular to the tusk axis wraps the mid-dentin region, where the MCF are oriented mainly along the tusk axis and arranged in a plywood-like structure with fiber orientations oscillating in a narrow angular range. This particular oscillating-plywood structure of the MCF and the birefringent properties of the collagen fibers, likely contribute to the emergence of the Schreger pattern, one of the most intriguing macroscopic optical patterns observed in mineralized tissues and of great importance for authentication issues in archeology and forensic sciences. Statement of SignificanceElephant tusks are intriguing biological materials as they are composed of dentin (ivory) like teeth but have mineralized collagen fibers (MCF) similarly arranged to the ones of lamellar bones and function as bones or antlers. Here, we showed that ivory has a graded structure with varying MCF orientations and that MCF of the mid-dentin are arranged in plywood like layers with fiber orientations oscillating in a narrow angular range around the tusk axis. This organization of the MCF may contribute to ivory’s mechanical properties and, together with the collagen fibers birefringence properties, strongly relates to its optical properties, i.e. the emergence of a macroscopic checkerboard pattern, well known as the Schreger pattern.

Application of confocal laser scanning microscopy for the diagnosis of amyloidosis.

We analysed specificity and sensitivity of confocal laser microscopy (CLSM) on tissue sections for a diagnosis of amyloidosis, in an attempt to reduce technical errors and better standardise pathological diagnosis. We first set up a protocol for the use of CLSM on this type of specimen, using a group of 20 amyloid negative and 20 positive samples. Of all specimens, 2, 4 and 8-μm sections were cut. Sections were stained with Congo red (CR) and thioflavin-T (ThT) and observed by cross-polarised light microscopy (CR-PL), epifluorescence microscopy (CRF-epiFM and ThT-epiFM) and CLSM (CRF-CLSM and ThT-CLSM). To validate the method in a diagnostic setting, we examined tissue samples from 116 consecutive patients with clinical suspicion of amyloidosis, selected from the period 2005 to 2014 from the database of the Pathology Unit of the University of Padua. The results were compared with those of transmission electron microscopy (TEM), which we consider as reference. We found that with CRF-CLSM, the false negative rate was reduced from 17 to 5%, while the sensitivity of detection increased to 12%. The results were in complete agreement with those of TEM ThT-CLSM; both sensitivity and specificity were 100%. Finally, ThT-CLSM results did not vary with section thickness, and small amounts of amyloid could even be detected in 2-μm sections. In conclusion, we found ThT-CLSM to be more sensitive as a screening method for amyloidosis than CR and ThT epifluorescence optical imaging. The method was easier to standardise, provided images with better resolution and resulted in more consistent pathologist diagnoses.

Progesterone solubilization within reverse hexagonal mesophase

The present work reports on the preparation and characterization of progesterone-loaded reverse hexagonal (HII) mesophase based on a glycerol monooleate (GMO)/tricaprylin (TAG)/water mixture for transdermal application.Several compositions into which progesterone was quantitatively solubilized were selected and used in the present study in order to evaluate the effect of the hormone on structural transitions and characteristics.A GMO/TAG of 90:1wt ratio with 20wt% water was found to form HII mesophases with relative high solubilization capacity of 4wt% progesterone and a shelf life of over one year. The structures were identified by cross-polarized light microscopy (CPLM). Structural confirmations were obtained from small-angle X-ray scattering (SAXS) measurements. The hormone is solubilized at the interface of the HII mesophase as confirmed by SAXS, rheology, FTIR, and DSC. The progesterone, which is loaded onto reverse hexagonal cylinders, leads to strengthening the hydrogen bonding and to better packing of the HII mesophase, suggesting that progesterone is a kosmotropic agent.The reverse hexagonal mesophase seems to be a promising carrier for solubilization of progesterone.

Antimicrobial Activity from Colistin–Heparin Lamellar-Phase Complexes for the Coating of Biomedical Devices

Infections arising in hospitalized patients, particularly those who have undergone surgery and are reliant on receiving treatment through biomedical devices, continue to be a rising concern. It is well-known that aqueous mixtures of oppositely charged surfactant and polymer molecules can self-assemble to form liquid crystalline structures, primarily via electrostatically driven interactions that have demonstrated great potential as tailored-release nanomaterials. Colistin is a re-emerging antibiotic used against multidrug-resistant Gram-negative bacteria. Its amphiphilic structure allows it to form micellar aggregates in solution. Thus, the aim of this study was to determine whether structured complexes form between colistin and negatively charged biopolymers, such as the highly sulfated anticoagulant, heparin. Cross-polarized light microscopy and synchrotron small-angle X-ray scattering were employed to visualize the appearance of birefringent structures and identify liquid crystalline structures, respectively, formed across the interface between solutions of colistin and heparin. A lamellar phase with a lattice parameter of ∼40 Å was formed upon contact between the oppositely charged solutions of colistin and heparin. In addition, in vitro release studies showed a slow release of colistin from the lamellar-phase gel complexes into the bulk media, and disk diffusion bioassays revealed antimicrobial activity against Pseudomonas aeruginosa. This system provides a novel, cost-effective, and simple approach to reducing the risk of infections by potentially applying the formulation as a coating for biomedical implants or tubing.