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Waste products in the bloodstream are filtered by the glomerular capillaries in the kidneys and excreted into the urine. When making a differential diagnosis of kidney diseases, structural assessment of glomeruli using histological, ultrastructural, and immunological studies is crucial. This study assessed the microscopic and ultrastructural morphometric parameters of glomerular capillaries and examined their correlation with serum creatinine and proteinuria. A total of 60 kidney biopsy cases received by the transmission electron microscope (TEM) laboratory for diagnosis were included in the study. Toluidine blue stained 300 nm thick sections of TEM tissue blocks were scanned for glomerular morphometry by a whole slide imaging system, and the estimation of Bowman's capsule (BC) area, glomerular capillary lumen diameter (GCLD), glomerular capillary density (GCD), glomerular capillary surface area density (GCSA), and percentage of glomerular capillary lumen space (%GCLS) was performed with QuPath software. TEM images of 70 nm thick sections were used for the evaluation of endothelial fenestration diameter (EFD), glomerular basement membrane (GBM) thickness, and podocyte foot process (PFP) effacement. Proteinuria and serum creatinine showed positive correlations with GBM thickness and PFP effacement. Negative correlations of serum creatinine were observed with EFD, %GCLS, and GCSA. Hence, glomerular filtration is greatly affected by the total area of the glomerular capillary surface and structural changes of GBM. Reduction of glomerulus filtration due to foot process effacement and thickening of GBM results in damage to the filtration barrier leading to the leakage of plasma protein into urine. [ABSTRACT FROM AUTHOR]

Previous studies have revealed that Propane-2-sulfonic acid octadec-9-enyl-amide(N15) exerts a protective role in the inflammatory response after ischemic stroke and in neuronal damage. However, little is known about N15 in Alzheimer's disease (AD). The aim of this study was to investigate the effects of N15 on AD and explore the underlying molecular mechanism. AD mice model was established by lateral ventricular injection with Aβ25–35. N15 was daily intraperitoneal administered for 28 days. Morris Water Maze was used to evaluate the neurocognitive function of the mice. The expression of PPARα/γ, brain-derived neurotrophic factor (BDNF), Neurotrophin-3 (NT3), ADAM10, PS1 and BACE1 were measured by qPCR. Aβ amyloid in the hippocampus was measured by Congo red assay. Toluidine blue staining was used to detect the neuronal apoptosis. Protein levels of ADAM10, PS1 and BACE1 were determined using immunoblotting. N15 treatment significantly reduced neurocognitive dysfunction, which also significantly activated the expression of PPARα/γ at an optimal dose of 200 mg/kg. Administration of N15 alleviated the formation of Aβ amyloid in the hippocampus of AD mice, enhanced the BDNF mRNA expression, decreased the mRNA and protein levels of PS1 and BACE1, upregulated ADAM10 mRNA and protein levels. N15 exerts its neuroprotective effects through the activation of PPARα/γ and may be a potential drug for the treatment of AD. [ABSTRACT FROM AUTHOR]

Bone marrow stem cells (BMSCs) are recognized for their robust proliferative capabilities and multidirectional differentiation potential. Ectopic endochondral ossification of BMSC-generated cartilage in subcutaneous environments is a concern associated with vascularization. Hence, devising a reliable strategy to inhibit vascularization is crucial. In this study, an anti-angiogenic drug, curcumin (Cur), was encapsulated into gelatin to create a porous Cur/Gelatin scaffold, with the aim of inhibiting vascular invasion and preventing endochondral ossification of BMSC-regenerated cartilage. In vitro wound healing tests demonstrated that a 30 μM Cur solution could inhibit the migration and growth of human umbilical vein endothelial cells without impeding BMSCs migration and growth. Compared to the gelatin scaffold, our findings verified that the Cur/Gelatin scaffold significantly inhibited vascular invasion after being subcutaneously implanted into rabbits for 12 weeks, as evidenced by gross observation and immunofluorescence CD31 staining. Moreover, both the porous gelatin and Cur/Gelatin scaffolds were populated with BMSCs and underwent in vitro chondrogenic cultivation to produce cartilage, followed by subcutaneous implantation in rabbits for 12 weeks. Histological examinations (including HE, Safranin-O/Fast Green, toluidine blue, and immunohistochemical COL II staining) revealed that the BMSC-generated cartilage in the gelatin group exhibited prominent endochondral ossification. In contrast, the BMSC-generated cartilage in the Cur/Gelatin group maintained cartilage features, such as cartilage matrix and lacunar structure. This study suggests that Cur-loaded scaffolds offer a reliable platform to inhibit endochondral ossification of BMSC-generated cartilage. [ABSTRACT FROM AUTHOR]

In tissue engineering bioresorbable polymers are commonly used to mimetize a damaged tissue characteristics. In spite of that, we did not find a comparative study of the response of the same cell to dense, porous, and fibrous biomaterials. The present work aimed to study the in vitro of pure polymers [poly (ε-caprolactone) (PCL) and poly (hydroxybutyrate-co-hydroxyvalerate) (PHBV)] and their 75:25 blend in dense, porous, and fibrous forms in Vero cells, a fibroblast cell line. The biomaterials were characterized morphologically by scanning electron microscopy (SEM). The MTT assay was used for analysis of possible cytotoxicity. Vero cells were cultured on biomaterials for 120 h and submitted to SEM, morphological and cytochemical analyses. The dyes used were crystal violet (morphology), toluidine blue at pH 4.0 (for nucleic acids and glycosaminoglycans) and 2.5 (for glycosaminoglycans only), periodic acid-Schiff reactive (for neutral sugars), xylidine ponceau at pH 2.5 (for total proteins), and picrosirius/hematoxylin (for selective collagen analysis). The biomaterials studied were not considered citocytotoxic. The cells were able to adhere to and proliferate on the substrates. The cytochemical data indicate that the cells have high functional activity, with high protein synthesis on the scaffolds but do not accumulate acid or neutral sugars. The cells on the fibrous polymers appear to be more elongated, following the shape of the fibers. Regarding the scaffold form (porous or fibrous), the in vitro data do not show great superiority of one type to the other. Then, all the polymers tested are suitable for tissue engineering. [ABSTRACT FROM AUTHOR]

Increased discharge of dyes into the environment and consequent contamination of water bodies have been issues of public health concern due to the toxicity of dyes. In this study, the ability of Stenotrophomonas sp. CFB-09 to decolorize lignin-mimicking dyes [Congo red (CR), toluidine blue (TB), methyl blue (MB), remazol brilliant blue R (RBBR) and indigo carmine (IC)] and produce ligninolytic enzymes were investigated. Dye decolorization activity was carried out qualitatively on plates and quantitatively by UV–Vis spectrophotometric analysis of broths. Influence of process parameters (initial pH, incubation temperature and media composition) on dye decolorization were also determined. Identification of biodegradation products was done by GC-MS. Results revealed that Stenotrophomonas sp. CFB-09 decolorized dyes by concomitantly producing ligninolytic enzymes and achieved ≥90% decolorization of 25 mg/L CR, MB, RBBR, and IC and 75.0–86.5% decolorization of 100 mg/L of IC, CR and MB at 120 h. Lignin peroxidase exhibited the highest activity among ligninolytic enzymes studied on all dyes. Optimum decolorization for all dyes was achieved at pH 8.0 and 35 °C in the presence of oil palm empty fruit bunch (OPEFB). Findings show that the ligninolytic bacterium, Stenotrophomonas sp. CFB-09 has potentials for application in bioremediation of dye containing effluents. [ABSTRACT FROM AUTHOR]

Retinal and optic nerve damage in glaucoma involves excitotoxicity via N-methyl-D-aspartate (NMDA) receptors. Since, trans-resveratrol (TR) is known to provide neuroprotection, we investigated its protective effects against NMDA-induced retinal and optic nerve injury. Sprague Dawley rats were divided into four groups which received vehicle (PBS), NMDA, and TR 0.4 or TR 4 nmol 24 h prior to NMDA, unilaterally and intravitreally. Seven days post-injection, rats were euthanized; eyeballs were enucleated and subjected to hematoxylin and eosin and terminal transferase dUTP nick end labeling staining while optic nerves were isolated for toluidine blue staining. Retinal morphometry showed that ganglion cell layer (GCL) layer thickness within inner retina (IR), retinal cell count (RCC) per 100-µm length of GCL, RCC per 100-µm2 area of GCL, and RCC per 100 µm2 of IR were significantly higher in both TR-treated groups compared to the NMDA group. No differences were observed between the two dose groups. Optic nerve morphology was in accordance with the retinal morphology whereby TR-treated groups showed significantly lesser degenerative changes compared to NMDA-treated group. TR protects against NMDA-induced changes in retinal and optic nerve morphology by preventing retinal cell apoptosis. [ABSTRACT FROM AUTHOR]

Mast cells in the brain are associated with increased inflammation during the acute period following exposure to infection; these cells are important for destroying the infectious agents. We investigated the relation between histopathological lesions and mast cells in sheep brains infected with Listeria. Pons and medulla regions from 17 infected and eight normal sheep brains were examined. Microabscesses and perivascular infiltration were assessed for histopathology. Mast cells were identified using toluidine blue and Listeria monocytogenes were investigated immunohistochemically. We found a significant increase in mast cells in infected sheep brains that was related directly to the extent of brain lesions. A strong correlation was found between mast cells and microabscess formation. A correlation between bacteria level and brain lesions also was observed, but not between bacteria level and mast cells. Our findings indicate that mast cells are increased following Listeria infection in sheep in proportion to the severity of brain lesions; the increase may contribute to acute inflammatory reactions and also may destroy bacteria directly. [ABSTRACT FROM AUTHOR]

Mast cells are immune cells of myeloid lineage, characterized by the presence of cytoplasmic granules. These cells play a significant role in multiple pathologies, such as urticaria and type 1 hypersensitivity reactions. Mast cells in tissue sections can be demonstrated with metachromatic stains, such as toluidine blue. Metachromatic staining of mast cells is influenced by the type of mast cell, pH of the staining solution and fixative employed. The objective of this study is to identify a simple, consistent, and reproducible toluidine blue staining method to quantify the mast cells in whole slide skin images using automated image analysis. Skin sections from naive mice and atopic dermatitis mice were stained with toluidine blue methods by Churukian-Schenk and Luna. Luna's toluidine blue staining method without the eosin counterstaining provided optimal staining of mast cells with a greater contrast to the background. The Churukian-Schenk toluidine blue method resulted in slight background staining which confounded the accurate detection and quantification of mast cells in mouse skin sections using an automated image analysis algorithm. Using the Luna toluidine blue stain, a 5-fold increase in the total number of mast cells was observed in atopic dermatitis skin samples as compared to naive mice. In summary, a simple, conventional, and reproducible toluidine blue method was identified to quantify the mast cells in mouse skin sections using an automated image analysis algorithm. [ABSTRACT FROM AUTHOR]

Cytology specimens and biopsy tissues are frequently small and pale, making them difficult to visualize grossly in paraffin. Ten dyes were assayed on small tissues to determine if specimen discernibility could be increased during the embedding and microtomy steps in the histological process. The ideal dye should not remain visible in a tissue section microscopically after subsequent staining and must not interfere with immunohistochemistry (IHC) assays. This study found that Harris hematoxylin and 1% aq. toluidine blue solution were the best labelers for gross tissue visualization and did not adversely affect post-processing staining and IHC assays. [ABSTRACT FROM AUTHOR]

Allergic dermatitis (AD) is a chronic inflammatory skin disease that a variety of immune cells are involved in the progression of AD. Among them, T cells are one of major players of AD pathogenesis. The V-domain Ig suppressor of T cell activation (VISTA) has been reported that it has a potential immunomodulatory for T cell response. This study aimed to investigate immunomodulatory of recombinant VISTA-Ig fusion protein in AD mice model. The model of AD was built with oxazolone (OXA) in BALB/c mice, then VISTA-Ig was used to treat AD by intraperitoneal (IP) injection. The ear thickness was measured by a digital thickness gauge. The ears tissues were collected and subjected to hematoxylin-eosin (H&E) and toluidine blue (TB) staining. The secretion levels of IL-4 and IgE in the serum were measured by enzyme linked immunosorbent assay (ELISA). The mRNA expression levels of inflammatory cytokines (IL-1β, IL-6, IL-12, and INF-γ) in ear tissues were measured using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). Treatment with VISTA-Ig successfully alleviated the symptoms of AD, such as erythema, horny substance, and swelling. The infiltration of inflammatory cells was significantly reduced following VISTA-Ig therapy. The secretion levels of IL-4 and IgE in the serum were significantly attenuated following treatment with VISTA-Ig. Additionally, VISTA-Ig observably down-regulated inflammatory cytokines expression in ear tissues. Taken together, our results showed that VISTA-Ig possessed the potential to be a novel immunomodulatory candidate drug against AD. [ABSTRACT FROM AUTHOR]

Aim: The aim of this study was to evaluate the effects of standard culture medium and chondrogenic differentiation medium with PRP on chondrogenic differentiation of rabbit dental pulp-derived mesenchymal stem cells (rabbit DPSCs) that are transfected with transforming growth factor-beta 1 (TGF-B1) gene, based on the hypothesis of TGF- B1 and PRP can be effective on the chondrogenesis of stem cells. Materials and Methods: Rabbit DPSCs were characterized by using flow cytometry, immunofluorescent staining, quantitative Real Time Polymerase Chain Reaction (qRT-PCR) and differentiation tests. For the characterization, CD29, CD44 and CD45 mesenchymal cell markers were used. Rabbit DPSCs were transfected with TGF-B1 gene using electroporation technique in group 1; with PRP 10% in group 2; with chondrogenic medium in group 3; with both chondrogenic medium and PRP in group 4. DPSCs were cultured in medium with 10% inactive PRP in group 5, chondrogenic medium in group 6, chondrogenic medium with PRP 10% in group 7. SOX9, MMP13 and Aggrecan gene expression levels were evaluated in 3, 6, 12. and 24. days by qRT-PCR. Results: The expression levels of SOX9, MMP13 and Aggrecan were higher in group 2, 3 and group 7 in 3th day however in 24th day group 7 and group 2 were found higher. The expression levels changed by time-dependent. The extracellular matrix of the cells in experimental groups were positively stained with safranin O and toluidine blue. Conclusion: The combination in culture medium of TGF-B1 gene transfection and 10% PRP accelerates the chondrogenic differentiation of DPSCs. [ABSTRACT FROM AUTHOR]

Iron-oxide nanoparticles are one of the most commercialized nanomaterials and have gained widespread acceptance in nanotherapeutics due to their ability for targeted drug delivery, bioimaging, and various other preclinical and clinical theranostic biomedical applications. However, the absence of regulations, guidelines, and harmonized standards as well as limitations associated with their use in clinical settings in the context of their safety and toxicity profiling necessitates in-depth understanding of their toxicological paradigms. Here we examine the toxicity of modified superparamagnetic iron oxide nanoparticles in Swiss albino mice in terms of body weight changes, organ coefficients, generalized and organ-specific biochemical, and various histological staining parameters after administration of bare (uncoated) magnetic nanoparticles (MNPs) and triple polymer-coated magnetic nanoparticles (MNP-AC-G2-pPEG). Both types of nanoparticles were administered intravenously, in three doses (5, 10, and 25 mg/kg body weight) and results of biochemical and histopathological assessment revealed that the highest dose of bare (uncoated) MNPs significantly altered biochemical and histoarchitectural aspects in vital organs, while coated NPs (MNP-AC-G2-pPEG) was found safe in almost all doses. Furthermore, results of toluidine blue (for mast cells) and Prussian blue (for iron deposition) staining also established that the highest dose administration of bare MNPs in animals significantly enhanced mast cell infiltration and iron deposition in tissue sections of most vital organs, while coated NPs did not demonstrate any such adverse effects. Overall, outcomes of this study aid in establishing that administration of uncoated (bare) magnetic NPs in vivo results in structural and functional toxicological modifications while the coating of these NPs with biocompatible and biodegradable polymers can significantly bring down the toxicity of these NPs. [ABSTRACT FROM AUTHOR]

Lecithin is a natural mixture of phospholipids and neutral lipids, and plays a vital role in the maintenance of cell-membrane integrity for all of the basic biological processes.This study aimed to evaluate the adhesion and growth of bone marrow-derived mesenchymal stem cells (BMSCs) on the three-dimensional (3D) electrospun lecithin-containing poly (L-lactic acid) (PLLA) fibrous scaffolds. In this study, PLLA and 0-15% of lecithin (wt.%) were dissolved in methylene chloride and thoroughly mixed to fabricate fibrous scaffolds by electrospinning technique. Surface morphology and hydrophilic properties of the scaffolds were characterized by scanning electron microscopy (SEM) and water uptake, respectively. The cytocompatibility of electrospun scaffolds and their potential cytotoxic effects on cells were studied over 8 days by seeding rat BMSCs on the scaffolds of PLLA containing 0-15% lecithin. Then cell proliferation was tested by CCK-8 assay and cell injury was assessed by measuring the release of intracellular lactate dehydrogenase (LDH). And cell morphology was studied by Toluidine blue staining, propidium iodide staining and SEM. The hydrophilicity was dramatically increased with an increase of lecithin content in the modified PLLA/lecithin scaffolds, as determined by rate of water uptake. In addition, the results obtained from in vitro assays suggested that the electrospun PLLA/5%lecithin scaffolds could possess the optimal hydrophilicity, higher adhesion and proliferation of BMSCs than PLLA, and they also did not lead to any significant toxic effects to cells. Moreover, BMSCs on PLLA/5%lecithin scaffolds tended to maintain their phenotypic shape and integrate with the microfibers to form a 3D cellular network, indicating a favorable interaction between this electrospun lecithin-based scaffolds and cells. The study demonstrated that the electrospun PLLA/lecithin fibrous scaffolds showed much better hydrophilicity and biocompatibility by introducing lecithin into the polymer in comparison with pure PLLA, which would offer a promising strategy for constructing tissue engnineered blood vessel. [ABSTRACT FROM AUTHOR]

Atopic dermatitis (AD) is a chronic skin inflammatory disease characterized by disequilibrium between Th1/Th2 lymphocytes. Helicobacter pylori neutrophil-activating protein (HP-NAP) has been reported that it has the potential immunomodulatory effect able to regulate the Th1/Th2 balance. This study aimed to investigate the therapeutic effect of HP-NAP in AD mice model. The model of AD was built with oxazolone (OXA) in BALB/c mice, then HP-NAP was used to treat AD by intraperitoneal injection. Ear thickness was measured by a digital thickness gauge. The ears tissues were collected and subjected to hematoxylin-eosin (H&E) and toluidine blue (TB) staining. The mRNA expression levels of inflammatory cytokines (IL-1β, IL-5, IL-6, and TNF-α) in ear tissue were measured using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). The secretion of IgE, IL-4, and IFN-γ was measured by enzyme-linked immunosorbent assay (ELISA). Treatment with HP-NAP successfully alleviated the symptoms of AD, such as erythema, horny substance, and swelling. The infiltration of lymphocytes and mast cells were significantly reduced following HP-NAP therapy. The secretion of IgE and IL-4 was significantly attenuated following treatment with HP-NAP. Additionally, HP-NAP observably downregulated inflammatory cytokine expression (e.g. IL-1β, IL-5, IL-6, and TNF-α) in ear tissues. Taken together, our results showed that HP-NAP possessed the potential to be a novel immunomodulatory candidate drug against AD. [ABSTRACT FROM AUTHOR]

Ephedrine has various side effects in the cardiovascular and nervous systems. However, the cellular mechanism of toxicity remains unknown, specifically on the retina. This study was to investigate effects of ephedrine on the retina and explore the underlying mechanisms. Sprague Dawley rats were treated with ephedrine (n = 10) or saline (n = 10) by oral gavage for seven days. The retinal morphology was evaluated by Toluidine blue staining. Apoptosis-related markers were detected in the retinal lysate. Enzyme-linked immunosorbent assays were used to measure neurotransmitters and oxidative stress markers. Real-time PCR and western blot were used to measure gene and protein expression, respectively. Our results demonstrated that ephedrine induced apoptosis in the retina, increased dopamine level as well as oxidative stress, and down-regulated the Jak2/Stat3 gene expression as well as protein expression of p-JAK2/p-STAT3. Our study indicated that ephedrine treatment caused retinal damage in SD rats, which may be associated with the JAK2/STAT3 pathway. [ABSTRACT FROM AUTHOR]

Purpose: Lysergic acid diethylamide (LSD) is a powerful hallucinogen with high potential for abuse. There is far less known about its effects on the retina, especially the underlying mechanisms. This study was to investigate the acute toxicity of LSD on the retina of C57 mice and its mechanisms of action. Methods: C57 mice were treated with LSD at progressively increasing doses (0.2–1.2 mg/kg) intraperitoneally two times daily for 5 days, mice treated with saline served as negative control. Electroretinography (ERG) was used to test the function of the retina. Toluidine blue staining was used to detect the morphology of the retina. Enzyme-linked immunosorbent assay (ELISA) was used to measure the apoptosis-related factors. Real-time PCR and western blot techniques were used to measure expression changes of genes and proteins, respectively. Results: LSD treatment caused retinal damage, as shown by a decrease in ERG response and the loss of photoreceptor cells. LSD treatment also increased apoptosis through up-regulating the expression of p-JAK1/p-STAT1. Conclusions: Our study indicated that intraperitoneal administration of LSD-induced retinal damage of C57 mice, at least partially through regulating the JAK/STAT pathway. [ABSTRACT FROM AUTHOR]

A new choline oxidase (ChO) and toluidine blue O (TBO) based amperometric choline biosensor was reported in this article. An amperometric choline biosensor with immobilization of TBO (as a mediator), ChO onto polypyrrole–polyvinylsulphonate (PPy–PVS) film was accomplished on the surface of a platinum electrode. ChO was immobilized on PPy–PVS film by cross-linking with glutaraldehyde (GA). TBO was used as the mediator. Choline is oxidized to betaine and hydrogen peroxide in an oxygenated environment by ChO. Mediator reduced by reaction with hydrogen peroxide. The amperometric response was based upon the electrocatalytic properties of TBO. Optimum pH and temperature values were 7.0 and 30 °C, respectively. There was linearity between 1.0 × 10−8 and 2.0 × 10−8 M (R2 = 0.9805). The detection limit of the biosensor was 1.0 × 10−9 M and response time of the biosensor was 200 s. The storage stability and reproducibility of the biosensor were also investigated. Interfering effect of several interferants such as ascorbic acid, uric acid, alanine, dopamine, paracetamol, cysteine, and glucose on the choline biosensor was examined. The developed biosensor was tested in determinations of content in a synthetic blood sample. [ABSTRACT FROM AUTHOR]

We developed a novel sectioning and staining method to make high contrast, high resolution sections of plant tissue for light microscopy. Specimens of teosinte (Zea mays L., ssp. mexicana) root tips were fixed and embedded in Technovit 7100™ plastic resin. Thin sections, 1−2.5 μm, were cut and mounted on glass slides. The sections were either treated with RNase or not, then stained with 0.1% toluidine blue O and observed through ∞/0 objective lenses. For light microscopy, the enzyme staining procedure increased resolution and contrast. High magnification ∞/0 objective lenses produced high quality images for digital photography without using a coverslip or immersion oil. Our slide preparation and microscopic analysis were less labor intensive and more rapid than previous methods and enabled rapid and precise alignment of serial transverse sections for both tracking cell lineages and tissue measurements. [ABSTRACT FROM AUTHOR]

Mast cells are large cells with granular cytoplasm that participate in wound healing, angiogenesis and defense against pathogens. They also contribute to inflammation by initiating innate and acquired immunity. The granules of these cells exhibit characteristic staining properties. We investigated toluidine blue, astra blue, Alcian blue-pyronin Y and May-Grunwald Giemsa stains for mast cells in various oral lesions and assessed the efficacy of each for identifying mast cells. Sections were obtained from 10 each of diagnosed cases of inflammatory fibrous hyperplasia, periapical cyst, mild dysplasia, oral submucous fibrosis and oral squamous cell carcinoma and stained using the stains listed above. Mast cells were assessed for their presence, contrast of the mast cell in the connective tissue background and number. We found that May-Grunwald Giemsa stain was the best for identification of mast cells, although toluidine blue staining is less time-consuming. Overall we obtained better results using May-Grunwald Giemsa and toluidine blue for staining mast cells. [ABSTRACT FROM AUTHOR]

Although hematoxylin and eosin (H & E) staining of sectioned embryonic insect material is widely used, it is time-consuming and may not provide sufficient information. We evaluated new staining procedures for embryonic whole mounts of the greater wax moth, Galleria mellonella. We compared a combination of toluidine blue and rhodamine B (TB-RB) to H & E; we also investigated calcofluor white (CFW) staining. TB-RB staining produced staining similar to H & E. TB-RB staining was less time-consuming and improved visualization of the blastoderm and its differentiation into the germ disk and serosa membrane. CFW enhanced details of mitosis in nuclei post-fertilization and stained the primary serosal membrane. Staining of whole mounts with TB-RB and CFW enabled embryonic staging that was more rapid, convenient and effective than the routine approach using H & E and fluorescent probes. [ABSTRACT FROM AUTHOR]

In the present study, the antimicrobial and antibiofilm efficacy of toluidine blue (TB) encapsulated in mesoporous silica nanoparticles (MSN) was investigated against Pseudomonas aeruginosa and Staphylococcus aureus treated with antimicrobial photodynamic therapy (aPDT) using a red diode laser 670 nm wavelength, 97.65 J cm−2 radiant exposure, 5 min). Physico-chemical techniques (UV-visible (UV-vis) absorption, photoluminescence emission, excitation, and FTIR) and high-resolution transmission electron microscopy (HR-TEM) were employed to characterize the conjugate of TB encapsulated in MSN (TB MSN). TB MSN showed maximum antimicrobial activities corresponding to 5.03 and 5.56 log CFU ml−1 reductions against P. aeruginosa and S. aureus, respectively, whereas samples treated with TB alone showed 2.36 and 2.66 log CFU ml−1 reductions. Anti-biofilm studies confirmed that TB MSN effectively inhibits biofilm formation and production of extracellular polymeric substances by P. aeruginosa and S. aureus. [ABSTRACT FROM AUTHOR]

Fibrous dysplasia (FD) and ossifying fibroma (OF) are fibro-osseous lesions (FOLs) having several overlaps that may make final diagnosis difficult by hematoxylin and eosin (H/E) alone. This study seeks to detect any association between Azan and Toluidine blue staining as compared with osteocalcin in FD and OF diagnosis. Forty formalin fixed paraffin embedded (FFPE) blocks of FD and OF were prepared for Azan, Toluidine blue and osteocalcin staining. Brown staining of calcified structures was considered as positive for osteocalcin. Scoring for Azan and Toluidine blue was evaluated based on intensity and localization. Level of agreement of original and revised diagnosis was determined. Six (40%) of 15 FD were corroborated by osteocalcin. Eight cases initially diagnosed as OF were revised to FD. There were 25 OF according to H/E, and 17 (68%) were validated by osteocalcin. Measure of agreement between histology and immunohistochemistry was 0.081; p =.608. Eleven (42.3%) OF expressed strong toluidine blue staining of the intervening fibrous connective tissue stroma while only 2 (14.2%) FD showed similar staining, this difference was statistically significant [p =.001]. Histomorphometric analysis with Toluidine blue may reduce diagnostic errors of OF and FD. [ABSTRACT FROM AUTHOR]

The aim of this study was to investigate the ability of Lemna minor for Toluidine Blue (TB) removal. Influence of the initial concentration over the removal process was considered. Experimental data have been analyzed using Langmuir, Freundlich, Dubinin-Radushkevich (D-R), and Elovich isotherm models. In addition, several kinetic models, pseudo-first-, pseudo-second-order, intraparticle and film diffusion models were considered. Langmuir and Freundlich isotherm suggested a favorable adsorption of TB by Lemna minor plants. From the D-R the mean free energy was calculated to be 11.18 kJ/mol, which indicates that TB adsorption was characterized by a chemisorption process. Kinetic studies showed that liquid film diffusion plays an important role during the process. Adsorption capacities of up to 26.69 mg/g and a high capacity of adaptation indicated that phytoremediation using Lemna minor could be a valuable alternative for dyes removal from wastewaters. [ABSTRACT FROM AUTHOR]

Feasible method of quantitative determination of anionic polymers in aqueous solutions was described using poly(styrene sulfonate) and poly(acrylate) sodium salts as an example. The method consists in absorbance measurements while titrating with thiazine dye. Volume of dye solution needed to attain specified value of absorbance is linearly dependent on polymer concentration. Optimal wavelengths are 666 nm and 640 nm for Methylene Blue dye and Toluidine Blue dye, respectively. Calibration plots are straight lines (r2 > 0.99) ensuring good accuracy of the method developed. Range of measurable concentrations is dependent on chosen value of end-point absorbance. [ABSTRACT FROM AUTHOR]

Gastric cancer has been strongly associated with presence of the bacterium Helicobacter pylori. To improve techniques in identifying H. pylori so that gastric cancer may be predicted early, this project was formulated to determine whether one particular stain is more effective in displaying H. pylori microscopically. In addition, this study attempted to determine whether the degree of inflammatory elements present in tissue could be used to predict the likelihood of H. pylori presence. Protocols for the staining techniques, Steiner and alcian yellow/toluidine blue (AY/TB), were employed on specimens to semi-quantitate H. pylori presence. Serial sections from the same specimens were stained with hematoxylin and eosin to determine the amount of inflammation. Spearman rho correlation was used to evaluate the association between amount of H. pylori and inflammation in each case. It was determined that AY/TB was more easily performed, more effective in demonstrating H. pylori, and more cost effective than the Steiner stain. Additionally, it was determined that a moderate positive association was indicated between high levels of inflammation and marked presence of H. pylori. [ABSTRACT FROM AUTHOR]

The interaction of toluidine blue (TB) with cyclodextrins (CDs), including ß-cyclodextrin (ß-CD) and carboxymethyl-ß-cyclodextrin (CM-ß-CD), and calf thymus DNA in aqueous solution is studied by ultraviolet-visible absorption and steady-state fluorescence technique. The interactive model of TB with double-stranded DNA has been investigated by means of the inclusive action of ß-CD and CM-ß-CD. Based on the changes of absorption, fluorescence and resonance light scattering (RLS) spectra, the intrinsic binding constant (Kap) and the binding site number (n) of TB with DNA and the inclusion complexes TB--CD with DNA are obtained in the case of 20 mmol L-1 Tris-HCl buffer solution (pH 7.2). According to the experimental results, it can be inferred that the interactive model of dimer TB with DNA is 'electrostatic binding', while the monomer TB with DNA is 'intercalative binding'. [ABSTRACT FROM AUTHOR]

Purpose of this study: To elucidate the origin of cell populations that contribute to rotator cuff healing, we developed a mouse surgical model where a full-thickness, central detachment is created in the supraspinatus. Materials and methods: Three different inducible Cre transgenic mice with Ai9-tdTomato reporter expression (PRG4-9, αSMA-9, and AGC-9) were used to label different cell populations in the shoulder. The defect was created surgically in the supraspinatus. The mice were injected with tamoxifen at surgery to label the cells and sacrificed at 1, 2, and 5 weeks postoperatively. Frozen sections were fluorescently imaged then stained with Toluidine Blue and re-imaged. Results: Three notable changes were apparent postoperatively. (1) A long thin layer of tissue formed on the bursal side overlying the supraspinatus tendon. (2) The tendon proximal to the defect initially became hypercellular and disorganized. (3) The distal stump at the insertion underwent minimal remodeling. In the uninjured shoulder, tdTomato expression was seen in the tendon midsubstance and paratenon cell on the bursal side in PRG4-9, in paratenon, blood vessels, and periosteum of acromion in SMA-9, and in articular cartilage, unmineralized fibrocartilage of supraspinatus enthesis, and acromioclavicular joint in AGC-9 mice. In the injured PRG4-9 and SMA-9 mice, the healing tissues contained an abundant number of tdTomato+ cells, while minimal contribution of tdTomato+ cells was seen in AGC-9 mice. Conclusions: The study supports the importance of the bursal side of the tendon to rotator cuff healing and PRG4 and αSMA may be markers for these progenitor cells. [ABSTRACT FROM AUTHOR]

Fabrication and transplantation of tissue-engineered bones in a rotating wall vessel bioreactor (RWVB) was studied in the present study aiming to repair segmental bone defects. Osteoblasts were transfected with green fluorescent protein prior to seeding on bio-derived porous bone scaffolds at a density of 1 × 106 cells/mL and cultured in an RWVB for one week. For comparison, constructs were also cultured in a static condition. Morphology and structure of fabricated bones were examined using an inverted microscope, scanning electron microscope and histology analysis via hematoxylin–eosin and toluidine blue staining. Moreover, an animal model for repairing segmental bone defects of a Zelanian rabbit was used to assess the efficacy and biosafety of fabricated bones. In conclusion, tissue-engineered bones grew favorably in RWVB. In animal study, a preliminary repair of bone defects was noticed only in the experimental group after 4 weeks of implantation. Using RWVB, the fabricated tissue-engineered bone constructs were approved with better bio-capability in repairing the segmental bone defect. [ABSTRACT FROM AUTHOR]

A new, highly specific toluidine blue assay for the determination of sulfated polysaccharides is reported. Sulfated polysaccharides may be specifically determined at pH 1 in the presence of carboxylated polysaccharides and phosphorylated molecules such as nucleic acids. In addition, performing the assay at pH 1 extended the linear dynamic range for fucoidan up to 3 g/L but lowered the sensitivity in comparison with pH 7. For a color change at neutral pH, at least a sulfated disaccharide is required, whereas at pH 1, the detection was negative even for a sulfated maltose. A comparison of the fucoidan concentration in crude algae determined by the toluidine blue assay at pH 1 and gel permeation chromatography confirmed the accuracy and specificity of the colorimetric method. The limit of quantitation for the toluidine blue assay at pH 1 was 0.62 g/L for fucoidan. For a deeper understanding of the interaction between the sulfated polysaccharides and the toluidine blue, spectroscopic constants were evaluated by Ketelaar plots for sulfate esters. The maximum value for the molar absorptivity was 4.07 × 104 L/(mol cm), indicating strong affinity. Similar behavior was obtained for fucoidan at pH 7 with a value of 4.46 × 103 L/mol. [ABSTRACT FROM AUTHOR]

The objective of this study was to evaluate the anti-biofilm efficacy of photodynamic therapy by conjugating a photosensitizer (TBO) with silver nanoparticles (AgNP).Streptococcus mutanswas exposed to laser light (630 nm) for 70 s (9.1 J cm−2) in the presence of a toluidine blue O–silver nanoparticle conjugate (TBO–AgNP). The results showed a reduction in the viability of bacterial cells by 4 log10. The crystal violet assay, confocal laser scanning microscopy and scanning electron microscopy revealed that the TBO–AgNP conjugates inhibited biofilm formation, increased the uptake of propidium iodide and leakage of the cellular constituents, respectively. Fluorescence spectroscopic studies confirmed the generation of OH•as a major reactive oxygen species, indicating type I phototoxicity. Both the conjugates down-regulated the expression of biofilm related genes compared to TBO alone. Hence TBO–AgNP conjugates were found to be more phototoxic againstS. mutansbiofilm than TBO alone. [ABSTRACT FROM AUTHOR]

NiFe2O4/Carbon nanotube composite materials have been prepared using in-situ chemical precipitation of metal hydroxides in the presence of multi-walled carbon nanotubes (MWCNTs) and followed by hydrothermal processing. The obtained composite powders were characterized using XRD and TEM. The adsorption efficiency of NiFe2O4/carbon nanotubes for toluidine blue at various temperatures was examined. The amount adsorbed increased with the NiFe2O4/CNTs dosage. The linear correlation coefficients and standard deviations of Langmuir and Freundlich isotherms were determined. The results revealed that Langmuir isotherm fitted the experimental results well. Kinetic analyses were conducted using pseudo first-order, second-order and the intraparticle diffusion models. The regression results showed that the adsorption kinetics were more accurately represented by a pseudo second-order model. Changes in the free energy of adsorption (ΔG0), enthalpy (ΔH0) and entropy (ΔS0), as well as the activation energy (Ea) were determined. ΔH0and ΔS0were 15.99 kJ/mol and 98 J/mol K, respectively. The activation energy was 8.2 kJ/mol. ΔH0, ΔG0andEavalues reflected that the adsorption of toluidine blue onto NiFe2O4/CNTs was by physisorption. [ABSTRACT FROM AUTHOR]

This study investigates the effects of the Fe2+concentration and synthetic iron oxide catalysts ono-toluidine degradation using a fluidized-bed Fenton process. The mineralization ofo-toluidine in the synthetic catalyst system is also examined. The H3.5 and H7.3 Fe/SiO2and A7.8 and A12.5 Fe/SiO2catalysts were successfully synthesized by adding H2O2and injecting air process, respectively. The optimum initial ferrous ion concentration for degradation of 1 mMo-toluidine was 1 mM. Experimental results reveal that 1 mMo-toluidine can be 100% degraded at 60 and 120 min in the modified fluidized-bed Fenton process with A7.8 Fe/SiO2and the conventional fluidized-bed Fenton process with SiO2carrier, respectively, when the optimum conditions of 1 mM Fe2+and 17 mM H2O2at pH 3 were used. The A7.8 Fe/SiO2catalyst had a stronger oxidation ability than the H3.5 Fe/SiO2, H7.3 Fe/SiO2and A12.5 Fe/SiO2catalysts, and was attributed to the high iron content on the surface of the SiO2support. The Fenton and Fenton-like reactions occurred in the A7.8 Fe/SiO2catalyst system. Degradation ofo-toluidine in the Fenton-like process follows pseudo-first-order kinetics. The A7.8 Fe/SiO2catalyst efficiently enhancedo-toluidine oxidation under the pH range of 2–4. [ABSTRACT FROM PUBLISHER]

Fungal stains on slides dewaxed by domestic dishwashing detergent in hot water were compared to xylene and alcohol dewaxing. The fungal stains were Grocott's Methenamine Silver (GMS), Periodic Acid Schiff's following diastase (DPAS), and Sulphation Toluidine Blue (STB). For detergent dewaxing, a 2% concentration of domestic dishwashing detergent in distilled water heated to 90°C was used and paraffin sections were treated twice for 1 minute each. Hot detergent dewaxing is not only faster, cheaper, and safer than xylene dewaxing, but also equivalent or better than xylene for detecting fungi on DPAS, GMS and STB. [ABSTRACT FROM AUTHOR]

We explored the relationship between sperm chromatin integrity, hormone levels, seminal plasma total antioxidant capacity (TAC), and routine sperm parameters in men with male factor (MF, n = 81) and non-male factor (NMF, n = 52) infertility. Semen and blood were collected and examined from men undergoing evaluation for infertility in the Avicenna Infertility Clinic. We have examined each patient for serum hormones (LH, FSH, E2, DHEA), sperm chromatin damage, level of protamination and seminal plasma TAC. Levels of FSH, LH, sperm chromatin damage, and abnormal protamination were significantly higher in MF vs. NMF groups ( p < 0.001). Sperm chromatin damage was correlated with percentage of CMA3- positive sperm (r = 0.64, p < 0.001) and with sperm concentration (r = −0.36, p < 0.001), motility (r = −0.21, p < 0.05), and morphologically normal spermatozoa (r = −0.29, p < 0.001). Linear regression showed sperm chromatin damage was related to percentage of CMA3- positive sperm ( p < 0.001) in ungrouped patients. It was related to both percentage of CMA3- positive sperm and serum DHEA in the MF group ( p < 0.001 and p < 0.05, respectively). Sperm chromatin maturity assessed by CMA3 test was inversely related to sperm chromatin damage assessed by the toludine blue assay. Male factor infertility associated with sperm chromatin damage may be related to sperm protamination and to serum DHEA. [ABSTRACT FROM AUTHOR]

Mast cells play a significant role in inflammatory diseases such as asthma, inflammatory bowel disease, and autoimmune diseases. Inhibition of c-kit receptor tyrosine kinase, a growth factor receptor, significantly reduces mast cell numbers. The purpose of this study was to determine the effect of Compound X (a c-kit inhibitor) on mast cell numbers in rats. Connective tissue mast cells (CTMCs) and mucosal mast cells (MMCs) have differing histochemical characteristics which presents a challenge when staining for quantification by semi-automated image analysis. CTMCs are present in tissues such as tongue and skin and will stain readily in tissues fixed routinely. In contrast, MMCs, such as those present in the intestinal mucosa, are sensitive to fixation. Brief fixation in Carnoy's solution, although seldom used due to its composition (a mixture of ethanol, chloroform, and acetic acid), was employed to fix tissues for MMC staining, while tissues for CTMC demonstration were fixed in 10% neutral buffered formalin. An enzyme histochemistry method, napthol AS-D choloroacetate (specific esterase), was briefly considered for staining; however, granulocytes stained along with mast cells, requiring manual identification and exclusion, thereby rendering the method incompatible with automated means of quantification. Instead, staining was performed using two different toluidine blue methods which have proven conducive to semi-automated image analysis techniques. CTMCs were stained using Luna's toluidine blue, while MMCs were stained with Matsson's toluidine blue modification. In summary, the selected methods, based upon a conventional stain, were easy to do and successfully identified both populations of mast cells for quantification by image analysis. [ABSTRACT FROM AUTHOR]

Histochoice™® is a proprietary nontoxic, non-cross-linking fixative designed by the manufacturer to replace formaldehyde based fixation protocols. We compared Histochoice and formalin fixation for several cartilaginous tissues including, articular and growth plate cartilage, meniscus and intervertebral disc. The tissues were stained with general histology stains including toluidine blue for tissue proteoglycans, picrosirius red to evaluate collagenous organization, and hematoxylin and eosin to assess cell morphology. The chondroitin sulfate and heparin sulfate substituted proteoglycans aggrecan and perlecan were also immunolocalized in some of the tissues to provide a comparison. Histochoice did not fix deep into the tissue blocks resulting in focal loss of aggrecan and other matrix components from the more central regions of the blocks. This was evident in toluidine blue stained sections of immature tibial articular cartilage where loss of glycosaminoglycan was significant in Histochoice™® fixed tissues. Histochoice fixation worked well, however, in the aggrecan and perlecan immunohistology applications where its non-cross-linking traits were conducive to epitope retrieval and identification by primary antibodies to extracellular matrix components. [ABSTRACT FROM AUTHOR]

A spectrofluorometric method was developed for the determination of total serum protein by exploring toluidine blue (TB) as the fluorescence probe. The fluorescence intensity of TB at 648 nm was significantly quenched in the presence of sodium dodecylbenzene sulfonate (SDBS) by forming a dimer of the dye, which can afterwards reconvert to monomer when proteins were added accompanied by the recovery of the fluorescence. This might be attributed to the modulated transferring of the dimer-monomer equilibrium of TB in the anionic surfactant caused by the addition of protein. A linear calibration graph was obtained in the range of 0.5-50 mg/l BSA, with a detection limit of 0.15 mg/l and a RSD of 1.3% (n=11, 5.0 mg/l BSA). Total proteins in human serums were analyzed by using the present procedure and the results agreed well with those obtained by the Biuret method. [ABSTRACT FROM AUTHOR]

Photogalvanic effect was studied in photogalvanic cells containing toluidine blue as photosensitizer, ethylenediamine tetra acitic acid as reductant and sodium lauryl sulphate, cetyl pyridinium chloride, Tween-80 as surfactants in different systems. The photopotential and photocurrent generated were 743.0 mV and 70.0 µA in EDTA-TB-NaLS system, 762.0 mV and 35.0 µA in EDTA-TB-CPC system, and 430.0 mV and 50.0 µA in EDTA-TB-Tween-80 system, respectively. The effects of different parameters on electrical output of the cell were observed, and a mechanism has been proposed for the generation of photocurrent in photogalvanic cell. [ABSTRACT FROM AUTHOR]

An improved microscopic slide preparation for staining of tissue-isolated and in vitro amyloid fibrils is described in detail. Specimens were prepared by slow drying of an amyloid fibril suspension on aminoalkyl-silanised glass slides underneath a coverglass. The purpose of this method is to obtain microscopically visible fibers. Polarization microscopy was performed on unstained fibers in different embedding media, on fibers stained with Congo red, toluidine blue, 1,9-dimethylmethylene blue, anisotropic PAS reaction and Sudan black. Results were identical in 8 tissue-isolated amyloid fibril preparations. The refractive index of amyloid was about 1.5, as expected for protein fibrils. Congo red-stained fibrils showed yellow or blue–green polarization colors, while staining with toluidine blue or 1,9-dimethylmethylene blue induced green or orange polarization colors. The latter dyes stained the fibrils even in buffers with low pH or high ionic strength, pointing to strong acidic groups like sulphates in amyloid. The anisotropic PAS reaction for selective demonstration of periodate reactive carbohydrates in tissue-isolated amyloid fibrils was positive. Thus, linearly ordered acidic groups and carbohydrate residues could be detected for the first time in tissue-isolated amyloid fibrils using an improved technique for microscopic slide preparation on one side and topo-optical reactions on the other. [ABSTRACT FROM AUTHOR]

The study of undecalcified bone by histological methods is essential in the field of bone research. Culturing skeletal tissues such as neonatal murine calvaria provides a reliable bridge between assessment of bone formation in vitro and anabolic activity in vivo and contains most of the essential elements of bone for studying bone formation. Neonatal calvarial assay, supported by histological methods, is used to study the anabolic effects of a wide variety of factors and compounds on bone tissue. To optimize visualization and histomorphometric measurements using neonatal calvaria, we developed a method that provides high quality tissue sections suitable for routine and histochemical staining. Undecalcified neonatal mouse calvaria were processed and embedded using a low temperature methyl methacrylate procedure. Various staining methods were performed on deplastisized and floated sections to examine mineralization and to identify cells. The Von Kossa stain counterstained with a modified H & E yielded precise images of unmineralized bone including mineralization sites, and distinct osteoblasts and osteoclasts. Toluidine blue, Ladewig's trichrome, tartrate-resistant acid phosphatase, Goldner, H & E and Villanueva stains also were tested on the undecalcified neonatal calvaria sections. [ABSTRACT FROM AUTHOR]

Based on a detailed study of the interaction of toluidine blue (TB) with nucleic acids, a novel resonance light scattering (RLS) method was developed for the determination of nucleic acids. The RLS intensity of TB at 400 nm was greatly enhanced in the presence of nucleic acids at pH 3.17. Linear relationships between the enhanced RLS intensity and the concentration of nucleic acids were observed in the range of 0.0–2.7 µg cm-3 for calf thymus DNA (ctDNA), 0.0–2.2 µg cm-3 for fish sperm DNA (fsDNA), and 0.0–0.5 µg cm-3 for yeast RNA (yRNA), respectively. The limits of detection (3σ) were determined to be 24.0 ng cm-3 for ctDNA, 22.7 ng cm-3 for fsDNA and 5.1 ng cm-3 for yRNA. The binding constants were evaluated to be 1.1 × 104 dm3 mol-1 for ctDNA, 1.8 × 104 dm3 mol-1 for fsDNA, and 1.7 × 104 dm3 mol-1 for yRNA. The maximum binding numbers (n) were 2.7, 5.5, and 15.0 for ctDNA, fsDNA and yRNA, respectively. The proposed method was applied for determination of the nucleic acids in chicken blood. The analytical results obtained by the present method were in good with that from a UV method. [ABSTRACT FROM AUTHOR]

Covalent immobilization of heparin has been developed to reduce the amount of heparin administered systematically during long-term dialysis. Recently, it was doubted partially because of the complexion during immobilization process. In this study, we investigated a novel method for specific immobilization of heparin on polysulfone (PSF) via free electron laser (FEL) irradiation. Laser wavelengths of 6.18 or 6.31 μm, the typical absorption bands of carboxyl groups of heparin and aromatic rings in PSF, respectively, were chosen to irradiate the thin heparin membrane formed on PSF surfaces. The amount of heparin immobilized on PSF was measured by the toluidine blue method. The binding of heparin on PSF was analyzed by X-ray photoelectron spectroscopy (XPS). The immobilization of heparin resulted in a hydrophilic surface on which decreased platelet adhesion was observed. The efficiency differences, depending on laser wavelengths, were discussed from the point of view of structural and environmental differences of light-absorbing groups. [ABSTRACT FROM AUTHOR]

Background: The oral tetracyclines, especially minocycline hydrochloride, are often used as an effective treatment for perioral dermatitis; however, they are sometimes difficult to use, especially in children, because of the side effects. Objective:The effectiveness of β-lactam antibiotics was evaluated in three cases of perioral dermatitis. Methods: Three Japanese patients with perioral dermatitis were treated with cefcapene pivoxil hydrochloride hydrate per os 100-300 mg/day. They were one girl (aged 10 years) and two adult women (aged 32 and 37 years respectively). One of the adult patients had a past history of Meniere's disease and the other had had a side effect, vertigo, from minocycline hydrochloride treatment. The presence of fusobacteria before and after the treatment was examined using the tape-stripping toluidine blue method. Results:These patients showed the improvement in 1-2 weeks and were much improved or cured after 2-5 weeks. No side effects were found during the treatment. Fusobacteria were positive before treatment but became negative after the treatment in all of them. Conclusion: The β-lactam antibiotics might be a useful treatment for perioral dermatitis, especially in cases who cannot take tetracyclines. [ABSTRACT FROM AUTHOR]

Methemoglobinemia usually results from exposure to oxidizing substances such as nitrates or nitrites. Iron within hemoglobin is oxidized from the ferrous (Fe 2+ ) state to the ferric (Fe 3+ ) state, resulting in the inability to transport oxygen and carbon dioxide. Clinically, this condition causes functional cyanosis. As methemoglobin levels increase, patients show evidence of cellular hypoxia in all tissues. Death usually occurs when methemoglobin fractions approach 70% of total hemoglobin. We describe the case of a 35-year-old female patient with severe life-threatening isobutyl nitrite-induced methemoglobinemia of 75% of total hemoglobin. Toluidine-blue was administered as first-line antidotal therapy immediately, followed by hyperbaric oxygenation. The patient recovered uneventfully and could be discharged 3 days later. [ABSTRACT FROM AUTHOR]

The article presents questions and answers related to the field of biology. Toluidine blue or alcian blue being used to identify calcium phosphate deposits. Toluidine blue can be used to stain either calcified or decalcified bone sections to identify cells and other components of the tissue. One might be able to see calcium phosphate in skin stained with toluidine blue, but the overall coloration will be shades of blue and purple, masking calcium deposits. The rabbit is one of several species with water soluble mast cell granules. To preserve them, one must use a non aqueous fixative such as Carnoy or an alcoholic Bouin mixture.