Dark Blue Spots Research Articles

Hysteroscopic evaluation of chronic endometritis and its relation to unexplained infertility

Background Hysteroscopy is considered the gold standard for detecting diseases of the uterine cavity. It is simple, safe, and can be performed in an outpatient clinic. Although endometritis is not clinically clear, it may lead to infertility. Moreover, there is strong evidence that endometritis may lead to a premature spontaneous birth. Aim This study aimed to assess the diagnostic value of diagnostic hysteroscopy in patients with chronic endometritis. Patients and method The current study was performed on 50 women with unexplained infertility. All patients presented to the outpatient clinic of Tanta University Maternity Hospital in the follicular phase of menstruation. Results Hysteroscopic findings in the form of hyperemia, edema, micropolyps, and dark blue spot suggesting chronic endometritis were found in 31 cases. Histopathological examination confirms the diagnosis of chronic endometritis in 28 cases by detection of plasma cells in the endometrial biopsy. Microbiological examination showed a positive growth in 17 cases by detection of organisms in the endometrial biopsy; 1 of them showed positive chlamydial infection and 4 cases of mycoplasma in the multiplex polymerase chain reaction test. There was a significant association between hysteroscopic picture suggesting chronic endometritis and its confirmation by histopathological examination of endometrial biopsy. Conclusion Hysteroscopy is a convenient examination for detecting chronic endometritis. It could be applicable for investigating asymptomatic infertile patients. Antibacterial, antichlamydial, and antimycoplasma drugs have a good result in chronic endometritis. Endometrial dyeing with methylene blue at hysteroscopy improves the detection of chronic endometritis. More studies are recommended for endometrial evaluation by hysteroscopy.

Visual detection of microRNA146a by using RNA-functionalized gold nanoparticles.

Gold nanoparticles of different sizes have been synthesized and surface-functionalized with selected RNA probes in order to develop a rapid, low-cost and sensitive method for detection of microRNA146a (miR146a). The strategy is based on the change of colour that can be observed visually after aggregation of the RNA modified-gold nanoparticles (AuNPs) in presence of miR146a. Experimental conditions have been carefully selected in order to obtain a good sensitivity that allows to perform visual detection of microRNA at the nM level, achieving a detection limit of 5nM. Good repeatability and selectivity versus other sequences that only differ from miR146a in 3 bases was achieved. miR146a has been described as one of the main microRNA involved in the immune response of bovine mastitis, being expressed in tissue, blood and milk samples. The method was successfully applied to the detection of miR146a in raw cow milk samples. The present scheme constitutes a rapid and low-cost alternative to perform highly sensitive detection of microRNA without the need of instrumentation and amplification steps for the early detection of bovine mastitis in the agrofood industry. Graphical abstract Schematic representation of the assay based on aggregation of RNA-modified gold nanoparticles (blue) in presence of microRNA146a generating a dark blue spot onto a solid support, versus a pink spot observed in absence of miR146a due to dispersed gold nanoparticles (red).

ABSENCE OR PRESENCE OF TAPETUM LUCIDUM: MACRO AND MICROSCOPIC INVESTIGATIONS IN DONKEY (EQUUS ASINUS), CAT (FELIS DOMESTICA) AND ONE-HUMPED CAMEL (CAMELUS DROMEDARIUS)

For explanation eye-shine phenomenon, we used both eyes of five healthy adult donkeys (Equus asinus), cats (Felis domestica) and one-humped camels (Camelus dromedarius). The eyes of a live animal of the three species were photographed under existing light and with a flash. The donkey's pupils appeared black centrally situated and horizontal in direction at daylight. Under flash, the light condensed centrally and the color changed to green or blue. The cat's pupils were oval vertically slit-like and colored yellow to red orange under flash. The camel's pupils characterized by presence of a small centrally spot of light with flash. Tapetum of the donkey was horizontal triangular in shape under the weak light colored indigo with dark blue spots changed into semicircular appearance under strong light. In the cat, Tapetum appeared semicircular in outline with yellow color under weak and strong light. The fundus of camel appeared divided into dark proximal half and lighter distal one under the weak light. Under flash, the two halves appeared transparent white. Microscopically, Tapetum lucidum of the donkey was fibrous in its texture while in cat, it was cellular. In camel, there was a brush's membrane and no Tapetum lucidum. In donkey, the thickness of the tapetal tissue and the degree of pigmentation in the retinal epithelium differed according to the region of Tapetum. The thick tapetal tissue and the unpigmented retinal epithelium combination created the greater reflectance of light. So the absence or presence of Tapetum, the tapetal tissue thickness, the degree of pigmentation in the retinal epithelium and the degree of illumination controlled the eye-shine phenomenon.

Development and validation of a high-performance thin-layer chromatographic—densitometric method for the quantification of guggulsterones E and Z in a polyherbal formulation containing Commiphora Mukul

The present article focuses on the quantitative analysis of a polyherbal formulation containing arjuna, guggul, and pushkarmoola in a combined dosage form which is used for its cardioprotective, anti-anginal, and hypolipidemic activities. Quantification of guggulsterones E and Z was done by developing and validating a high-performance thin-layer chromatography method after extraction by methanol. Densitometric evaluation was carried out on precoated silica gel G60 F254 high-performance thin-layer chromatography (HPTLC) plates using toluene—ethyl acetate (8:2, v/v) as the mobile phase. Based on the HPTLC results, the methanol extract of guggulsterone was chosen as it showed a better separation of spots as compared to another extract. After postderivatization of plates with anisaldehyde—sulfuric acid (10%) reagent, the methanol extracts gave different color spots (the arjuna extract gave a blue spot, the guggul extract gave orange and brown spot, and pushkarmoola gave dark blue spot). Scanning and densitome...

IDENTIFIKASI SENYAWA GOLONGAN FENOL DARI EKSTRAK ETANOL DAUN SIRIH HIJAU (PIPER BETLE LINN.) DENGAN METODE KLT-SPEKTROFOTODENSITOMETRI

Phenol compound in Piper betle leaves has several pharmacology activities such as antibacteria, antifungi and antioxidant. The pharmacology activities of a herbal drug are influenced by the phytochemistry content, so in order to do a quality determination that provides phytochemistry profile and consistent pharmacology activities, a standardization is required. Fingerprint is the main standard to perform quality control for herbal drug. TLC-spectrophotodensitometry was used as the method in order to provide fingerprint profile of phenol compound. In this experiment, Piper betle leaves samples were extracted by reflux method using ethanol 96% as the solvent. Identification of phenol compound was done using TLC-spectrophotodensitometry with Silica gel 60 F254 as the stationary phase, toluena: ethyl acetate (93:7 v/v) as the mobile phase, FeCl3 and Folin-Ciocalteau as the reagent. The fingerprint profile of phenol compund was shown in Rf value 0,19; 0,42; and 0,62. Positive results of phenol compound are black spot on FeCl3 colour test and dark blue spot on Folin-Ciocalteau colour test. Maximum wavelength of phenol compound was 283 nm.

Endoscopy of Aberrant Crypt Foci: The Expert Approach

Aberrant crypt foci (ACF) were found to be possible precursors of adenoma and cancer, as well as useful surrogate lesions for chemoprevention of colorectal cancer. The aberrant crypts are stained more darkly with methylene blue and have a thicker epithelial lining and a larger pericryptal zone than normal crypts. Recently, advanced endoscopic imaging techniques have been found to be more helpful to improve detection and characterization of ACF in the colon. For inspection of ACF, optimal bowel preparation and a high-resolution endoscope equipped with a magnifying function is required. After spraying plenty of methylene blue, the mucosal surface should be washed with plenty of water to remove the sticky mucus. Once the dark blue spot is recognized with low magnification, zooming up to the maximum point should be performed. This article is part of an expert video encyclopedia.

Cost effective quantification of camptothecin and a comparative study of its content in Nothapodytes foetida and Ophiorrhiza mungos sourced from selected geographical locations

Abstract High-performance liquid chromatography is theprevalent method used to estimate Camptothecin quantita-tively. In the present study thin layer chromatography andUV spectrophotometric method was established as a sensi-tive, economical and accurate method of detection and esti-mation of Camptothecin. The method was developed onTLCaluminumplates(60F 254 fromE-MerckLtd.)precoatedwith silica gel using solvent system ethyl acetate: methanol(7.5:2.5, v/v) which gives a dark blue spot of Camptothecinat 254 nm (Rf value 0.46 cm). Spectrometric analysis ofCamptothecin was carried out at the absorbance of 254 nm.Leaves and stems of Nothapodytes foetida and Ophiorrhizamungos were extracted, subjected to TLC and UV spectro-photometric analysis and maximum absorption of CPT inethyl acetate was observed at 254 nm. The quantification bythis method was comparable to data obtained by HPLC. Theoptimized method was simple, repeatable, precise and costeffective, useful when there is large number of crude sam-ples.Inthepresentstudythismethodwasusedtoscreenhighyielding lines of CPT producing plants.Keywords Camptothecin .Nothapodytesfoetida .Ophiorrhizamungos .ThinlayerchromatographyAbbreviationsCPT CamptothecinTLC Thin layer chromatographyHPLC High performance liquid chromatographyIntroductionCamptothecin (CPT) is a cytotoxic quinoline alkaloid(Fig. 1), first isolated by Wall and Wani (1996) fromCamptotheca acuminate belonging to family Nyssaceae.Camptothecin and its derivatives was found to inhibit DNAtopoisomerase activity and at present the semi syntheticwater soluble CPT analogs known as topotecan andirinotecan are used as anti-cancer agents (Aimi et al. 1989;Arisawa et al. 1981; Hsiang et al. 1985; Yasuyo Yamazakiet al. 2003). Among the various plant sources for anti-tumorcompounds like Vinca alkaloid Vincristine (Noble 1990),Etoposide derived from Podophyllum peltatum (Thurstonet al. 1989), Flavopiridol a synthetic flavone derived fromthe plant alkaloid rohitukine which was isolated from theleaves and stems of Amoora rohituka and later fromDysoxylum binectariferum (Kelland 2000). A number ofadditional plant derived agents are currently under investi-gation. Homoharringtonine is an alkaloid isolated from theChinese tree, Cephalotaxus harringtonia has shown efficien-cyagainstvariousleukemia,4-Ipomoeaonolisapneumotoxicfuran derivative isolated from sweet potato Ipomoea batatas(Rehm and Devor 1993). It is believed that CPT is thethird most important alkaloid sought after by pharma-ceutical companies around the world which was initiallyisolated from the bark and wood of Camptotheca acu-minate (Nyssaceae) and act by inhibiting topoisomerase

Microstructural changes of starch in cooked wheat grains as affected by cooking temperatures and times

The changes of starch microstructure in soft and hard wheat grains after cooking in a pressure cooker were investigated using a scanning electron microscope (SEM) and a light microscope (LM) in conjunction with image analysis. The conditions studied included variation of the cooking times (20–120 min) at the constant temperature (120 °C) and with variable temperatures (110–140 °C) for constant time (40 min). SEM images showed that steam induced changes in the intact starch granules of cooked wheat grains, with the microstructure becoming more in mud-like structure, with increased cooking time or temperature. The swelling and gelatinization of starch granules could contribute to this change. Suspensions of cooked wheat flour were selectively stained with iodine, for observation using the LM. The image analysis software was used for quantitative analysis of the images captured from the LM, providing both the number and area of the starch granules (determined from the dark-blue spots) per fixed image size. With increased cooking time or temperature, the number of starch granules decreased and the area increased as a result from the swelling and melting of starch granules. This study demonstrated the use of SEM and LM for the investigation of starch granules in cooked wheat grains without prior starch isolation.

TLC identification and quantification of coenzyme Q10-β-cyclodextrin complex

TLC is often set aside as a result of unjustified simplification of analytical knowledge and, at the same time, over-emphasis of the use of metrological solutions. Some analysts assume that techniques with smaller measurement uncertainty automatically improve the reliability of their analytical results. This is not true. The reliability of some TLC techniques is demonstrated by analysis of an inclusion complex of coenzyme Q10 (CoQ10) with β-cyclodextrin, which was prepared because it is a water-soluble food additive. The products obtained were analyzed by one-dimensional, two-dimensional, and multi-dimensional TLC (according to Ebel). These inexpensive, rapid, and very informative methods were used in-line during the preparation steps. Separation was performed in two steps, with dioxane-water, 50 + 50 (v/v), as the first mobile phase and chloro-form-methanol, 55 + 45 (v/v), as the second. The development distance was 7.5 cm. The separated spots were visualized in two steps, first with 5% phosphomolybdic acid in EtOH and drying at 110°C, then with 50% H2SO4 and heating at 120°C for 5 min. After the first treatment blue spots on yellow background were observed for CoQ10. After spraying with H2SO4 dark blue spots of CoQ10 were observed on a light blue background. Images were acquired with a scanner and quantification was performed by use of TLC software. The synthetic yield was calculated from the concentrations determined. The results obtained were later confirmed by use of IR, NMR, and HPLC-MS.

B19 parvovirus induced fetal hydrops: rapid and simple diagnosis by detection of B19 antigens in amniotic fluids

In our study we describe the direct detection of parvovirus B19 capsid antigens in amniotic fluid samples for the rapid and simple prenatal diagnosis of B19 induced fetal hydrops. The assay was performed on amniotic fluid specimens from fetal hydrops dotted on nylon membranes. The two capsid antigens, VP1 and VP2, which represent four per cent and 96 per cent of the capsid, respectively, were detected using a pool of monoclonal antibodies directed against these two proteins and the complex was visualized by immunoperoxidase staining. The assay could be performed in about four hours and positive results were revealed at the end of the reaction as dark blue spots on the nylon membrane. We analysed 26 amniotic fluid samples from 26 selected cases of non-immune hydrops for the presence of B19 antigens. Out of these 26 samples, 13 had previously proved positive for B19 DNA, detected by dot blot hybridization and/or in situ hybridization and/or nested PCR, and 13 had proved negative. The results obtained with our assay were compared with results obtained for the presence of B19 DNA and a close agreement was found. The method is simple and rapid to perform, does not require costly instruments, and all the reagents used in the assay are commercially available. The assay described can thus be useful for a prompt counselling and management of B19 fetal infection. © 1998 John Wiley & Sons, Ltd.

B19 parvovirus induced fetal hydrops: rapid and simple diagnosis by detection of B19 antigens in amniotic fluids

In our study we describe the direct detection of parvovirus B19 capsid antigens in amniotic fluid samples for the rapid and simple prenatal diagnosis of B19 induced fetal hydrops. The assay was performed on amniotic fluid specimens from fetal hydrops dotted on nylon membranes. The two capsid antigens, VP1 and VP2, which represent four per cent and 96 per cent of the capsid, respectively, were detected using a pool of monoclonal antibodies directed against these two proteins and the complex was visualized by immunoperoxidase staining. The assay could be performed in about four hours and positive results were revealed at the end of the reaction as dark blue spots on the nylon membrane. We analysed 26 amniotic fluid samples from 26 selected cases of non-immune hydrops for the presence of B19 antigens. Out of these 26 samples, 13 had previously proved positive for B19 DNA, detected by dot blot hybridization and/or in situ hybridization and/or nested PCR, and 13 had proved negative. The results obtained with our assay were compared with results obtained for the presence of B19 DNA and a close agreement was found. The method is simple and rapid to perform, does not require costly instruments, and all the reagents used in the assay are commercially available. The assay described can thus be useful for a prompt counselling and management of B19 fetal infection.

The determination of the antiperspirant activity of aluminium-chlorhydrate by digital image-analysis

Synopsis The method of Minor to visualize by iodine-starch-reaction the activity of sweat glands was used in combination with image-analysis to quantify sweat-secretion. Aqueous aluminium chlorohydrate solutions (ACH) (1, 2, 4, 8, 16%) were applied twice daily for 6 days to defined areas of the lower volar forearms. After 3 and 6 days of treatment perspiration was induced by thermal stress. The dark blue spots on top of active glands which are formed by iodine-starch-complexes were counted and their projection area was measured by an image analysing system. Increasing concentrations of ACH lower the number of active sweat glands, while the size of the dark spots is not dependent on the concentration of the active ingredient.

Dot immunoperoxidase assay using monoclonal antibody for direct detection of cytomegalovirus in urine samples.

A rapid, simple dot immunoperoxidase assay for the direct detection of cytomegalovirus in clinical urine samples was developed. The assay was performed on nitrocellulose paper dotted with urine pellets free of cellular debris. Cytomegalovirus was detected with a monoclonal antibody to the capsid antigen, and the complex was visualized by immunoperoxidase staining. Positive reactions appeared as well-defined dark blue spots. Of the 87 urine samples examined, 10 proved positive in the dot immunoperoxidase assay, and 77 proved negative. The results agreed completely with the detection of cytomegalovirus-induced antigens in cell cultures inoculated with clinical specimens.

A dot immunobinding assay (DIA) on nitrocellulose membrane for the serological detection of antibodies to Entamoeba histolytica

A rapid, cheap, simple and specific serological test of adequate sensitivity for detecting IgG antibodies against Entamoeba histolytica antigen is described. Axenically cultured amoebic antigen was used to precoat the nitrocellulose membrane. The strips were incubated with test samples and later with horseradish peroxidase (HRPO) labelled protein-A conjugate. A dark blue spot was obtained by treatment with peroxidase substrate, 4-chloro-l-naphthol, in positive samples. Serum samples from 32 healthy controls, 45 patients with acute amoebic liver abscess and 10 asymptomatic E. histolytica cyst passers were tested. This test was positive in 93% of cases of amoebic liver abscess, 3% of healthy controls and none of the cyst passers; its sensitivity (97%) and specificity (93%) were as good as that of the ELISA test and, because it is simple, quick and cheap it is recommended as a serological test of choice for the diagnosis of invasive amoebiasis.

Study of accompanying impurities in methyltestqsterone by a combination of physicochemical methods

In the present work, we report the results of studies on the structure and content of possible accompanying impurities in methyltestosterone (I). To solve the problem, we first revealed the presence of foreign steroids by the method of thin layer chromatography (TLC) and by the method of high-performance liquid chromatography (HPLC). To study the structure of the impurities, we used the methods of UV spectroscopy and chromato-mass spectrometry (CMS). The quantitative content was determined by the HPLC method. EXPERIMENTAL The analysis was carried out on serially produced samples of methyltestosterone and standard samples from the World Health Organization (WHO), European Pharmacopoeia, British Pharmacopoeia, and the National Formulary of the U.S.A. For a preliminary estimation of the quality of the compounds studied, we used the TLC method. The studies were carried out on 150  150 mm ready-for-use Silufol UV-254 plates (CSSR) in hexane-acetone (70:30) system of solvents [2]. The adsorption zones of the compounds on the chromatogram were detected by examination Under an ultrachemiscope with a light filter at a wavelength of 254 nm and using a 10% solution of phosphomolybdic acid in acetone as developer. The compounds on the chromatogram were developed in the form of darkblue spots on a yellow background by heating to 80~ When 200 ~g of the sample studied were deposited on the starting line of the chromatographic plate under the above conditions, we observed three additional zones (Fig. i). In several samples one hardly visible spot present near the solvent front was noted. In view of the small amount of the compound, its structure could not be established. We have already shown that the final compound does not contain either the starting material or the intermediate products of its synthesis [I]. The same samples of methyltestosterone were then examined by the HPLC method on a chromatograph from the firm Waters Associates, model 240 with a UV detector at 254 nm. In the investigation, a 250  0.46 mm column was used, filled with sorbent Zorbax CN, with a particle size of 5-7 ~m. For the mobile phase, a heptane-chloroform mixture as used with an experimentally selected ratio of 91:9, by volume. The flow rate of the mobile phase was 2.5 ml/min, the velocity of movement of the diagram tape was 0.25 cm/min, and the time of analysis 20 min. The temperature of the column was 20~ As the result of chromatography, in some samples, besides the three above impurities, another impurity was found, which is eluted on the chromatogram after the principal substance (Fig, 2). To clarify the possible nature of the impurities by the TLC and HPLC methods, we isolated four fractions under the above conditions. These fractions were concentrated in a nitrogen current at 30~ and analyzed by the CM$ and UV spectroscopy methods.

A ‘dot-immunobinding assay’ on nitrocellulose membrane for the determination of the immunoglobulin class of mouse monoclonal antibodies

A rapid and convenient procedure for the determination of the immunoglobulin class of mouse monoclonal antibodies is described. Hybridoma supernatants or other dilute solutions of monoclonal antibodies are spotted onto nitrocellulose membrane strips, and each strip is incubated with a different rabbit antiserum specific for a mouse Ig class. The strips are then incubated with protein A conjugated to peroxidase, and finally with the peroxidase substrate 4-chloro-1-naphthol. A positive reaction produces a dark blue spot. The procedure is sensitive, economical, fast, and safe. Many monoclonal antibodies can be tested at one time, and most of the reagents can be re-used. The procedure has been applied to a new series of monoclonal antibodies recognizing plasma membrane antigens of rat thymocytes.

Improved Histology for the Chick-Quail Chimera

A simple modification of nuclear staining after acid hydrolysis has been made which provides easy identification of quail nuclear markings in a chick-quail chimera. This method also improves the histologic detail normally seen with hematoxylin and eosin when compared to the more commonly used Feulgen reaction. Embryonic tissues can be fixed in Zenker's or Helly's solution and the sections obtained are hydrolyzed in acid (3.5 N HCl at 37 C for 40-50 min). After acid hydrolysis the sections are stained with hematoxylin and eosin rather than Schiff reagent and fast green. The interphase nuclei of chick cells show homogeneous or mottled purplish blue staining, while quail nuclei contain a dark blue spot. This staining corresponds to the reddish purple staining of the quail's heterochromatin seen adjacent to the nucleolus in the standard Feulgen stain. This new technique facilitates identification of quail cell types in the chick host and provides superior histology of the chick tissues by demonstrating cytoplasmic detail.

Direct Fluorescent Detection of Organothiophosphorus Pesticides and Some of Their Sulfur-containing Breakdown Products After Thin Layer Chromatography

Abstract A rapid, simple, convenient, and widely applicable method for the direct fluorescent detection of organothiophosphorus pesticides and some postulated breakdown products of these compounds is presented. The coinpounds were spotted on thin layer chromatographic sheets, developed in ethyl acetate :nhexane, and made visible by exposure to bromine vapor followed by spraying with ferric chloride and 2-(o-hydroxy phenyl) benzoxazole. Of the 47 compounds tested, 32 compounds produced fluorescent blue spots vinder longwave ultraviolet light; these consisted of 25 organothiophosphorus pesticides, 5 sulfur-containing breakdown products, and phosphoric and hypophosphorous acids. A superimposed Congo red spray destroyed the fluorescence and resulted in dark blue spots against a red backgrovind. The sensitivity of this method is in the range of 0.2 to 5.0 μg, depending on the specific compound.

On the colouring matter of the black bronchial glands and of the black spots of the lungs

It is well known that in men the bronchial glands that are situated near the bifurcation of the trachea are of a very dark colour; that in infancy the lungs in general have little or no such colour; that at the age of eighteen or twenty they have a mottled or marbled appearance, from black or dark blue spots, lines and points disseminated through them immediately under the membranous covering, and that this appearance becomes darker and darker as persons advance in age; and that the same black spots are also observable throughout the whole interior substance of the lungs when cut. It has been conjectured by some persons that this appearance is owing to sooty matter taken in with the air during respiration, and accumulated in proportion to the duration of life; but in reply to this supposition it has been objected by others, that the same degree of blackness is not observable in brute animals, and that the theory is not supported by any accurate observations of the proportional want of intensity in those persons who might be supposed to have lived less exposed to soot by permanent residence at a distance from any large town, and from any other considerable consumption of fuel.