The use of harlequin staining to assess radiation effects on the in vitro cell cycle kinetics of human lymphcoytes.
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Azure Stains↗
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Amplification of Pneumocystis carinii DNA on specimens scraped from slides.
A method for the detection of Pneumocystis carinii by polymerase chain reaction using specimens obtained by scraping bronchoalveolar lavage or tissue impression smears is described. The smears were scraped into water and then absorbed onto a glass-fiber filter. After fixing with methanol, the specimen on the filter was digested with proteinase K. The digestion mixture was then clarified, and a portion of the clarified supernatant was used as a template for the amplification of a portion of the mitochondrial rRNA gene of P. carinii. Using this method of sample preparation, we were able to amplify P. carinii DNA from both unstained and Giemsa stained smears.
Azure Stains↗
Viability and acrosome staining of stallion spermatozoa by Chicago sky blue and Giemsa.
A simple trypan blue-neutral red-Giemsa staining procedure for simultaneous evaluation of acrosome, sperm head, and tail membrane integrity and morphology has been used to evaluate equine spermatozoa. Some special characteristics and problems have arisen in evaluating stallion semen. One problem was the differentiation of intact vs. damaged sperm tails primarily in frozen and thawed samples. After freezing and thawing, a high percentage of spermatozoa with an unstained head and stained tail were observed. These cells are considered immotile. Therefore, unambiguous differentiation of intact vs. damaged sperm tail membrane is very important for evaluating semen quality. The aim of our study was to develop a method especially for stallion sperm to distinguish more accurately the different cell types. We compared Chicago sky blue 6B (CSB) to trypan blue (TB) for viability staining. CSB/Giemsa staining showed good repeatability and agreement with TB/Giemsa measurements. For densitometry analysis, individual digital images were taken from smears stained by CSB/Giemsa and by TB/Giemsa. A red-green-blue (RGB) histogram for each area of spermatozoa was drawn. Differences of means of RGB values of live vs. dead tails and separate live vs. dead heads from each photo were used to compare the two staining procedures. CSB produced similar live/dead sperm head differentiation and better tail differentiation. TB can be replaced by CSB and this results in more reliable evaluation. After staining with 0.16% CSB and 4 min fixation, 2-4 h Giemsa staining at 25-40 degrees C is recommended for stallion semen.
Acrosome↗
A method for analysis of cell morphology, banded karyotype, and immunoperoxidase identification of lymphocyte subset on the same cell.
The authors describe a method that allows three staining procedures to be applied to the same mitotic cell, namely Giemsa staining for morphology, immunoperoxidase staining for surface markers, and banding of the chromosomes for karyotype. After culturing of the blood lymphocytes, colchicine was added to arrest the cells in mitosis. After mild hypotonic treatment, the cells were cytocentrifuged to glass slides. The mitotic cells of different lymphatic cell subsets were identified with monoclonal anti-sera against T-helper-, T-suppressor-, and B-cells using the immunoperoxidase technic. The cells were counterstained with Giemsa for morphologic identification. After removal of Giemsa stain and peroxidase, the chromosomes were G-banded.
Azure Stains↗
Diagnosis of cerebral toxoplasmosis using fluorescein-labeled antitoxoplasma monoclonal antibodies.
The direct immunofluorescence technique using monoclonal antitoxoplasma antibodies was used to detect Toxoplasma organisms in 13 brain touch preparations and six smears of cerebrospinal fluid. For comparison, Giemsa-stained smears and histologic sections of the same cases were also studied. Positive results were obtained from two cases of acquired immunodeficiency syndrome (AIDS) with all three methods. Toxoplasma cysts, pseudocysts, and tachyzoites were easily identified by the immunofluorescence technique, and no false-positive results were encountered in nine cases of AIDS with different kinds of encephalitides. It was difficult to identify tachyzoites in histologic sections and time-consuming to search for pseudocysts and cysts. The parasite was even more difficult to recognize in Giemsa-stained brain touch preparations due to the easy distortion of the organisms. In the present study, immunofluorescence technique was more sensitive than hematoxylin--eosin or Giemsa staining in terms of the number and range of forms demonstrated in tissue. Therefore, immunofluorescence technique using monoclonal antibodies appears to be the preferred method for a rapid diagnosis of cerebral toxoplasmosis as compared to Giemsa staining and histology.
Antibodies, Monoclonal↗
[Detection of Helicobacter pylori in gastric biopsy and resection specimens].
AIM: To compare the sensitivity of detecting H. pylori in gastric biopsy and resection specimens using modified Giemsa stain and immunohistochemistry, using a commercially available anti-H. pylori antibody (Dako, Denmark). METHODS: Gastric antral biopsy specimens showing chronic gastritis (28 cases) together with tissue blocks from gastrectomy specimens for duodenal ulcer (2 cases) were stained with modified Giemsa and immunoenzymatic alkaline phosphatase - anti-alkaline phosphatase (APAAP) method, and were carefully examined for the presence of H. pylori. RESULTS: Using a modified Giemsa stain, the spiral shaped bacteria of H. pylori stained blue, were attached to the brush border of the gastric foveolar epithelial cells. However, the specificity of modified Giemsa stain depended on the morphological appearance of H. pylori. The specificity of immunostaining permitted detection of low numbers or even single organisms. In all cases bacteria were more prominent and easier to detect in immunostained preparations. H. pylori was identified in 22 (73.3%) of 30 sections stained with modified Giemsa stain, but it could be identified with greater frequency in sections stained with APAAP, in 27 (90%) of 30 sections. CONCLUSION: Immunohistochemical identification of H. pylori was better than Giemsa stain for detecting that organism.
Adult↗
Staining of different tissues in thick epoxy resin-impregnated sections of human fetuses.
Sections of undecalcified human fetuses, fixed in formaldehyde, embedded in the epoxy resin Biodur E 12 and cut on a diamond-wire saw were stained according to a slight modification of the method described by Laczkó and Lévai. The sections were immersed in a methylene blue/azure II solution at 90 C for at least 3 min and counterstained with a basic fuchsin solution at the same temperature. Differential staining was as follows: bone stained pinkish; cartilage, violet; collagen fibers, blue-violet; elastic fibers, red and muscle fibers, green-blue. Most other tissues were stained blue-violet against the transparent background of the embedding epoxy resin. Thanks to the distinct and differential staining of each tissue, contrast is sufficient for black and white as well as for color photography.
Azure Stains↗
GIEMSA STAIN FOR THE DIAGNOSIS OF BOVINE BABESIOSIS. II. CHANGES IN ERYTHROCYTES INFECTED WITH BABESIA BIGEMINA AND B. ARGENTINA.
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Animals↗
Topographic examination of sister chromatid differential staining by Nomarski interference microscopy and scanning electron microscopy.
BrdU-substituted Chinese hamster chromosomes were treated with a hog Na2HPO4 solution and stained with Giemsa to produce sister chromatid differential staining (SCD). The process of SCD was examined with the Nomarski differential interference microscope and the scanning electron microscope. After the Na2HPO4 treatment alone, unifilarly BrdU-substituted (TB) chromatids appeared somewhat more severely collapsed than the bifilarly substituted (BB) chromatids. Subsequent Giemsa staining, however, brought about pronounced piling up of the Giemsa dye on the TB-chromatids but not on the BB-ones, causing highly distinct differential Giemsa staining as well as a marked differentiation in surface topography between the sister chromatids. Removal of the Giemsa dye from the differentially Giemsa stained chromosomes resulted in a disappearance of such a pronounced topographic differentiation.
Animals↗
A simple procedure for obtaining autoradiographs of G-banded chromosomes.
Chromosomal preparations from cells flash-labeled with 3H-dT were Giemsa-banded following trypsin digestion, allowed to air-dry, and then were coated with a layer of 1% Formvar. The slides were subsequently coated with radiographic emulsion (NTB2) and processed for autoradiography. The resulting chromosomes had distinct G-banding and radiographic labeling patterns. Chemographic grain formation in the emulsion, normally caused by Giemsa stain, was prevented by the film of Formvar, allowing a very low background grain level.
Animals↗
Selective staining of the same set of nucleolar phosphoproteins by silver and Giemsa. A combined biochemical and cytochemical study on staining of NORs.
Gel electrophoresis of nucleolar isolates from Zajdela ascites hepatoma cells followed by various staining procedures revealed a common set of bands that stained selectively with silver and Giemsa. The gel bands, corresponding to molecular weights of 104, 78, 37, and 29 kilodaltons (kd), appeared to contain phosphoproteins that were at least partly associated with oligo-deoxyribonucleotides. Enzyme digestion studies showed that the Giemsastainability was due to the phosphorylated state of the proteins. The positive selective silver-staining reaction in gels could be most likely attributed to the high content of carboxyl groups present in these phosphoproteins. The significance of these findings in relation to cytological results produced by selective silver staining of nucleolus organizing regions (NORs) and by Giemsa N-banding is discussed.
Animals↗
A sequential staining technique for the chromosomal analysis of the interspecific mouse/hamster and mouse/human somatic cell hybrids.
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Animals↗
Comparison of two staining and evaluation methods used for computerized human sperm morphology evaluations.
The purpose of the study was to analyse the agreement between computer analysed (Hamilton Thorne, IVOS Dimensions Version 3) normal sperm morphology and values obtained from 97 slides stained according to the Papanicolaou and Diff-Quik method. Liquefied semen samples were washed once by centrifugation and air dried smears on slides were made, which were stained according to the Papanicolaou and Diff-Quik method and analysed by computer. The paired t-test was used to assess whether any bias existed between the two methods. The limits of agreement were calculated using the Bland and Altman approach and a modification of this approach (mean-dependent limits). A significant bias of 1.6% was obtained in favour of higher normal sperm morphology percentages when using the Diff-Quik method. The standard limits of agreement were -13.4% to 16.6%, whereas the mean-dependent limits of agreement were 1.6% [5.8 + 0.6 (mean percentage normal morphology)]. Statistically, the Diff-Quik and Papanicolaou staining methods produce different normal sperm morphology profiles. These inherent differences may, therefore, require the establishment of new normal sperm morphology thresholds for male fertility, based on clinical data, when using the Diff-Quik staining method in conjunction with computerized analysis.
Azure Stains↗
Visualization of surface immunoglobulin of human B lymphocytes using microsophere-immunoglobulin conjugate in Giemsa-stained preparations.
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Antibodies, Anti-Idiotypic↗
Differential Giemsa staining of sister chromatids after extraction with acids.
Chromosomes of Chinese hamster strain cells were air-dried on slides after BrdU substitution for two or three rounds of replication. The preparations were treated with 20% PCA at 55 degrees C for 20-30 min, or 5N HCl at 55 degrees C for 15-20 min. After staining with Giemsa, unifilarly BrdU-substituted chromatids stained faintly and bifilarly substituted chromatids stained darkly. Such a pattern of sister chromatid differential staining was confirmed by the examination of metaphase cells grown with BrdU for three rounds of replication.
Azure Stains↗
Comparison of four techniques for the routine diagnosis of Trichomonas vaginalis infection.
Specimens from 495 patients attending Johannesburg hospitals and family planning clinics were examined for Trichomonas vaginalis by microscopy of Giemsa (GS), Papanicolaou (PAP), and acridine-orange (AO) stained smears, and by culture in Feinberg-Whittington medium. Culture, Pap and GS stained smears from vaginal swabs yielded fewer positives than AO stained smears. Although Pap-stained cytological smears gave the highest number of positives, in 30% of these cases the presence of T.vaginalis could not be confirmed by examination of vaginal swabs. Of the positive AO-stained smears, 93% were also positive by at least one other technique.
Acridines↗
Banding analysis of the somatic chromosomes of the domestic dog (Canis familiaris).
The chromosomes of the domestic dog (Beagle) were investigated by several different staining techniques. G-banding, Q-banding, and the bis-benzimidazol derivative Hoechst 33258, make possible the identification of all 39 chromosome pairs. Constitutive heterochromatin (C-bands) was present on a few chromosomes as distinctive, large stained areas; on the other autosomes there was little or no heterochromatin detectable.
Animals↗
Leishman-Giemsa cocktail: an effective Romanowsky stain for air-dried cytologic smears.
OBJECTIVE: To develop an economical, quick, readily available and simple alternative to May-Grünwald-Giemsa (MGG) stain and so to explore the combination of Leishman and Giemsa stain (LG cocktail). STUDY DESIGN: One wet-fixed and 2 air-dried smears were prepared from 720 cases during the period January 2003-November 2004. The LG cocktail and MGG stain were used on the air-dried smears and Papanicolaou stain in wet-fixed smears. Diagnoses of the cases were made using the LG cocktail-stained smears, and then its diagnostic efficacy was cross-checked with the MGG- and Papanicolaou-stained smears by the same cytopathologist. A comparative study of the LG cocktail and MGG-stained smears was done. RESULTS: The results achieved with the LG cocktail-stained smears were comparable to or even better than those with the MGG-stained smears, with excellent diagnostic efficacy. CONCLUSION: As compared to MGG stain, the 1-step LG cocktail is cheaper, easier to standardize and quicker.
Adenocarcinoma↗