Reliable Staining Research Articles

Toluidine Blue-Basic Fuchsin Stain for Glycolmethacrylate Embedded Tissue

AbstractThis report presents a rapid, simple, and reliable redlblue staining method for glycolmethacrylate embedded tissue sections using a combination of toluidine blue and basic fuchsin. The end product is comparable that of the hematoxylin and eosin stain, and the technique works well with most ofthe plastic embedding media in routine use. (The J Histotechnol 16:139, 1993)

Production and characterization of a monoclonal antibody recognizing a cytoplasmic antigen of equine mononuclear phagocytes

An IgG1 mouse monoclonal antibody, designated 1.646, is described which recognizes a cytoplasmic antigen of equine mononuclear phagocytes. Indirect fluorescent antibody staining of peripheral blood leukocytes reveals a granular cytoplasmic staining, predominantly in adherent blood mononuclear cells. Indirect fluorescent antibody staining is positive for alveolar and peritoneal macrophages. In some horses, a few neutrophils are also stained. In equine tissue samples stained by immunohistochemistry, the distribution of positive cells is consistent with the distribution of tissue macrophages. The most intense and reliable staining occurs with splenic and lymph node macrophages. Hepatic Kupffer cells also stain with antibody 1.646, although the intensity of that staining is somewhat variable between horses. A granular pattern of staining typical of lipofuscin deposition is also seen in liver sections. There is also pale staining of some biliary and renal tubular epithelium. Equine erythrocytes, platelets and lymphocytes are not recognized by this antibody, and neither are monocyte/macrophages of human, canine or feline origin. Antibody 1.646 recognizes two proteins (150 and 30 kDa) of equine monocyte-derived macrophages when assayed by Western immunoblot. Because of the distribution of staining (tissue mononuclear phagocytes, lipofuscin-containing storage granules, biliary and renal tubular epithelium, and some neutrophils) we hypothesize that antibody 1.646 recognizes a cytoplasmic antigen that is closely associated with lysosomal membranes.

Pitfalls in the histochemical demonstration of alpha-glucan phosphorylase activity in glycogen-depleted skeletal muscle fibres.

In the present communication, an investigation is described into the reliability of histochemical methods for the demonstration of alpha-glucan phosphorylase activity in glycogen-depleted skeletal muscle fibres. Human skeletal muscles with glycogen-depleted fibres from patients with diseases of the neuromuscular system and from subjects who had suffered from malignant hyperthermia were used for the study. The location of phosphorylase activity and glycogen was demonstrated with histochemical techniques. Biochemical techniques were used to assay the activity of phosphorylase and the content of glycogen. Biochemical determinations of phosphorylase activity did frequently not reveal significant differences between glycogen-depleted and non-depleted skeletal muscle fibres. In contrast, all histochemical methods investigated, showed little or no phosphorylase activity in the glycogen depleted fibres, indicating that none of the existing histochemical methods revealed reliable staining results in these fibres. Owing to the invalid staining results of the histochemical methods for glycogen-depleted muscle fibres, it is necessary that for metabolic studies a biochemical assay for phosphorylase activity is also to be performed.

A method for reliable and permanent intracellular staining of retinal ganglion cells

We have developed a method for reliable, permanent, high-resolution intracellular staining of ganglion cells in mammalian retinas. Living ganglion cells in the isolated retina are impaled in vitro and injected intracellularly with both Lucifer Yellow (LY) and biocytin. After fixation and aggressive pretreatment of the retina with detergents, the LY is tagged immunohistochemically with biotin using a commercially available anti-LY antibody and a biotinylated secondary antibody. A conventional avidin-biotin procedure is then used to visualize both the biocytin and the biotinylated bridge antibody, yielding complete Golgi-like filling of the soma, dendrites and axon. Advantages of the method include the ease and speed of dye injection, the reliable recovery of stained cells, the large number of cells which can be stained in single retinas, and the high resolution and permanence of the stain, which permit prolonged examination and quantitative analysis.

Viability and acrosome staining of bull, boar and rabbit spermatozoa.

A practical and reliable staining procedure was developed to distinguish the viability and acrosomal status of bull, boar and rabbit spermatozoa. The first stain with trypan blue or Congo red is rapid and avoids artifacts. This stain is precipitated by neutral red during the 2 min required for fixation. The precipitate gives a high contrast black color, resistant to the subsequent rinsing and persists during the time required for staining the acrosome with Giemsa. Ten classes of spermatozoa are distinguished (live or dead with intact acrosomes, loose acrosomes, damaged acrosomes, no acrosome, or with no acrosome and no postacrosomal ring). The intact acrosomes are purple, the loose acrosomes are dark lavender and the damaged acrosomes are pale lavender. The anterior part of the head of live spermatozoa with no acrosome is white or light pink and the same area of dead spermatozoa is white or pale gray. The postacrosomal ring is red. The postacrosomal area of the head of live spermatozoa is white or light pink and the same part of dead spermatozoa is black, dark violet or gray. The procedure did not give satisfactory results for stallion spermatozoa.

Rapid, Reliable and Economical Silver Stain for Neurofibrillary Tangles and Senile Plaques

AbstractA simple, rapid, sensitive, consistent and economical method for plaques and tangles of Alzheimer's disease is presented. Sections were sensitized in 0.5% ammoniacal silver nitrate, followed by treatment in gum mastic and immersion in a physical developer consisting of hydroquinone, gum mastic and a very low concentration of silver nitrate. They were then treated with gold chloride and sodium thiosulfate. The procedure gives comparable results to those obtained with Bielchowsky and its modifications in about 30 minutes. (The J Histotechnol 14:39, 1991)

Staining Plant Cells with Silver. I. The Salt-Nylon Technique

A technique is described for selectively silver staining nucleoli, active nucleolus organizers, nucleolar material attached to chromosomes, kinetochores, synaptonemal complexes, and chromosome cores in plant cells. The technique, called salt-nylon silver staining, involves spreading cells on glass slides, treating the cells with a solution of saline sodium citrate, and incubating the cells in a silver nitrate solution covered with nylon screen. Selected variables important for achieving reliable silver staining are considered.

A Multichromatic Stain for Lowicryl K4M Embedded Tissues

A staining procedure using celestine blue and chromotrope 2R for Lowicryl K4M embedded tissues is presented. The stain produces a reliable multichromatic stain for light microscopy on Lowicryl embedded semithin sections.

Rapid, Reliable and Economical Silver Stain for Neurofibrillary Tangles and Senile Plaques

Rapid, Reliable and Economical Silver Stain for Neurofibrillary Tangles and Senile Plaques

NB84: A new monoclonal antibody for the recognition of neuroblastoma in routinely processed material

A new monoclonal antibody NB84 recognizing an uncharacterized molecule of 57 kD was produced using human neuroblastoma tumour tissue as a source of antigen. The antibody was selected for its ability to give reliable staining on conventional formalin-fixed, paraffin-embedded material. In the present study, NB84 was assessed for its usefulness in the differential diagnosis of paediatric small cell tumours. It gave reliable staining of 17/19 (90 per cent) neuroblastomas and was negative on all other tumours apart from Ewing's sarcomas, of which 9/14 (64 per cent) showed mainly focal positivity. It is concluded that in association with a panel of markers against leucocyte, muscle, epithelial, and neural markers NB84 provides considerable assistance in the recognition of neuroblastoma and thus is important in enabling the most appropriate therapy to be given.

Microwave Assisted Staining of Nerve and Muscle Biopsy Tissue

This paper reports a technique using microwaves to assist penetration of stains into biopsy sections of muscle and peripheral nerve. The technique results in more consistent and reliable staining of tissue sections for examination by light microscopy.

A Rapid and Reliable Sequential Staining Method for Bovine Muscle

Various different combinations of histochemical stains at differing pH levels on single sections of bovine skeletal muscle were evaluated. The staining sequence that was the most consistent and reliable of those studied included an initial NADH-TR (reduced nicotinamide adenine dinucleotide-tetrazolium reductase) followed by an acid ATPase stain at pH 4.15. This staining combination resulted in easily discernable type I, II A, and II myofibers. Thus, a marked saving of expended time, labor and materials was realized.

A reliable method for simultaneous demonstration of two antigens using a novel combination of immunogold-silver staining and immunoenzymatic labeling.

We have developed a reliable and sensitive immunohistochemical staining technique which allows the simultaneous demonstration of two different antigens expressed in or on the same cell (referred to as mixed labeling), together with the evaluation of the general histopathological appearance of the tissue. The staining procedure combines a three-step (streptavidin-biotin) immunogold-silver staining (IGSS) with a three-step immunoenzymatic labeling. For this purpose, we investigated the compatibility of IGSS with various substrates of peroxidase or alkaline phosphatase (AP). Highly reliable and discernible mixed labeling was achieved only after initial labeling with IGSS followed by AP labeling using the substrates naphthol AS-MX phosphate/Fast Blue or naphthol AS-BI phosphate/New Fuchsin, respectively. To ensure utmost specificity, we applied FITC-conjugated mouse monoclonal antibodies and rabbit anti-FITC immunoglobulins visualized by AP-labeled immunoglobulins and the respective substrate in a final step. This novel approach provides an excellent means for demonstration of immunocompetent cells and unequivocal determination of the percentage of specific cell subsets in infiltrated tissue. The advantages of this method, as compared with double immunofluorescence or double immunoenzymatic labeling, were investigated and are discussed.

A Microwave Oven Method for the Combined Alcian Blue-Periodic Acid-Schiff Stain

A rapid microwave modification of alcian-blue-periodic acid-Schiff stain is described. The method gives consistent and reliable staining of acid and neutral mucins within 15 minutes. (The J Histotechnol 12:295, 1989)

A Simple, Reliable and Inexpensive Silver Stain for Nerve Fibers in Bleached Skin

A procedure for silver staining is described which leads to the selective and reliable impregnation of nerve fibers in bleached skin of vertebrates and invertebrates. In combination with osmium, the protocol enhances the staining of secondary sensory cells of mechanosensory and electrosensory organs so that the innervation pattern of each organ and the number of sensory cells per organ can easily be evaluated. The technique can be also used for staining nerve fibers in whole embryos.

LR-White and LR-Gold resins for postembedding immunofluorescence staining of laminin in mouse kidney.

This work describes a method for the immunolocalization of laminin on 1 micron-thick tissue sections using a postembedding immunofluorescence technique. Embedding of unfixed or formaldehyde-fixed mouse renal cortex in either of the acrylic resins LR-White or LR-Gold permitted reliable postembedding immunofluorescence staining for laminin. LR-White was heat-cured at 50 degrees C whereas LR-Gold was polymerized at -25 degrees C. A stronger immunostaining for laminin was obtained from tissue embedded in polymerized LR-Gold compared with the staining from tissue embedded in LR-White. Prerequisites for adequate postembedding immunostaining are the partial dehydration of the tissue (maximum ethanol concentration, 70%) and pepsin treatment of the tissue sections prior to performing the immunostaining reactions.

The use of tetracycline hydrochloride as a rapid fluorescent stain for myelin membranes in vertebrates and invertebrates

We report here the use of tetracycline as a fast and reliable histological stain of myelinated nerves. These results are significant in that they point to tetracyclines as potentially important probes to study the role of divalent and trivalent cations in myelinated nerves. The usefulness and specificity of tetracycline as a supravital probe for calcium and iron metabolism in myelinated nerves is currently under study.

Eastwood Congo Red Method For Demonstrating Amyloid

A modification of Eastwood's Congo red method is described. The method, which substitutes iron hematoxylin in place of alum hematoxylin, appears to enhance the staining of amyloid when viewing with light microscopy, at the same time quenching background fluorescence when viewing with fluorescence microscopy. The method gives consistent and reliable staining results, and the Congo red staining solution has excellent stability. (The J Histotechnol11:105, 1988.)

A Rapid and Reliable Silver Impregnation Method forPneumocystis Cariniiand Fungi

AbstractA modified Grocott's methenamine-silver method for the rapid demonstration of Pneumocystis Carinii and fungi is described. Tissue sections are imbued in a dilute methenamine silver solution, then impregnated in a microwave oven, which is used as the initial heating source to promote rapid deposition of metallic silver in tissue structures. The sections are then transferred to a conventional oven to promote even, consistent, rapid, and reliable staining. (The J Histotechnol 11: 27, 1988.)

Histopathology of experimental Aspergillus fumigatus keratitis

Histopathological studies in rabbit's eyes, 7 and 14 days after intracorneal inoculation with 1 X 10(5) Aspergillus fumigatus conidia have been performed. Similar lesions were found in both periods with fungal hyphae in the anterior third of corneal stroma, round cell infiltration from the sclero-corneal edge and in the anterior chamber and, neovascularization. No lesions were found in the Descemet's membrane. Gomori silver-methenamine stain with hematoxiline-eosine counter-stain was found to be the most reliable stain to detect fungal presence in corneal stroma, and Masson's trichromic stain in the study of pathological changes in ocular elements.