Combined Staining Method Research Articles

Cytoskeletal F-actin patterns in whole-mounted larval and adult salivary glands of the fleshfly, Sarcophaga bullata

The patterns of filamentous actin were analysed in different larval, pupal and adult stages in the salivary glands of the fleshfly Sarcophaga bullata. Using the rhodamine labelled phalloidin staining method in combination with detergent extraction specific actin filament distribution was detected. The salivary glands which are histolysed during the process of metamorphosis show distinct cellular morphology and actin filament patterns in larvae and adults. The large third instar larval salivary gland cells contain a well developed apicolateral microvillar zone. In third instar larvae this microvillar zone invaginates and expands in the basal part of the lateral membranes. Larval salivary gland cells also contain numerous parallel basal actin bundles. The larval glands are histolysed during metamorphosis and adult glands are formed out of the imaginal cell group. At the onset of metamorphosis these basal actin bundles form a network of crossing bundles. The filamentous actin patterns of the proximal part of adult gland cells is confined to the apicolateral microvillar membranes. The cells in the distal, tubular part of the adult salivary glands show intense staining of their folded lateral membranes.

Small Bleeding Hemangiosarcoma of the Jejunum: Case Report and Immunocytochemical Findings

AbstractA case of a 68-year old man with intestinal bleeding caused by a small hemangiosarcoma in the jejunum is reported. Tissue specimens were examined by light microscopy with classical histochemical techniques and the immunogold-silver staining method in combination with silver acetate autometallography. Monoclonal and polyclonal antibodies to various tissue type markers and lymphocyte associated antigens were employed. Factor VIII-related antigen was negative in the neoplastic cells but positive in most endothelial cells of veins and some capillaries of the adherent tissue. However, antibodies to vimentin, the endothelial marker BMA 120, and Ulex europeus I agglutinin (UEA I) lectins gave strongly positive reactions throughout the lesion and in endothelial cells of all vessels. T lymphocytes as well as neuronal and glial structures were detected between the proliferating cells. (The J Histotechnol 13:141, 1990)

Analysis of activated human lymphocytes membrane antigens by dual laser flow cytometry

Cell surface antigens of normal human peripheral lymphocytes after activation with mitogenic lectins; phytohemagglutinin (PHA), concanavalin A (Con A), pokeweed mitogen (PWM) were analysed by dual laser flow cytometer.Normal human peripheral blood mononuclear cells were isolated by Ficoll-Conray sedimentation method. The cells were washed three times with RPMI-1640 medium and were resuspended into RPMI-1640 medium supplemented 10% fetal calf serum. The cell suspension was adjusted to a concentration of 1×106/ml and PHA, Con A and PWM were added to the cell suspension at an optimal concentration of the lectins respectively. The cells were then cultured at 37 C° humidified 5% CO2 atmosphere for 72 hrs.After 72 hrs, the cells were harvested, washed three times with phosphate buffered saline (PBS) and cell surface markers were analysed by direct, indirect and combined immunofluorescent staining methods. The combined staining method was carried out first, by indirect staining with tetramethyl rhodamine isothiocyanate (TRITC) conjugated goat anti-mouse IgG serum as the secondaly reagent and second, by direct staining with fluorescein isothiocyanate (FITC) conjugated T4 and T8 mouse monoclonal antibody.Activated T cell antigen recognized with anti-Tac monoclonal antibody, other T cell antigens recognized with T4, T8, OKT-9, OKT-10 monoclonal antibodies and immune associated (Ia)-like antigen recognized with I2 monoclonal antibody were markedly expressed on the cell surface of lectin stimulated normal humal peripheral lymphocytes.Through the dual laser flow cytometric analysis, all T4 positive cells exhibited the Tac antigen. The T8 positive cells also exhibited the Tac antigen but not on all cells. The Tac negative but the T8 positive and double marker cells representing both T4 and T8 positive were also demonstrated.Acute lymphoblastic leukemia antigen (CALLA) with J5, B cell antigen with Bl and myelomonocyte antigen with Mo 2 monoclonal antibody were not recognized on the distribution of two membrane surface antigens on the cells with a single procedure.

Analysis of nucleolar components by combined silver staining and serial sections procedures

The nucleolus in Sertoli cells, spermatogonia and spermatocytes of rabbit testis was studied under electron microscopy using a silver staining method in combination with serial section reconstruction. Fibrillar centers (Fc) as part of the nucleolar organizer region (NOR) are usually observed in these nucleoli and appear poorly stained after silver method which stain nucleolar acidic pr o teins. Silver stains preferentially part of the dense fibrillar components of the nucleolus which appears like strings inside the nucleolar mass emerging from part of the fibrillar center. The quantity and extent of the silver reaction seems to be correlated to the degree of transcriptional activity of each type of nucleolus. Moreover, the methods used allow the topography of the different nucleolar components to be inferred.

Types of enzymatic overdosing in trisomy 21: erythrocytic superoxide dismutase-AJ and phosphoglucomutase.

A comparative study was carried out on the hemolysates of 6 trisomic 21 and 6 normal subjects, by electrophoresis in starch gel, determining by a combined staining method both SOD-A (former IPO-dimer) and PGM activity. The enzymes were found statistically to be in a hyperactive status, the ratio of trisomic to normal values being approximately equalt to 1.4. SOD-A supraactivation is the effect of a genic dose, as demonstrated in earlier works (Sichitiu, 1973; Sichitiu et al., 1974; Sinet et al., 1974), whereas PGM hyperactivity appears to be modified secondarily, the same as the activity of other cellular enzymes in Down's disease.

An Alcoholic Eosin and Methylene-Blue Staining Method

ABSTRACT Aqueous or alcoholic solutions of eosin and aqueous solutions of methylene-blue have long been individually and successively employed for the double staining of sections. (It is not intended here to deal with the mixed or combined staining method of Romanowsky (8), Ehrlich (1), and others.) The various methods now in use (Schafer (9), Sims Woodhead (10), Miller (5), etc.) may be generally described as follows :