A Selective Stain for Renal Basement Membranes

Paraffin sections from human kidneys fixed in Carnoy's fluid No. 2 were treated consecutively with periodic acid-sodium bisulfite and stained with resorcin-fuchsin. Basement membranes were colored black in cross sections, dark gray in tangential sections. Cytoplasm, nuclei, reticulin and collagen fibers remained unstained or were only lightly colored, depending on duration of fixation. Elastic fibers were colored black. In sections counterstained with Kernechtrot, the sharp black coloration of basement membranes and the pink staining of nuclei facilitated the study of glomerular lesions. After counterstaining with Van Gieson's picro-fuchsin, the black basement membranes contrasted well with the red reticulin and collagen fibers. Because this method does not require differentiation, it gave uniform results in the hands of different users. No fading was observed in section stored for 3 yr.     

Alkaline Bismuth Stain as A Tracer for Golgi Vesicles of Plant Cells

An alkaline solution of bismuth subnitrate reacts well with carbohydrate-rich components of Golgi bodies in sections prepared from plant leaves fixed with glutaraldehyde and osmium tetroxide and embedded in Epon. The metal deposits formed are so fine that the stain is appropriate to ultrastructural observation at high magnification. The Golgi vesicles show polarity with respect to the localization of the reactive deposits. Golgi vesicles that had migrated farther from the Golgi cisternae showed greater reactive deposits and higher membrane contrast than those close to the Golgi cisternae. These results indicate that the alkaline bismuth stain is an excellent tracer for Golgi bodies of plant cells.     

Phosphotungstic Acid-Eosin Combined with Hematoxylin as A Stain for Negri Bodies in Paraffin Sections

Experimentation with the Papanicolaou stain in this laboratory led to the discovery that the eosin, combined with phosphotungstic acid, was responsible for differential staining of Negri bodies. Eosin prepared as in EA 36 was used, but without the light green and Bismarck brown. Paraffin sections of hippocampus from brains of animal affected with rabies were fixed in 10% formol or in a mixture of 2 volumes of saturated aqueous HgCl₂ and 1 volume of absolute alcohol. They were stained first with hematoxylin and then with eosin. This procedure gave better results than staining with other types of eosin or by the original EA 36 mixture. The Negri bodies were well stained and their structure easily visible. The best results were obtained from material fixed with the HgCl₂ solution.     

Pinacyanol Erythrosinate as A Stain for Mast Cells

An alcoholic solution of the compound dye, pina-cyanol erythrosinate when diluted to the optimum dissociation point is a differential tissue stain which, in addition, selectively stains and differentiates mast cells. It can be made up and used like any other compound dye (e.g., Bowie's stain, neutral gentian, etc. or like a blood stain). It can be used after any of the common fixatives and has the advantage of selectively staining all types of mast cells in their various functional phases, even in those species (notably rabbit and man) in which they may be difficult to demonstrate with other mast cell stains after aqueous fixatives.     

Congo Red as a Simple Stain for Beta Cells of the Hypophysis

Bennhold's Congo red method for amyloid has been found to provide a simple, one step, differential staining technique for cells of the anterior lobe of the hypophysics, and to be applicable to routine use with formalin-fixed tissue. The beta cells stain orange-red and the alpha cells yellow, while the chromophobe cytoplasm remains unstained. This method can be used on material from humans and laboratory animals. Ordinary degrees of post-mortem change do not affect the staining reaction.     

Congo Red as a Simple Stain for Beta Cells of the Hypophysis

Bennhold's Congo red method for amyloid has been found to provide a simple, one step, differential staining technique for cells of the anterior lobe of the hypophysics, and to be applicable to routine use with formalin-fixed tissue. The beta cells stain orange-red and the alpha cells yellow, while the chromophobe cytoplasm remains unstained. This method can be used on material from humans and laboratory animals. Ordinary degrees of post-mortem change do not affect the staining reaction.     

Enzymology of Plasma Membranes of Insect Intestinal Cells

The enzymology of insect intestinal cell plasma membranes isa field of scientific research that is in the earliest stagesof development. In this paper the few published studies specificallydesigned to isolate plasma membranes from insect intestinalcells and determine the enzymes associated with them are reviewedin light of both older studies that approached these problemsless directly and recent results from our laboratory. In the past few years reliable methods have been developed forthe isolation of specific portions of plasma membranes fromthe epithelial cells of the midguts of a few insect larvae.These membrane preparations have been assayed for a varietyof enzyme activities. Alkaline phosphatase, leucine aminopeptidaseand -glutamyl transpeptidase have shown promise as potentialmarkers for the plasma membranes of insect larval midgut cells.However, only the latter enzyme currently stands unchallengedas a marker for the apical portion of the plasma membrane ofinsect midgut columnar epithelial cells. No enzymes can yetbe considered to be even tentatively established as markersfor the basal or lateral portions of insect intestinal cells.     

The Cationic Chelate of Chromium and Aluminon as a Selective Nuclear Stain

The chelate k prepared by adding 4.5 gm of aluminon and 100 gm of chrome alum to 200 ml of distilled water, boiling gently for 20 min., filtering, and allowing the filtrate to drop into 3.5 liters of absolute alcohol. The alcoholic suspension is filtered and its precipitate is dried at room temperature. To prepare the staining solution 3 gm of chelate are dissolved in 100 ml of 3% HCI. Hydrated sections—paraffin, frozen, or celloidin—are stained for 30 min to 18 hr at room temperature. The stain is self-limiting and requires no differentiation. Since the stain is not removed by alcohol or weak acids, a large variety of counterstains my be used.     

Basic Fuchsin and Picro-Indigo Carmine: A Selective Stain for Cells in Mitosis and for Autoradiography

Selective staining of dividing nuclei is accomplished as follows: paraffin sections, after hydration, are stained 15 min in a saturated aqueous solution of basic fuchsin, washed, then stained 1.5 min in an equal-volumes mixture of indigo carmine saturated in 70% alcohol, and saturated aqueous picric acid. Removal of excess dye with 3 changes of 70% alcohol, dehydration, clearing and covering in a resinous medium completes the process. Nuclei of dividing cells are stained red; cytoplasm and interphase nuclei, light green. This method has been used successfully for determining the mitotic activity of skin, kidney, liver and other rabbit and mouse tissues. Tissue sections previously prepared as autoradiographs may be stained by this method to facilitate the determination of radioactive labeling of mitotic cells.     

The Use of Picric Acid with the Gram Stain in Plant Cytology

The technic described involves the use of a saturated solution of picric acid in absolute alcohol in the process of dehydration following the gentian-violet-iodine stain as applied to plant cytological material. The method is suitable for both paraffin sections and smears of pollen mother cells fixed in Navashin's or Flemming's solutions. Differentiation in clove oil is very easy since cytoplasm destains immediately, while chromatic material destains very slowly following picric acid. Chromosomes are stained more distinctly than with the usual Gram stain and do not fade.     

Glucose Transporter in Plasma Membranes of Cultured Neural Cells, as Characterized by Cytochalasin B Binding

Identification of hexose transporter sites by cytochalasin B binding was conducted with a centrifugation assay. The determination of KD and Bmax values by LIGAND computer analysis provided binding data that are similar in primary astrocytes (238 nM and 14 pmol/mg protein) and neuroblastoma cells (179 nM and 13.6 pmol/mg protein). In contrast, only an insignificant number of transporter sites was detectable in C6 glioma cells, irrespective of whether membrane fractions were obtained by a two-phase polymer system or by a latex phagocytosis technique yielding inside-out plasma membranes. The latter membrane preparation was utilized to identify and quantitate the transporter molecules at the inner membrane surface of primary astrocytes, i.e., 160 nM (KD) and 5.8 pmol/mg protein (Bmax), respectively.     

Rhodanile Blue; a Rapid and Selective Stain for Heinz Bodies

Equal volumes of heparinized or EDTA-treated blood and a 0.5% solution of rhodanile blue (E. Gurr, Michrome No. 1156) in 1% NaCl were mixed and allowed to stand for 2 rain. Thin smears were then prepared, air dried and examined under oil. Heinz bodies stained deep purple and contrasted well with the yellow-orange to blue-green cytoplasm. Durable mounts could be made by applying a cover glass with a resinous mediiun to the dry smear (D. P. X. was used). The reticular material in reticulocytes did not stain in 2 min but could be stained by allowing the stain to act 5 min.     

Hematoxylin as a Pituitary Stain

The accurate picture of the acidophil granules of the anterior pituitary which is provided by iron hematoxylin can be combined with the differential staining of the basophils by either the periodic acid-Schiff (PAS) or combined aldehyde-fuchsin-PAS procedures. To accomplish this the two stages of the iron hematoxylin technique are separated so that mordanting in iron alum precedes and application of hematoxylin follows the basophil procedures.     

C. I. Acid Red 52: A New Stain for Cells of Granulocytic Origin

Using the xanthene dye C.I. acid red 52 (CI. 45100) as a single agent stain applied to coverslip preparations of blood and bone marrow, primary and secondary granules in cells of neutrophilic origin stained brilliant pink. In eosinophils, granules stained dark red. In leukemic myeloblasts that also stained with Sudan black B and demonstrated myeloperoxidase and specific esterase activity, a few bright red staining granules were visualized with acid red 52- In some leukemic promyelocytes, Auer rods stained bright red. In leukemic lymphoblasts, no red granules were seen. Of a wide variety of dyes tested so far, acid red 52 is the most sensitive stain for primary and secondary granules of granulocytes in blood and bone marrow.     

DAPI as a Useful Stain for Nuclear Quantitation

A simple-to-use fluorescent stain, 4',6-diamidino-2-phenylindole (DAPI), visualizes nuclear DNA in both living and fixed cells. DAPI staining was used to determine the number of nuclei and to assess gross cell morphology. Following light microscopic analyses, the stained cells were processed for electron microscopy. Cells stained with DAPI showed no ultrastructural changes compared to the appearance of cells not stained with DAPI. DAPI staining allows multiple use of cells eliminating the need for duplicate samples.     

DAPI as a Useful Stain for Nuclear Quantitation

A simple-to-use fluorescent stain, 4′,6-diamidino-2-phenylindole (DAPI), visualizes nuclear DNA in both living and fixed cells. DAPI staining was used to determine the number of nuclei and to assess gross cell morphology. Following light microscopic analyses, the stained cells were processed for electron microscopy. Cells stained with DAPI showed no ultrastructural changes compared to the appearance of cells not stained with DAPI. DAPI staining allows multiple use of cells eliminating the need for duplicate samples.