Differences In Staining Intensity Research Articles

DEOXYRIBONUCLEIC ACID CYTOPHOTOMETRY OF STAINED HUMAN LEUKOCYTES I. DIFFERENCES AMONG CELL TYPES

The deoxyribonucleic acid (DNA) content of individual human leukocytes was estimated cytophotometrically using visible light and spreads stained either with gallocyanin-chrome alum following ribonuclease digestion or with the Feulgen reaction. When the cells were measured on a scanning cytophotometer, significant differences in stain intensity were found among slides. Significant differences also were found among the leukocyte types. In gallocyanin-chrome alum preparations, monocytes measured 16% higher than small lymphocytes and 13% higher than neutrophilic granulocytes. In Feulgen preparations, monocytes measured 4% higher than small lymphocytes and 6% higher than neutrophils. These differences among cell types were independent of donor and stain intensity. Measurements of cells within types and within slides frequently showed close agreement, but it is only in this very limited context that the data are consistent with the hypothesis of DNA constancy. Measurements made on a two-wavelength cytophotometer showed a divergence of only 2.1% relative to similar measurements made on the scanning cytophotometer, which suggests that the differences observed among cells and types are unlikely to be artifacts of the instruments. Over-all, the data indicate either that there is variability in DNA content or that DNA is not being expressed correctly by the measured stain content.

An in situ study of the preoptico-neurohypophysial complex of the freshwater teleost Ciarias batrachus (L.)

The complicated architectural pattern of the preoptico-neurohypophysial neurosecretory system of Clarias batrachus is described using bulk staining procedure. The nucleus preopticus is U-shaped; its limbs constitute the PMC and PPC. The difference in staining intensities noticed among the PMC and PPC might suggest a possible functional difference within the system. Axons of the PMC diagonally pass through the PPC and emerge along with those of the latter to form the right and the left main tracts. The folds and recurrent curves of the neurosecretory axonal tracts, the formation of several pairs of lateral tracts and their union to form median tracts might be devices to increase the storing and releasing capacity for the neurosecretory material. The presence of prominent Herring bodies, and diffuse areas in the neurosecretory tract suggest possible sites of release of neurohormones along the highly vascular hypothalamic region. The common neurosecretory tract is divided into several smaller branches at the level of the pars distalis prior to entering the pars intermedia. Branches do penetrate into the pars distalis, but those entering the rostral component of the pituitary are relatively scarce.

Polymorphism of plasma esterases in flounder and plaice.

Esterase activity was detected in starch-gel electropherograms of plasma samples from two species of teleost: flounder and plaice. In both species polymorphism was found. In flounder its genetic origin was indicated by population genetical data. Differences in staining intensity of the individual bands were found to be correlated with length of the animals.

NUCLEOLONEMA AND PARS AMORPHA IN ROOT MERISTEMATIC CELLS OF VICIA FABA

In osmium-fixed, Epon-embedded root meristematic cells of Vicia faba stained according to the Feulgen – methylene blue procedure, the interphase nucleolus is seen to consist of a metachromatic cabbage-green colored material in which are embedded numerous unstained vacuole-like structures of varying size. By using an orange Wratten filter in the illumination system, it is observed that the metachromatically stained nucleolar material is, in fact, made up of two components, each segregated into zones distinguishable by differences in staining intensity. The first more lightly stained component is found in the central and bordering regions of the nucleolus as well as in a number of areas of varying width extending radially in between them. The second more intensely stained component is observed in areas of varying length and width usually occupying the intermediate regions of the nucleolar mass. Under electron microscopy, the first component is seen to consist of densely packed fibrils and granules some 150 Å in diameter; in places, those constituents are assembled into seemingly coarse, thread-like structures approximately 0.1 μ in diameter. The second component, on the other hand, is made up mainly, if not exclusively, of tightly packed fibrils 60 to 100 Å in diameter. Judging from their topographical distribution and ultrastructural characteristics, it would seem as if the two nucleolar components just described correspond respectively to what is usually referred to as nucleolonema and pars amorpha in animal nucleoli. In addition to nucleolonema and pars amorpha, two ultrastructurally and topographically distinct types of vacuole are recognizable within the interphase nucleolus of Vicia faba. The vacuoles of the first type are usually large or medium-sized, contain loosely and uniformly scattered fibrils and granules approximately 150 Å in diameter, and are observed exclusively within the nucleolonematic zones of the nucleolus. The vacuoles of the second type are small, contain only loosely dispersed fibrils, 60 to 100 Å in diameter, and are confined to the pars amorpha of the nucleolus.

Poly(vinyl alcohol)–iodine complexes

AbstractThe poly(vinyl alcohol)–iodine blue color reaction in dilute aqueous solution has been investigated, and extinctions at the absorption maximum have been measured as a function of the concentrations of polymer, iodine, iodide ions, and boric acid. Depending upon the reaction conditions, the main absorption maximum can be made to appear at any wavelength between 580 and 700 mμ, with the longest wavelengths and highest extinctions per vinyl alcohol group showing up at high boric acid concentrations. Poly(vinyl alcohol) samples from various sources displayed, under identical reaction conditions, great differences in staining intensities. This can be ascribed to differences in the regularity of the polymer chain structures. To account for the behavior of poly(vinyl alcohol) towards iodine under a wide variety of reaction conditions, as well as for a number of irreversible or only slowly reversible phenomena, a model for the mechanism of the PVA–iodine complex formation is proposed involving a dual process of helix formation and intramolecular helix association. There is evidence to indicate that at full iodine saturation of the polymer, 12 vinyl alcohol residues supported by one boric acid molecule form a single turn of a helix which enwraps one iodine atom, out of a long polyiodide chain nested in the interior of a poly(vinyl alcohol) helix.

A Self-Limiting, Highly Selective Chromium-Hematoxylin Stain

A compound, which is probably a cationic chelate, can be isolated as a dry powder from a hematoxylin-chrome alum lake. In aqueous acid solution this compound is an excellent nuclear stain which is extremely selective, very resistant to acids and alcohols, and self-limiting. Staining time may vary from 20 min to 16 hr without causing significant differences in staining intensity. To prepare the dry stain, dissolve 10 gm of hematoxylin, 10 gm of NaOH and 70 gm of chrome alum in 600 ml of distilled water, boil 20 min, cool and filter, allowing the filtrate to drop into 3.5 liters of absolute alcohol. Filter off the precipitate formed in the alcohol, and air dry it at room temperature. The staining solution is prepared by dissolving 3 gm of the dried precipitate in 100 ml of 3% HCl.

Ontogeny of the Shoot Apex of Seedlings of Pinus ponderosa

Dormant seeds of ponderosa pine (Pinus ponderosa) were stratified and then planted in the greenhouse. Changes occurring in the shoot apex during germination and growth of the seedlings were observed and are described. Cell divisions in germinating embryos are first noted on the flanks of the apex near the cotyledons. From these loci, mitotic reactivation proceeds up the flanks of the apex and inward toward cells of the rib meristem. Cells are dividing actively in all regions of the shoot apex of seedlings 12 days after planting, but no cytohistological zonation is evident due to differences in staining intensity and in cell and nuclear size. In 64-day-old seedlings the mitotic rate is reduced and the characteristic zonation of pine apices begins to appear. Zonation is even more evident in older seedlings. These observations are discussed in relation to the concept of initials and to theories concerning factors which regulate growth in the shoot apex.

ONTOGENY OF THE SHOOT APEX OF SEEDLINGS OF PINUS PONDEROSA

Dormant seeds of ponderosa pine (Pinus ponderosa) were stratified and then planted in the greenhouse. Changes occurring in the shoot apex during germination and growth of the seedlings were observed and are described. Cell divisions in germinating embryos are first noted on the flanks of the apex near the cotyledons. From these loci, mitotic reactivation proceeds up the flanks of the apex and inward toward cells of the rib meristem. Cells are dividing actively in all regions of the shoot apex of seedlings 12 days after planting, but no cytohistological zonation is evident due to differences in staining intensity and in cell and nuclear size. In 64‐day‐old seedlings the mitotic rate is reduced and the characteristic zonation of pine apices begins to appear. Zonation is even more evident in older seedlings. These observations are discussed in relation to the concept of initials and to theories concerning factors which regulate growth in the shoot apex.

Interrelationship of aldehyde reactions and the einerson staining method in intraepithelial and invasive squamous carcinoma of the uterus

Histochemical reactions are performed on tissue sections of 17 cases of malignant cervical epithelium with normal controls, to stain cell nuclei with different aldehydestaining methods (Feulgen, Feulgen NAH, and ammoniacal silver). These are accompanied with the Einerson staining procedure as a direct nucleic acid staining method. When the four methods are compared closely, approximating characteristics are found as regards differences in staining intensities of normal and malignant cervical tissue. In all cases the malignant tissue was stronger stained in a similar degree of intensity. No principal staining differences were found in invasive cancer as compared with cancer in situ.

Cholinesterase content of certain regions of the spinal cord as judged by histochemical and cartesian diver technique.

Summary.1. The cholinesterase content of neurones belonging to the antero‐lateral group of the anterior horn of the spinal cord of rat has been investigated histochemically by means of a modified thiocholine method and microchemically by means of a Cartesian diver technique.Biochemical controls with the inhibitor (Mipafox) run in solutions similar to those used histochemically have been made.2. When rat brain homogenate was used as source of enzyme, with Mipafox (4 × 10‐6 M) as inhibitor, and ace'ylthiocholine as substrate (0.004 M), the inhibition curve fits between those of AcChE (electric tissue) and BuChE (Cohn's serum fraction).3. A histochemical survey of a section of the cervical cord shows that only anterior and lateral horns are stained. The large motor neurones are heavily stained and the product of the histochemical reaction is clearly visible along the neurites. The difference in staining intensity among various cells is not sufficiently marked to allow a quantitative appreciation of cholinesterase activity.4. By measuring the cholinesterase activity of single neurones by means of the Cartesian diver technique two significantly different groups of cells can be demonstrated. In one group the total enzyme activity per cell is approximately 4 times higher than in the other. All the cells investigated have a measurable cholinesterase activity. Experiments with a selective inhibitor demonstrated that the enzyme present in these cells is acetylcholinesterase.5. The results are discussed in the light of what is known about the morphology and physiology of the motor cell groups of the spinal cord.