AN IMMUNOGLOBULIN-ENZYME BRIDGE METHOD FOR LOCALIZING TISSUE ANTIGENSMASON, T. E.; PHIFER, R. F.; SPICER, S. S.; SWALLOW, R. A.; DRESKIN, R. B.
doi: 10.1177/17.9.563pmid: 4186250
An immunohistochemical method not requiring conjugation of a label to antibody is described for visualizing tissue antigens. The method depends on binding of an enzyme label to the tissue antigen through the antigen-antibody reactions of an immunoglobulin-enzyme bridge. This bridge is composed of the following components in order: (a) specific antiserum to the tissue antigen, (b) antiserum against the Immune globulin of the species for component a, (c) specific antiserum prepared against the enzyme label in the same species as component α and (d) enzyme label. The bridge method employing horseradish peroxidase as the enzyme label has been applied to localization of cells forming growth hormone and chorionic gonadotrophin in the human adenohypophysis and placenta respectively.
ELECTRON MICROSCOPIC DEMONSTRATION OF ADENOSINE TRIPHOSPHATE PHOSPHOHYDROLASE ACTIVITY IN HERRING GULL SALT GLANDSABEL, JOHN H.
doi: 10.1177/17.9.570pmid: 4186251
Sites of adenosine triphosphate phosphohydrolase activity were demonstrated in herring gull salt glands with a cytochemical lead technique, utilizing fixed and unfixed tissue sections and a modified Wachstein and Meisel incubation media. The principal modifications in the procedure as recommended by Wachstein and Meisel included shortening the time of fixation and decreasing lead concentration while increasing the substrate concentration of the incubation media. Several different types of controls were run. In fixed tissues tested with an unmodified media, reaction product was deposited in capillary endothelial cells, tubular lumens and intercellular channels of central canal cells and nucleoli. Evidence is presented that none of these reactive sites contain a true adenosine triphosphate phosphohydrolase. With a modified procedure adenosine triphosphate-specific reaction product was deposited within the matrix of mitochondria, along the inner surface of the plasma membrane that lines the base of the principal salt-secreting cells and on the centriolar tubules. The possible validity of these reaction product localizations is discussed.
LIGHT MICROSCOPIC STUDY OF THE PEROXIDATIC ACTIVITY OF CATALASE IN FORMALDEHYDE-FIXED RAT LIVERHIRAI, KEI-ICHI
doi: 10.1177/17.9.585pmid: 4186252
The peroxidatic activity of rat liver catalase was demonstrated by histochemical staining with 3,3'-diaminobenzidine as hydrogen donor. The activity was so weak that its location was hard to identify in formaldehyde-fixed cells, although high catalatic activity was present, as evidenced by the production of bubbles upon the addition of hydrogen peroxide to the incubation medium. Pretreatment of fixed sections for 60 min at 37°C with formamide, urea or trypsin enhanced the peroxidatic activity significantly. The reaction granules scattered throughout the cytoplasm of the parenchymal cells probably correspond to the peroxisomes.
CHANGING NUCLEAR HISTONE PATTERNS DURING DEVELOPMENT III. THE DEOXYRIBONUCLEIC ACID CONTENT OF SPERMATOGENIC CELLS IN THE CRAB EMERITA ANALOGAVAUGHN, J. C.; LOCY, R. D.
doi: 10.1177/17.9.591pmid: 4186253
The deoxyribonucleic acid (DNA) content of spermatogenic cells of the decapod crab, Emerita analoga, has been cytophotometrically determined at various stages of development, using Feulgen-stained nuclei. The haploid DNA value is found to be 2.9 x 10–12 g, regardless of the nuclear histone content, which drops to cytochemically indetectable levels by sperm maturity. In determining this value, a precise extinction coefficient for Feulgen-stained nuclei was first determined using chicken, frog and toad erythrocyte nuclei and also nuclei from various rat tissues. The effect of naturally occurring nuclear histone depletion on the quantitative Feulgen reaction has also been examined. Identical hydrolysis curves and Feulgen spectral absorption curves are found for somatic nuclei, which contain a full histone complement, and for mature sperm nuclei, which do not contain histones. Loss of nuclear histone does not lead to a change in total Feulgen dye bound per nucleus, as early, middle and late spermatids have equal DNA contents as reflected by Feulgen binding, and primary spermatocytes contain 4 times this value, as expected from the DNA constancy law. The effect of histones (and other proteins) on quantitative Feulgen microspectrophotometry is discussed.
CHANGES IN ENZYME ACTIVITIES OF VARIOUS MUSCLE FIBER TYPES IN RAT INDUCED BY DIFFERENT EXERCISESKOWALSKI, K.; GORDON, E. E.; MARTINEZ, A.; ADAMEK, J.
doi: 10.1177/17.9.601pmid: 4186254
Mutability of enzyme activities (phosphorylase, succinic dehydrogenase, cytochrome oxidase) of red and white fibers was studied in rat quadriceps subjected to normal physiologic chronic exercises. A rise in phosphorylase activity was found in weight lifting and to a greater extent in running rats when muscle was taken as a whole, but both exercises resulted in equal increments for succinic dehydrogenase and cytochrome oxydase activities. Intraregional comparisons, however, revealed the greatest relative rise of succinic dehydrogenase activity in those fibers that were regarded as predominantly anaerobic in type. This effect was seen only with running and not with weight lifting. Statistically unproved but frequently observed after running in some of the preparations was a rise of phosphorylase activity in red fibers, although to a lesser degree than in white. Thus, in contrast to the dichotomy apparent in electrophysiologic events in nerve and its dependent muscle, metabolic demands may alter what is regarded normally as fixed fiber enzyme patterns. Whole muscle cannot be studied as a biochemical entity because of diverse regional responses to the same stimuli. Endurance exercise (running) and brief, forceful exercise (weight lifting) produced quantitatively different regional changes in succinic dehydrogenase and probably in phosphorylase activities.
LEAD ION AND PHOSPHATASE HISTOCHEMISTRY III. THE EFFECTS OF LEAD AND ADENOSINE TRIPHOSPHATE CONCENTRATION ON THE INCORPORATION OF PHOSPHATE INTO FIXED TISSUEROSENTHAL, ALAN S.; MOSES, HAROLD L.; TICE, LOIS; GANOTE, CHARLES E.
doi: 10.1177/17.9.608pmid: 4186255
This communication attempts to separate and define the relationships between lead inhibition of tissue adenosine triphosphatase activity, lead. catalyzed adenosine triphosphate hydrolysis and reaction product localization when the Wachstein-Meisel reaction is applied to kidney. Using a radiochemical assay of adenosine triphosphatase activity and varying the concentration of lead nitrate or adenosine triphosphate, the quantity of phosphate bound to and released from tissue was determined. Depending on the relative concentrations of lead and adenosine triphosphate, two situations may exist. With low lead or high adenosine triphosphate concentrations, phosphate release by tissue exceeds phosphate trapped by tissue and substantial quantities of phosphate are lost to the medium. With low adenosine triphosphate or high lead concentrations more phosphate is bound in tissue than can be attributed to tissue enzyme activity. Possible explanations for these phenomenon are discussed.