The development of new techniques has permitted new scientific assaults on the understanding of living bodies. The application and combination of both classical and modern techniques offer new ideas for several kinds of research. The technique of whole-body autoradiography, which was first introduced in 1954 by Sven Ullberg, has been employed to examine the distribution of drugs and other chemicals within the animal body. This technique makes it possible to examine and compare the radioactivities of many organs and tissues under identical conditions.
For over a decade, the distribution of labelled sugars in the mammalian body and the uptake of their metabolites by the tissues and organs have been studied in our laboratory. This article will focus on the whole-body autoradiographic studies of this area. As we have been trying to develop techniques of whole-body sectioning for histological and histochemical analysis for the purpose of confirming and extending the information obtained from whole-body autoradiographs, these techniques are also reviewed. Several applications of the whole-body sections will be discussed, and an idea concerning the metabolic cooperation of tissues and organs in the whole-body will be proposed.

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Using the endogenous peroxidase reaction with the diaminobenzidine, I was possible to identify various types of leukocyte populations in the intercellular spaces of the junctional epithelium (JE) and adjacent connective tissue of healthy rat gingivae. Namely, neutrophils predominated, but monocytes and lymphocytes were few and eosinophils were very rare. No basophils were observed. Each leukocyte was identified by the number and shape of the peroxidase-positive and -negative granules at the electron microscopic level. Peroxidase was localized only in the azurophil granules of mature neutrophils and monocytes. The mature eosinophils also contained peroxidase only in the granules, which consisted of a peroxidase-positive matrix and a peroxidasenegative single discoid core in an equatorial position. No organellae of lymphocytes showed peroxidase-positive activity. In the inhibitory experiments for peroxidase of the identified leukocytes, 0.1 and 1M KCN inhibited the peroxidase moderately and almost completely, but 0.01M KCN and 0.01, 0.1 and 1M 3-amino-1, 2, 4-triazole (AMT) had no inhibitory effect. The endogenous peroxidase in the granules of the leukocytes, mainly neutrophils, was sensitive to KCN but insensitive to AMT. Although I found no evidence of a direct effect of the peroxidase on foreign stimuli, my findings suggest that the various leukocytes always patrol the JE and adjacent connective tissue of the healthy rat gingivae to protect against foreign stimuli.

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Using blood-group-specific lectins conjugated to horseradish peroxidase, distribution of A, B and H antigen was examined in formalin-fixed, paraffin-embedded pancreases from 27 human autopsy cases. Dolichos biflorus agglutinin, Griffonia simplicifolia agglutinin I-B₄, and Ulex europaeus agglutinin I or Lotus tetragonolobus agglutinin were found to be satisfactory to demonstrate A, B and H antigen, respectively in acinar cells. No exception was encountered to the correspondence between blood type and lectin staining. In blood group O individuals, acinar cells producing H antigen were evenly distributed throughout the gland. However, in other blood groups, the distribution of antigens was not homogeneous. In blood group A individuals, some acini contained A antigen but not H antigen, others contained only H antigen and a few contained both A and H antigen. Such a mosaic distribution of antigens was also observed in B and AB individuals. Furthermore, in AB individuals, independent production of either A or B antigen in some acini was observed. The blood group antigens in acinar cells were demonstrated irrespective of secretor status of the tissue donor. Although Ulex europaeus agglutinin I reacted ubiquitously with vascular endothelia independently of the blood group, other blood-group-specific lectins could not react with the endothelia of any given blood group.
The present study further demonstrated that the staining patterns of some blood-group-nonspecific lectins exhibit certain dependence on the blood group of the tissue donor. That is, soybean agglutinin reacted with the cells secreting A and/or H antigen but not those secreting B antigen; peanut agglutinin reacted with many acinar cells in all the nonsecretors examined, but in secretors, existence of such cells was very rare.

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In avian smooth muscle, desmin is a major protein composed of intermediate filaments and is localized at the Z-line of myofibrils. It has been reported that in liver tissue, desmin is restricted to certain smooth muscle cells of the large blood vessels in the portal triads. We have recently identified desmin in fat-storing cells (Ito cells) of the Wistar rat liver by means of immunocytochemical techniques using rabbit IgG antibody to chicken gizzard desmin. We studied the immunologic cross-reactivity of this desmin antibody in fat-storing cells of vertebrates. Fresh liver tissues were obtained from a cyclostome (lamprey), fish (carp, eel, young yellowtail, saurel), amphibians (bullfrog, newt), reptiles (striped snake, viper, soft-shelled turtle), birds (quail, linnet) and mammals (guinea pig, mouse, rat, pig, cat, dog, rhesus monkey, man). Desmin was identified in fat-storing cells by indirect immunofluorescent microscopy of frozen tissue sections. Fat-storing cells of a cyclostome, amphibians, reptiles and birds clearly reacted with this desmin antibody. In mammals, however, fat-storing cells of the rhesus monkey and man were not stained by immunoperoxidase and immunofluorescent techniques. Similarly, the fat-storing cells of fish were not stained.
Essentially, these immunocytochemical studies using chicken gizzard desmin antibody provide evidence of the immunologic cross-species reactivity to the desmin found within fat-storing cells of many vertebrates. However, the fat-storing cells of fish, the rhesus monkey and man do not appear to demonstrate cross-reactivity. This finding would suggest either that desmin is immunologically heterogeneous or that there are differences in the accessibility of the antigenic determinants of desmin within the fat-storing cells of the latter species.

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Immunohistochemically detectable S-100 protein in 76 salivary pleomorphic adenomas was found in two types of myoepithelium derived neoplastic cells; spindle-shaped anastomosing cells and round cells. Spindle-shaped tumor cells displayed a strong presence of S-100 protein-like immunoreactive substance (LIS), and they were usually located in the outer layer of duct-like structures and showed a continued proliferation into the stroma. Round tumor cells abundantly gave a S-100 protein LIS staining, and they were distributed in the outer layer of duct-like structures. Chondroid cells and chondrocytes in hyalinous and myxomatous tissue were also reactive for S-100 protein LIS. In rare cases, stromal connective tissue fibers exhibited an intense S-100 protein LIS staining. Peripheral nerve fibers and myoepithelial cells in the affected salivary glands were also positive for S-100 protein LIS.

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A relative cytophotometry was effectively applied for quantitative determination of succinate dehydrogenase (SDH) activity in rat liver and skeletal muscle section. The sections were stained first for SDH activity and then superimposed with protein staining by naphthol yellow S (NYS). The absorbance at 570nm and 450nm were measured cytophotometrically for the amount of TNBT-formazan and tissue protein content. The final result was expressed as a relative activity (RA) based on the ratio of the amount of TNBT-formazan to tissue protein content for the compensation of uneven section thickness. The linear relationship was obtained between the time of incubation and RA value of SDH in rat muscle section, as well as in rat liver section.
In application, the present method has revealed the different frequency distribution profiles of RA of SDH in type I and II muscle fibers which were identified on the basis of the myofibrillar actomyosin ATPase activity. The RA value was less scattered in type I fiber than in type II fiber and the type II fiber could be subclassified into two groaps according to the RA-frequency profile for SDH in muscle fibers. This observation was almost in agreement with that in other studies on SDH activity-frequency profile for type II fiber.
Thus, the present method was applicable to the cytophotometric determination of SDH activity in tissue sections.

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The distribution of α-D-N-acetylgalactosamine residues on the membranes of axon-myelin-Schwann cell complexes of rabbit and rat sciatic nerves was visualized by means of a one-step method with HRP labelled soybean agglutinin. The compact myelin was found to be very rich in lectin binding sites whereas mesaxons and split myelin lamellae, as well as the Schwann cell plasma membrane were almost negative. The labelling of the axolemma was most pronounced in the nodal and paranodal regions. The smooth endoplasmic reticulum and mitochondria in axons and glial cells were also labelled.

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An attempt has been made to obtain evidence for the localization of superoxide dismutase (SOD) activity in rabbit alveolar macrophages (AMφ). SOD localization was investigated with two different superoxide (O^-₂)-generating systems, the potassium superoxide (KO₂) and the xanthine-xanthine oxidase system. The hydrogen peroxide (H₂O₂) generation site, which indicates SOD activity, was electron cytochemically investigated by using the cerium method introduced by Briggs et al.
In the KO₂ system, the formation of cerium perhydroxide serves as an indication of SOD activity, and its presence was confirmed throughout the cytoplasm and mitochondria, but not in the nucleus and granules. Under the action of diethyldithiocarbamate and potassium cyanide, which act as Cu, Zn-SOD inhibitors, it was found that the cytoplasmic SOD activity was virtually completely suppressed, and that complete inhibition of SOD activity occurred in the presence of catalase (a scavenger of H₂O₂) or in the absence of KO₂. In the xanthine-xanthine oxidase system, cerium deposits were observed in the cytoplasm, but rarely in the mitochondria. Occasionally some reaction product was seen in the nucleus and granules.
From the results, it is concluded that KO₂ system is an excellent method for investigation of the localization of SOD activity.

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To make a biophysical standard for histological identification of fish teeth, fluorescence and Raman-spectra are employed for typical teeth of various fish, such as the sea-bream, shark, trout, and hagfish in comparison with human teeth. The tooth tissues of sea-bream, mainly containing hydroxyapatite, resemble those of human teeth. In the teeth of the trout and hagfish, no difference can be found between the surface and deep layers and they appear like decalcified human cement. Shark teeth, rich in fluoroapatite, are much different from both human enamel and dentin. According to the similarity of Raman-band patterns, it is possible to arrange them in order from trout to sea-bream through the human enamel and finally to the shark. These suggest that the structural characteristics off teeth in the fish kingdom do not coincide with differentiation of human tooth tissues.

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Potassium pyroantimonate staining has been utilized to localize calcium (Ca) in biological specimens at the electron microscopic level. This method, however, has been criticized by many investigators because of its specificity to Ca. X-ray microanalysis (EDX) of that reaction product, pyroantimonate-Ca, was an attempt to confirm Ca in it. There is a technical probrem in distinguishing Ca and antimony (Sb) by EDX because the characteristic X-ray emission energies of Sb and Ca are too close each other to be distinguished.
Therefore, the reaction product localized in the cytoplasm and over the plasma membrane of type II cells in taste buds of four wild Japanese monkeys was analysed with an electron energy loss spectroscope (EELS) combined with STEM image mode. Ca peak was recognized in the EELS spectrum with the addition of 2% K-pyroantimonate to 4% glutaraldehyde. Similar addition of K-pyroantimonate to osmium tetroxide was not essential to preserve Ca for EELS survey. Thus, EELS appeared to be more advantageous as compared to EDX in detection of Ca in histochemical reaction product.

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This paper describes the age-related and hormonal bursectomy-induced changes in the frequency of appearance of α-naphthyl butyrate esterase (ANAB)-positive cells in chicken peritoneum. The phagocytic ability of macrophages was determined with sensitized and non-sensitized sheep erythrocytes (SRBC).
The increase in the number of ANAB-positive cells ranged from 65% at 1 week of age to 90% at 2 and 4 weeks of age in controls. With testosterone propionate (TP) treatment during embryonic development, 98% of the cells taken were positive for ANAB at 1 week of age.
Some difference was found in the non-specific phagocytosis, but not in the specific phagocytosis, between macrophages harvested from the normal and TP-treated groups. TP treatment induced enhancement of the non-specific phagocytic activity of macrophages. No correlation was found in the present study between non-specific phagocytosis and the appearance of ANAB-positive cells.

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A mouse monoclonal antibody produced by a hybridoma method and immunization with isolated normal human epidermal cells reacted with the cell surface of the basal cells of the stratified squamous epithelia and appendages but not with the basal laminar surface of these cells. The antibody showed cross-reactivity with the vascular components such as endothelial cells and pericytes of the capillaries and capillary endothelium of the renal glomeruli, hepatic sinusoids and alveolar walls of the lung.

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