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Cytoplasmic protein extracts from chronic myelogenous leukemia (CML) cells contained an activity that altered the electrophoretic mobility of complexes formed between nuclear proteins and the transcriptional enhancers of interferon (IFN)-inducible genes. Exposure of CML cells to IFN-alpha diminished the effect of the CML cytoplasmic proteins on these nuclear protein-DNA complexes. The presence of clinical responsiveness to IFN-alpha correlated with the sensitivity to the IFN-induced change in the electrophoretic mobility of nuclear protein-DNA complexes. These data suggest that the action of IFN-alpha in CML may be linked to a pathway that can result in posttranslational modification of nuclear proteins.

An immunocytochemical investigation was carried out on round and spreading hemocytes of Planorbarius corneus by using 20 antisera to vertebrate bioactive peptides. The immunotests showed the presence of alpha 1-antichymotrypsin-bombesin-, calcitonin-, CCK-8 (INC)-, CCK-39-, gastrin-, glucagon-, Met-enkephalin-, neurotensin-, oxytocin-, somatostatin-, substance P-, VIP-, and vasopressin-immunoreactive molecules in the spreading hemocytes. The round hemocytes were only positive to anti-bombesin, anticalcitonin, anti-CCK-8 (INC), anti-CCK-39, anti-neurotensin, anti-oxytocin, anti-substance P and anti-vasopressin antibodies. No immunostaining was observed with anti-CCK-8 (Peninsula), anti-insulin, anti-prolactin, anti-thyroglobulin and anti-thyroxin (T4) antibodies. As probably in vertebrates, these bioactive peptides may modulate immuno cell function.

Basic fibroblast growth factor (bFGF) is thought to be of major importance for fibrosis and angiogenesis. Despite intensive studies dealing with the biochemistry and multiple biologic effects of bFGF, the cellular distribution is virtually unknown. Therefore, using the indirect immunoperoxidase technique, we examined the effect of bFGF on a large pattern of normal, inflammatory, and tumorous human tissues. Staining was performed on cryostat sections with a highly specific affinity-purified antiserum. In normal tissues, especially those of the thymus and placenta, mainly dendritic cells contained the growth factor. High levels of bFGF were also detected in basal cells and gland epithelial cells of skin biopsies. A conspicuous expression was observed in chronic inflammatory tissues corresponding to a generally pronounced proliferation of fibroblasts and endothelial cells in these situations. Tumors revealed a very heterogenous staining pattern. In some lesions, bFGF was predominantly present in infiltrating and endothelial cells. In several, neoplasms tumor cells exhibited an intensive staining. In some, especially vascular tumors, bFGF could not be detected. From the staining results it is concluded that angiogenesis is not simply controlled by the presence of bFGF but is mediated by a balance of several angiogenic inducers and inhibitors.

Megakaryoblasts of bone marrow differentiate into megakaryocytes that in turn are the source of blood platelets. We have raised monoclonal antibodies to a megakaryoblast-like cell line derived from rat bone marrow (RPM cells). One antibody (Mab 213) and the corresponding antigen has been characterized by Western blotting and immunohistochemistry. Biosynthetic labeling with [35S]methionine showed that this antigen is synthesized by the RPM cells. In Western blots the antibody recognized proteins of about 90 kDa and 160 kDa in Triton extracts of RPM cells, whereas it recognized proteins of about 160 kDa and 200 kDa in Triton extracts of rat platelets and one of about 200 kDa in Triton extracts of various rat tissues (kidney, lung, intestine, and heart). By immunohistochemistry, the antigen was localized to the apical part of the epithelium lining certain parts of kidney tubuli, bronchi and large intestine.

Calcium-binding phosphoprotein particles are the most abundant extracellular proteins in the hemolymph of heterodont bivalves, and granular hemocytes are the most abundant cells in the same fluid. In this study, the hemocytes of Rangia cuneata were examined ultrastructurally and probed with anti-phosphoprotein IgG to demonstrate that the granulocytes are a probable source of the hemolymph phosphoprotein. The granulocyte cytoplasm is laden with large vesicles containing an amorphous homogenous matrix and variable numbers of electron-dense particles; the latter are ultrastructurally similar to the extracellular phosphoprotein. The vesicle particles and matrix are related forms of the hemolymph phosphoprotein as evidenced by heavy gold labeling when Lowicryl sections were sequentially treated with rabbit-anti-phosphoprotein IgG and colloidal gold-anti-rabbit IgG. The vesicles may be the loci for posttranslational phosphorylation and eventual secretion of the calcium-binding phosphoprotein, or alternatively the vesicles may be digestive structures which degrade internalized phosphoprotein.

Neutral and acid glycosphingolipids of Echinococcus multilocularis metacestodes that were obtained after intraperitoneal infection of Meriones unguiculatus have been analyzed by thin layer chromatography. Neutral and acid glycosphingolipids accounted for 95% and 5% of total glycosphingolipids, respectively. 12 different fractions were observed in the neutral glycosphingolipids extracts of the parasite. The most important was a monohexosylceramide fraction accounting for 56.4% of neutral glycosphingolipids. 9 different fractions were detected in gangliosides (acid glycosphingolipids). The fact that these glycosphingolipids were specific to the parasite was established by the analysis of different cell populations of the host. Glycosphingolipids were purified from control and parasite-infected gerbil blood cells as well as from peritoneal exudate cells of healthy gerbils after a non-specific immunostimulation. The chromatograms obtained with these extracts were totally different from the parasite. In addition, parasitosis was found to have no effect on the host blood cell glycosphingolipids.

Concentrations of free amino acids were measured concurrently in plasma, erythrocytes, granulocytes, and lymphocytes in umbilical cord blood, neonates, children, and adults. In each age group, the patterns of free amino acids were fairly similar in plasma and erythrocytes except for aspartic acid which was more abundant in erythrocytes. Of the amino acids in granulocytes, 71-77% was taurine; in lymphocytes taurine, aspartic acid, and glutamic acid comprised 35-44%, 18-24%, and 20-28%, respectively, of the total in all age groups. Leukocytes may contribute to the interorgan transport of amino acids to about 10% of the erythrocytes' contribution. Postnatally, the levels of glutamic acid and tyrosine in plasma; threonine plus glutamine, serine plus asparagine, and tyrosine in erythrocytes; histidine in granulocytes; and glutamic acid in lymphocytes were significantly increased (p less than 0.001); while the levels of phenylalanine and lysine in plasma; taurine in erythrocytes; valine and phenylalanine in granulocytes; and threonine plus glutamine, tyrosine, and phenylalanine in lymphocytes were significantly decreased (p less than 0.001). After the neonatal period concentrations of taurine and aspartic acid in erythrocytes, taurine and valine in granulocytes, and tyrosine and phenylalanine in lymphocytes increased gradually with age; while concentrations aspartic acid in plasma, histidine in granulocytes, and glycine in lymphocytes decreased gradually with age. The levels of glycine and valine in plasma, alanine and valine in erythrocytes, serine plus asparagine, glycine, alanine, and tyrosine in granulocytes, and aspartic acid, serine plus asparagine, and alanine in lymphocytes remained constant in all age groups.(ABSTRACT TRUNCATED AT 250 WORDS)

An anion-exchange high-performance liquid chromatography method has been used to quantitate the intracellular purine and pyrimidine nucleotides in extracts of pure lymphocytes, monocytes, neutrophils, eosinophils, erythrocytes, and platelets isolated from the blood of healthy human donors. For accurate and reproducible measurements of the nucleotide profiles in different types of pure leukocytes, the cell suspensions have to be free of platelets and erythrocytes. Incubation of the purified leukocytes for 1 h at 0 degrees C did not alter the nucleotide concentrations but reduced the interdonor variation to 10%. Incubation of purified lymphocytes for 1 h at 37 degrees C caused considerable changes in the relative concentrations of the adenine, guanine, uracil, and cytosine nucleotides. During this incubation the cell viability, the cell number, and the ATP:ADP ratio decreased. Incubation of monocytes and granulocytes for 1 h at 37 degrees C caused considerable loss of cells and/or cell death. For erythrocytes and platelets reproducible nucleotide concentrations were obtained after extraction of freshly isolated cells. During storage of erythrocytes, both at 0 degrees C and at 37 degrees C, a decrease in the ATP:ADP ratio was detected. In all cell types the predominant nucleotides were purine nucleotides, especially adenosine triphosphate. The relative concentrations of the adenine, guanine, uracil, and cytosine nucleotides were very reproducible per cell type and appeared to be characteristic for each cell type. The total nucleotide content was nearly the same for all cell types except erythrocytes, when expressed per microgram of protein. The described methods for purification and storage of blood cells will be useful for comparison of blood cells from healthy donors with those of patients, for example, leukemia patients, in which deviations of the purine and pyrimidine metabolic enzymes have already been described.

This article attempts to introduce the concept of cellular affinity chromatography as it departs from molecular affinity chromatography. Special emphasis is placed on the selectio of a solid support as well as on the role of lectins as affinity ligands. Our major goal was to bring to light the basic principles involved, multiple options of ligands and matrices, and different techniques, which may be applied to separate the complex cell population as well as cell membranes. It is hoped that further developments in the field, especially in the selection of proper experimental conditions, ligands and matrix material may provide better results. We have tried to identify some of the potential problems which should be considered before these approaches can be used on a routine basis. Although the review deals primarily with the affinity chromatography of cells and cell membranes, examples are presented for diverse systems such as cell organelles, viruses and phages. A table summarizing the use of cellular affinity chromatography is included. It lists more than 80 examples covering the literature up to December, 1979.

Blood cells of the colonial tunicate Botryllus were separated by density gradient centrifugation in Percoll. Unseparated blood cells were used to immunize mice for development of hybridoma cell lines producing anti-Botryllus monoclonal antibodies. These antibodies identify specific subpopulations of blood cells, indicating possible divisions of these cells into defined subgroups sharing particular differentiation antigens. Additional studies utilizing fluorescein- conjugated lectins also revealed differential binding to density- and monoclonal antibody-defined blood cell fractions. These methods allow separation of the different Botryllus blood cell types for functional studies.

The cholera toxin-colloidal gold-labeled IgG-F(ab')2 anticholera toxin ultrastructural immunocytochemical procedure was used for the localization of GM1 monosialoganglioside on the surface of human blood cells. The number of gold particles per micron of cell surface was counted and the data subjected to statistical analysis. Cholera toxin (CT) binding characteristics assessed in several subjects showed consistent labeling patterns for the various hemic cells, although some quantitative differences were noted in surface labeling densities between subjects. Neutrophils were invariably the most heavily labeled of the hemic cells, while lymphocytes, erythrocytes, and platelets exhibited only limited CT labeling. Exposure of hemic cells to neuraminidase induced a major increase in surface CT labeling that proved to be directly related to cell type and differed in many respects with the CT labeling pattern noted in nonenzyme treated cells. Newly exposed CT binding sites attributed to "masked" GM1 and/or to neuraminidase-transformed GD1a or GT1 gangliosides, showed that the number of new binding sites were nearly twice as abundant on platelet and monocyte sufaces as on the surfaces of neutrophil, lymphocyte, and erythrocyte populations. However, ratios of new CT binding sites to those normally available for CT binding were approximately 10:1 for erythrocytes, approximately 3--7:1 for lymphocytes, monocytes, and platelets, and approximately 1:1 for the neutrophil group. Exogenous GM1 was incorporated into the cell surface of the hemic cells in a differential manner. Platelets showed a dramatic increase in surface CT labeling, viz. approximately 12- to 20-fold, compared to that of other hemic cells; however, neutrophil and erythrocyte GM1 uptake was limited. Our studies have demonstrated that distinct differences exist in the extent of surface CT labeling of the various types of blood cells. They further indicated that the ability of the cell surface to incorporate exogenous GM1 may represent a differential expression of the physiochemical properties of the surface of the individual cell types.

The method of impulse cytophotometry was applied for determination of DNA content of blood cells in patients with various types of leukaemia. It was found that at the time of improvement in acute leukaemia the proportion of cells in the non-proliferative stage increases. In chronic leukaemias the improvement was associated with shifts in reverse direction.

We have outlined a modified method of DNA extraction from blood and bone marrow that can be used for gene rearrangement studies of leukemia and lymphoma and can consistently give excellent yields of DNA from as little as 1 x 10(6) cells. The method is based on the use of small sample vessels, a marked reduction in solution volume and minimal handling of the specimen. By using known cell numbers from eight leukemic bone marrow samples and four leukemic cell lines, the modified extraction procedure consistently had better yields of DNA, compared with a standard procedure. These findings were confirmed by further comparisons of DNA yields from 14 hypocellular clinical specimens that had been divided into two equal parts for side-by-side DNA extraction with both methods. The quality of the DNA from the modified extraction method was similar to standard methods on subsequent Southern blot analysis and hybridization with immunoglobulin and/or T-cell receptor gene probes. This modified DNA extraction method is a procedure that complements standard extraction methods and expands the number and type of specimens that can be evaluated by the clinical molecular genetics laboratory.

The backscattered electron imaging (BEI) mode of scanning electron microscopy (SEM) has been applied to study various histo-cytochemical reactions in biological specimens since the early seventies. Due to numerous, recent technical improvements the BEI mode of SEM now belongs to the routine of many SEM laboratories. For cytochemistry, BEI has been mainly used to: visualize intracellular structures and organelles; recognize the different cell types in heterogeneous populations or tissues; study the correlations between enzymatic activities and cell surface features. We have evaluated the most relevant results obtained in the study of blood cells and the possible future applications of these techniques.

Commerically available Romanowsky blood stains are variable mixtures of thiazein dyes and brominated fluorescein derivatives with varying degrees of metallic salt contamination in a number of different solvent systems. There is a need for standardized Romanowsky stains of constant composition, which, when used in conjunction with a carefully controlled specimen preparation technique, should give consistent performance. Such a preparation system would be of great value to hematologists in general and would be essential to the validity of data obtained by the digital processing of blood cell images. It is possible to prepare standardized Romanowsky stains as mixtures of two or three dye components, namely, eosin Y, azure B and methylene blue, although azure B has only recently become commercially available at an acceptable degree of purity. The logistic problems of stain standardization are discussed.

A series of fluorescein-conjugated lectins (Sophora japonica agglutinin, Helix pomatia agglutinin, peanut agglutinin, the erythroagglutinin from Phaseolus vulgaris, pokeweed mitogen, wheat germ agglutinin and the fucose-binding lectin from Lotus tetragonolobus) were analyzed for their binding to human peripheral blood cells with a fluorescence-activated cell sorter. Most of the lectins showed increasing cell binding in the order erythrocytes less than lymphocytes less than monocytes less than neutrophils and the degree of fluorescence was not related to the major blood groups. The fucose-binding lectin (FBP) was the main exception in that it appeared to bind only to blood neutrophils. From an analysis of the binding of this lectin to human bone marrow cells it was found that the degree of binding within the granulocytic series increased with progressive differentiation. Marrow monocytes and nucleated erythroid cells bound to FBP in contrast to monocytes and non-nucleated red cells in the peripheral blood which showed negligible binding. Lymphocytes both in the marrow and blood displayed negligible binding of FBP. These properties allowed an enrichment of haemopoietic progenitor (colony-forming) cells from human marrow cell suspensions and a depletion of colony-inhibiting cells, when these were present, by selection of cells with the appropriate fluorescence intensity.

The relationship between blood cells and plasma concentrations of cyclosporin A (Cy A) determined by radioimmunoassay, was investigated in 12 heart and 12 kidney transplant recipients. The decision between a linear and nonlinear model was made according to a standardized residuals plot. We observed high blood cells-plasma concentration ratios in the two groups, indicating a high affinity of Cy A for blood cells. The distribution of Cy A between blood cells and plasma was ascribed to a nonlinear saturable model in the two groups. According to our results we have simulated the blood-plasma concentration ratio of Cy A as a function of plasma Cy A concentration and hematocrit.

1. A study was undertaken to investigate the distribution of ascorbic acid between various cellular components of blood, in normal individuals, and its relation to the plasma concentration. Forty-one unsupplemented individuals and sixteen supplemented (2 g/d for 5 d) individuals were studied. 2. Granulocytes, mononuclear leucocytes, platelets and erythrocytes were separated by differential sedimentation and centrifugation. Ascorbic acid contents were measured by the dinitrophenylhydrazine method. 3. Ascorbic acid content per cell was higher in mononuclear leucocytes and granulocytes than in platelets and erythrocytes. Intracellular ascorbic acid concentrations, calculated from published values for cell volumes, when compared with the plasma concentration showed a marked ability to concentrate ascorbic acid in mononuclear leucocytes (80 times), platelets (40 times) and granulocytes (25 times). 4. Erythrocytes showed little ability to concentrate ascorbic acid over the normal range of plasma concentration but because of their relative numbers they and the plasma fraction accounted for most of the blood-borne ascorbic acid (greater than 70%). 5. The ascorbic acid content of granulocytes, platelets and erythrocytes showed a significant positive correlation with the plasma concentration and supplementation with ascorbic acid significantly increased the content of these cell types. Mononuclear leucocytes in contrast did not show any such relationship. 6. The ability of the mononuclear leucocytes to maintain the highest levels of ascorbic acid in the cell types studied, despite variation in plasma availability, warrants further study, particularly in view of the importance of these cells in immunocompetence.

Histochemical techniques have demonstrated an increase in the content of some physiologically active substances (catecholamines, serotonin and histamine) and acetylcholinesterase activation in the formed elements of the blood plasma of patients with acute pneumonia. The development of acute experimental pneumonia and pulmonary edema in rabbits has been shown to be accompanied by inhibition of serotonin inactivation in the lungs. It may be assumed that the increase in the blood content of physiologically active substances is caused by abnormality of the non-respiratory functions of the lungs in acute pneumonia.

The present paper refers to statistically ensured normal values of cytochemical findings in leukocytes of the peripheral blood, of the leukocytes concentrate and the bone-marrow which were obtained in determining the activity of peroxydase, alkaline and acid phosphatase as well as the content of polysaccharides and lipids in 814 healthy persons of various age and sex. The data were calculated according to the formulae of Astaldi and Verga [2].

X-ray histospectrographic analysis at the scanning and transmission electron microscope (STEM) are made on the blood cells of Phallusia mamillata Cuvier and Ciona intestinalis, to study the 'direct' intracellular sites of accumulation of vanadium. The results show a clear accumulation of vanadium on the membrane and in the granules of vacuoles of amebocytes, signet ring cell, compartment cell and traces of metal in the 'vanadophores' of vanadocytes.

Cellular lectins (CLs) of Phallusia mamillata were demonstrated in protein preparations obtained by salt fractionation from hemocytes sonicated in a suitable medium. Since the lectins from the precipitated fraction bind sugars containing D-galactosyl groups, they were purified by affinity chromatography on Sepharose. SDS-PAGE under reducing conditions showed that CLs are formed of two components of apparent MWs approximately 36,900 and 35,090 and thus differ from serum lectins (SLs) (MW about 62,200). The "shrinkage" observed when SLs were examined under nonreducing conditions suggest the presence of intrachain disulphide bonds which can affect the molecular structure of the SLs. CL-SL differences were also revealed by the nonidentity reaction of the immuno-precipitate in immunodiffusion using an anti-SL immune serum. The capacity of hemocytes to form rosettes or clumps with erythrocytes demonstrated that they possess alpha-lactose specific CLs on their surfaces.

The numbers of glucocorticoid receptors in human alveolar macrophages and peripheral blood cells were measured with 3H-prednisolone. Alveolar macrophages, which constituted 89.0 +/- 5.9% of broncho-alveolar cells, obtained by broncho-alveolar lavage from normal volunteers had much larger numbers of specific glucocorticoid receptors than peripheral blood cells. The numbers of glucocorticoid receptors in peripheral polymorphonuclear leucocytes, lymphocytes and lymphocyte subpopulations (B cells, T cells, TG cells and TnonG cells) were nearly equal. In patients with idiopathic pulmonary fibrosis, in whom alveolar macrophages amounted to over 85% of the broncho-alveolar cells, the number of glucocorticoid receptors in alveolar macrophages was significantly decreased, but the numbers in their peripheral blood cells were normal. This finding suggests that the number of glucocorticoid receptors in alveolar macrophages may change specifically during disorders of the lung.

A novel compound, for which the trivial name tunichrome is proposed, was isolated from the vanadium-rich blood cells of the tunicate Ascidia niga. Preliminary structural studies suggest a molecular weight of about 390, the presence of conjugated vinyl groups, and an acidic group, possibly carboxyl, with an apparent pKa of 3.0. Elements C, H, N and O comprise 98.4% of the sample weight, the number of atoms per mol of tunichrome being 14.1, 22.2, 1.5 and 10.6 respectively, which indicates some heterogeneity in the sample. Tunichrome readily reduces Fe(III) and V(V). In an initial fast step, 2 mol of V(V) are reduced, or 4 mol of Fe(III)-phenanthroline per mol of tunichrome; in a further slow reaction, another 9 mol of Fe(III)-phenanthroline or Fe(III)-bipyridine are reduced. The initial reaction is first-order with respect to tunichrome and Fe(III). Above pH 3.5, tunichrome is rapidly hydrolysed, 13 mol of OH- being consumed per mol of tunichrome. The hydrolysis involves polymerization and loss of the characteristic absorption peak at 325 nm. It is suggested that the presence of tunichrome may be linked to vanadium accumulation by the blood cells. The mechanism involves entry of vanadate via an anionic channel into vacuoles of the blood cells, where it is reduced to V(IV) or V(III), both of which, being cationic, cannot escape from the vacuole.

The mass fraction of calcium (Ca), magnesium (Mg), manganese (Mn), iron (Fe). zinc (Zn), strontium (Sr) and copper (Cu) in isolated granulocytes, erythrocytes and platelets was measured using the nuclear microprobe technique. The cellular profile of metal variation was conspiciously altered in patients (n = 29) with ankylosing spondylitis compared with the profile found in age and sex matched healthy controls. Ca was accumulated in erythrocytes and granulocytes but decreased in platelets (p less than 0.001). Mg was increased in granulocytes but appeared in reduced concentrations in platelets and erythrocytes (p less than 0.001). The cellular amounts of Mn was increased in granulocytes (p less than 0.001), normal in platelets (p greater than 0.05) and subnormal in erythrocytes (p less than 0.001). Zn was reduced in all 3 cell types (p less than 0.001). Fe accumulation was evident in granulocytes and platelets (p less than 0.001). Sr was only measurable in granulocytes from patients. Cu was below the detection limit in the different cell types isolated from patients, but appeared in measurable amounts in erythrocytes and platelets from controls. The granulocyte amounts of Ca, Mg, Mn and Sr were strongly related to the acute phase reaction. Negative correlations were found between erythrocyte Mg and Zn and the inflammatory activity. The patients had increased serum levels of Cu, normal levels of Ca, Mg and Sr and decreased levels of Zn, Fe and Mn. No relationship was found between the serum concentrations of these elements and their respective cellular stores, except for a weak negative correlation between granulocyte and serum Fe.(ABSTRACT TRUNCATED AT 250 WORDS)

The simultaneous determination of serum CA and their receptors in blood cells offers a possibility to evaluate disturbances of the DA and NA neuronal systems in man. High affinity binding sites for 3H-yohimbine in platelets, 3H-DHA on granulocytes and 3H-spiperone in lymphocytes from healthy control persons, unmedicated (n = 28), and medicated (n = 8) schizophrenics as well as from an unmedicated psychiatric control (n = 14) were investigated. Furthermore, the actual concentration of the circulating CA was determined with HPLC-ECD. In unmedicated schizophrenics as compared to controls, specific binding of 3H-spiperone to lymphocytes was markedly elevated in capacity and less in affinity as compared to controls. For beta2-receptors a significant decrease was found in capacity with no change in affinity. The changes in alpha2-receptors with a slight decrease in capacity were less distinct. The concentrations of circulating CA ranged from normal values up to a more than 3 fold increase in NA and DA, whereas A concentrations were practically unchanged. No overall change in these data was found in the medicated schizophrenic patients. 3H-spiperone binding was characteristically increased only in schizophrenics, but did not elevate above control data in the non-schizophrenic psychiatric control group. Preliminary experiments of family studies suggest that this model could be valuable as a vulnerability marker.

An antigen immunochemically indistinguishable from plasma alpha 2-antiplasmin, the primary plasmin inhibitor, was detected in human platelets. By radioimmunoassay, 33-114 ng alpha 2-antiplasmin antigen was quantitated in the detergent-soluble extract of 10(9) washed human platelets from 10 normal donors with a mean level of 62 +/- 24 ng/10(9) platelets. Plasma alpha 2-antiplasmin, either in the platelet suspending medium or on the surface of the platelets, could account for less than 8% of the antigen present in the platelet extracts. When stimulated with thrombin, the platelets released alpha 2-antiplasmin antigen without cell lysis, and greater than 85% of the alpha 2-antiplasmin antigen was released at a high thrombin dose. At a lower dose of thrombin, alpha 2-antiplasmin and platelet factor 4 were partially released without concomitant secretion of serotonin. No alpha 2-antiplasmin antigen was detected in extracts or red blood cells, polymorphonuclear leukocytes, and adherent and nonadherent mononuclear cells. Thus, the platelet is the only peripheral blood cell containing significant amounts of alpha 2-antiplasmin.

The distribution of lipid, glycogen, peroxidase, alkaline phosphatase and acid phosphatase has been studied in the cells of blood and bone marrow smears from young chickens. Chicken heterophil granules react differently from those of mammalian neutrophils. A strongly positive peroxidase reaction was given by developing erythrocytes in chickens, unlike mammals. The significance of these species differences is not yet clear.

The main results obtained by the author and collaborators in 25 years' work about the type of sialic acid determined in some materials from the animal kingdom are summarized, with the respective references to the original papers and the indication of the reviews in which they have generally been quoted. The high number of publications on sialic acids from the time of their discovery reflects the increasing interest on these acids in areas such as their chemical composition (16 different sialic acids are now recognized), their distribution in Nature (a not yet well explained topic), their metabolism (which shows differences between bacteria and animal cells) and their biological roles, mainly as a part of glycoproteins and gangliosides (as receptors, markers of the cytoplasmic membrane, viscosity agents of the glycoproteins, life regulators of some substances, etc.).

It was our purpose to study the immunological activity of circulating fluid cells in the blattoid insect Leucophaea maderae L. These cells are generically called haemocytes; after a preliminary morphological study under optical microscope, they were treated with anti-heteroagglutinins serum marked with fluorescein isothiocyanate. Thus, it was possible to show the presence of heteroagglutinins into the cytoplasm, as regards one group of haemocytes, and on the cell membrane for the second group.

Prussian blue has been widely used to localize iron in a variety of tissues at the light and electron microscopic level. In the present study, thin sections of human marrow and blood cells and rat duodenal cells were exposed to silver proteinate (SP) after staining en bloc with acid ferrocyanide (AF), with and without prior iron saturation using iron nitrilotriacetate (FeNTA). Silver deposition was observed over Prussian blue-reactive sites and significantly enhanced sites of minimal AF and FeNTA-AF staining. AF-SP stain deposits were present in the cytoplasmic matrix, granules, and occasionally on the surfaces of macrophages, monocytes, and erythroblasts. FeNTA-AF-SP stained additional cytoplasmic and surface sites in erythroblasts and stained neutrophil granules intensely. Duodenal epithelium from iron-loaded rats demonstrated strong AF-SP staining of ferric iron in microvilli, apical cytoplasmic matrix, and lateral membranes. Similar preparations from iron-replete rats stained sparsely; however, intense AF-SP staining was observed after iron saturation with FeNTA. SP similarly enhanced luminal ferrous iron deposits stained with acid ferricyanide in rats given intraluminal ferrous iron. AF-SP stain deposits were removed by exposure of thin sections to NH4OH, KCN, or HNO3 but were not affected by prior exposure to HIO4 or NaBH4, consistent with a silver cyanide or complex stain precipitate rather than reduced silver or silver ferriferrocyanide. SP enhancement of Prussian blue allows identification of reactive sites not readily visualized with AF or FeNTA-AF alone, and offers the potential for differentiating AF staining from other deposits or organelles of comparable density.

A methodological problem with the normal phase high performance liquid chromatography (HPLC) of hydroxylated products of docosahexaenoic and arachidonic acids is described. Diacylglycerols present in lipid extracts of rat retina co-elute with monohydroxy derivatives of docosahexaenoic or arachidonic acid, when samples are applied to uPorosil columns and eluted with hexane/isopropanol/acetic acid. Analysis of fatty acid composition of diacylglycerols which were acetone-extracted from the incubation medium showed a profile similar to diacylglycerols extracted from the tissue by hexane/isopropanol, although acetone extraction resulted in extremely variable recovery of diacylglycerols. This co-elution of diacylglycerols with monohydroxy polyunsaturated fatty acids can lead to a significant error in estimation of lipoxygenation activity by conversion of radiolabeled precursors, because the incorporation of fatty acids into diacylglycerols is very active in many tissues. An alternative extraction method and reverse phase HPLC procedures that result in the complete separation of hydroxy fatty acids and diacylglycerols are described.