Human Blood Smears Research Articles

Enzyme activity of peripheral blood cells in experimental chronic myocarditis linked with persistence of Coxsackie a virus

Human and animal blood smear staining with PAPh has revealed mononuclear leukocyte-erythrocyte aggregates. Administration of retinoic acid increased concentration and dimensions of these aggregates and was followed by preferential accumulation of PAPh-negative osmotically unstable erythrocytes. Similar changes were detected in the blood samples of women engaged in the production of retinoic acid. Aggregate concentration showed positive correlation with erythrocyte sedimentation rate and no correlation with prothrombin time.

Enzyme-histochemical Differentiation of Neuroglia and Micro ia: A Contribution to the Cytogenesis of Microglia and Globoid Cells: Review of the Literature

The classification of microglia is uncertain. An ectodermal origin and similarity with neuroglial cells as well as a mesodermal origin and similarity with blood monocytes are discussed. The classification of globoid cells in globoid cell leukodystrophy is unclear as well.The literature dealing with the enzymatic activity of neuroglial cells, microglial cells, globoid cells, and blood monocytes was studied and reviewed. Some of the author's observations concerning enzymatic activity in human brain tissue acquired during autopsy, human blood smears, and animal brain tissue are integrated; in addition to the oxidoreductases, several other. enzymes were studied. The following cell types were considered: oligodendrocytes, astrocytes, microglial cells, globoid cells, and blood monocytes. Two functional stages could be distinguished in the nonneuronal brain cells: the resting stage and the reactive.stage.The following relationships could be established: The enzyme patterns vary considerably for resting microglia and neuroglial cells; for resting microglia and reactive microglia, globoid cells as well as blood monocytes. The resting microglia are striking because enzymes which are present in varying amounts in the other cells mentioned (i. e., oxidoreductases, butyryl cholincstrase, phosphorylase, acid phosphatase, peroxidase, alpha-naphthyl acetate esterase, 5-nucleotidase) are absent in resting microglia. Nucleotide phosphatases however were demonstrated in resting microglia in an intensity observed only in reactive microglia, globoid cells, and blood monocytes.The findings are only hypothetically applicable to the problem of the classification of cell types because differing enzyme patterns are not necessarily indicative of different classifications. On the other hand, however, the observation of the identity of enzyme patterns in reactive microglia , globoid cells and blood monocytes agrees with observations made with other methods (particularly immunohistologic and autoradiographic methods). It therefore may be concluded that the identical enzyme-histochemical findings of reactive microglia, globoid cells, and blood monocytes, tends to indicate identical origin for these cells; it however does not contradict their clzissification in the mononuclear phagocyte system.

Human lymphocyte subpopulations identified by bacterial adherence are functionally different

Previously, two B (B 1 and B 2)- and four T (T 1, T 2, T 3, T 4)-lymphocyte subpopulations have been identified in human blood smears by bacterial adherence. Here, to study the functional differences between these subpopulations the T 1T 2 cells were separated from T 3T 4 cells by selective adherence to Escherichia coli-2 4 4 This strain of E. coli was deposited in the ARS Culture Collection under No. B-11010, at Northern Regional Research Center, U.S. Department of Agriculture, Peoria, Ill. 61602. monolayers. The adherent cells (T 1T 2 cells) responded well to concanavalin A in 3-day cultures and in mixed lymphocyte culture (MLC) in 6-day cultures and developed into cells specifically cytotoxic for allogeneic lymphocytes. The nonadherent cells (T 3T 4 cells) cultured for the same length of time were poorly responsive to concanavalin A, variably responsive in MLC, and poorly active in specific cytotoxicity. The T 3T 4 cells were naturally cytotoxic for allogeneic lymphocytes and for a normal lymphoblastoid cell line. We concluded that the T cells that bind E. coli-2 (T 1T 2 cells) are functionally different from those that do not bind (T 3T 4 cells).

A Measurement Technique for Cell Adhesiveness

Abstract Determination of the percentage of cells in clumps on a stained smear of human peripheral blood porvided a useful, accurate technique for measuring cell adhesiveness. Smears of human peripheral blood drawn with EDTA were prepared on a blood slide centrifuge, stained, and examined under a light microscope. Statistical analysis showed that the method resulted in a Poisson distribution of particles on the slide, where a particle was considered to be a simple cell, or two or more cells which appeared to be touching, Analysis of the distributions of erythrocytes and leukocytes showed that clumps were formed before the cells were deposited on the slide. When adhesiveness of erythrocytes or leukocytes was increased by incubation with antiserum to the corresponding cell type, the percentage of that cell type in clumps increased proportionately, Preliminary results using the method showed that normal human donors had similar to 1% of their erythrocytes and 1- 5% of their leukocytes in clumps. In chronic myelocytic leukemia, as many as 60% of the leukocytes were in clumps.

A measurement technique for cell adhesiveness

Determination of the percentage of cells in clumps on a stained smear of human peripheral blood porvided a useful, accurate technique for measuring cell adhesiveness. Smears of human peripheral blood drawn with EDTA were prepared on a blood slide centrifuge, stained, and examined under a light microscope. Statistical analysis showed that the method resulted in a Poisson distribution of particles on the slide, where a particle was considered to be a simple cell, or two or more cells which appeared to be touching, Analysis of the distributions of erythrocytes and leukocytes showed that clumps were formed before the cells were deposited on the slide. When adhesiveness of erythrocytes or leukocytes was increased by incubation with antiserum to the corresponding cell type, the percentage of that cell type in clumps increased proportionately, Preliminary results using the method showed that normal human donors had similar to 1% of their erythrocytes and 1-5% of their leukocytes in clumps. In chronic myelocytic leukemia, as many as 60% of the leukocytes were in clumps.

A modified technique of preparing mouse-liver nuclei for the detection of antinuclear antibodies (ANA) by immunofluorescence

Smears of homogenized mouse liver were used for the detection of antinuclear antibodies (ANA) by immunofluorescence test. Using this technique sixty serum samples from human patients suffering from systematic lupus erythematosus (SLE), rheumatoid arthritis (RA) and various autoimmune and collagen diseases and sixty normal human sera were studied. The results were comparable with those obtained using smears of human peripheral blood as source of nuclei, but with the latter material the incidence of ANA in normal sera was higher. Moreover, by using smears of mouse liver several kinds of antinuclear antibodies can be distinguished. Results obtained with the modification proposed were compared with data obtained by using the conventional method (cryostat section). The modified technique has two advantages: 1) the nuclear figures and morphological details can be better observed; 2) it is easier to perform.

An Elution Procedure for Visualization of Adult Hemoglobins in Human Blood Smears

Abstract A procedure is described for visualization of normal and mutant adult hemoglobins in human blood smears. After extraction of blood smears with a concentrated potassium phosphate buffer (2.76 M, pH 7.2), erythrocytes that had adult hemoglobins stained bright red with erythrosin, whereas cells that had only fetal hemoglobin appeared as clear ghosts. Analyses of cord blood from newborn infants indicate that, although most erythrocytes contain only Hb F and a few contain only Hb A, many contain both hemoglobins A and F.

Evaluation of glucose-6-phosphate dehydrogenase in single erythrocytes in human blood smears.

A cytochemical method has been described for the specific and direct staining of glucose-6-phosphate dehydrogenase (G-6-PD) in red blood cells (RBC), by using the tetrazolium salt Nitro-BT (NBT). This method allows a semiquantitative estimation of G-6-PD activity in single cells. The results of such estimations were found to be correlated with spectrophotometric determination of G-6-PD activity in RBC lysates. The activity of G-6-PD was shown to decline with aging of the cell. In cases of severe G-6-PD deficiency, where the spectrophotometric assay revealed no activity, staining of ‘young’ RBC showed appreciable activity, while the ‘old’ cells showed no activity.

Analysis and diagnostic significance of leucine aminopeptidase activity in human blood and bone marrow smears

Analysis and diagnostic significance of leucine aminopeptidase activity in human blood and bone marrow smears

Scanning Microscopy of Human Blood and Marrow Smears

The scanning electron microscope (SEM) permits direct observation of standard light microscope slide and cover slip preparations. Single cells can be located and identified in the two instruments and the higher resolution, three dimensional image of the SEM can be correlated with the classic cytological information contained in the light microscope image.Samples of peripheral blood and bone marrow were obtained from patients with chronic lymphatic leukemia and polycythemia. A drop of either blood or bone marrow from the syringe used for drawing the specimen was placed on alcohol cleaned, 22 mm square cover glasses. A second cover glass was placed over the drop of blood or marrow and the two cover glasses drawn apart in the manner usual for preparing blood and marrow for examination by light microscopy. These cover slips were rapidly air dried and then stained for examination by light microscopy. The staining procedure consisted of flooding the cover glass with Jenner's stain for 5 minutes, adding distilled water to cover the glass for another 5 minutes and then washing in tap water. Following this a Giemsa stain made up freshly from the stock solution was applied to the cover glass for 10 minutes. The cover glass was then washed and air dried. A thin coating of platinum-paladium was applied in a vacuum evaporator to facilitate electrical conduction.

Antinuclear factors and human leucocytes: reaction with granulocytes and lymphocytes.

SummaryWe compared the reaction of antinuclear factor (ANF) with nuclei of lymphocytes, granulocytes and liver cells by retesting III sera known to have given a positive test for antinuclear factor with human blood smears. These sera could be placed in three groups, as follows: Group 1, 47 sera, reacted to equal titre with the nuclei of lymphocytes, granulocytes and rat liver nuclei; Group 2, 25 sera, reacted to much higher titre with the nuclei of granulocytes than with nuclei of lymphocytes and liver cells; Group 3, 39 sera, reacted only with the nuclei of granulocytes. Thus, reactive sera contained separate antinuclear factors for nuclei of lymphocytes and liver cells, and for granulocytes.The presence of high titre lymphocyte‐reactive ANF characterized systemic lupus erythematosus, and was associated with lymphopenia (but not granulocytopenia) and a positive result to the LE cell test. Granulocyte‐specific ANF occurred in 4% of healthy subjects, but only to low titre (10 or less) ; high titres occurred in a range of diseases considered to be “ autoimmune ”, but had little diagnostic specificity and were not associated with cytopenias.The fixed blood film has some advantages for ANF tests: it is inexpensive and readily available, the incidence and titre of reactions with lymphocyte nuclei and tissue nuclei are similar, and the granulocyte‐specific ANF can be recognized. However, it should be specified whether positive reactions are with all nuclei or only granulocyte nuclei.

Elution Procedure for the Demonstration of Methæmoglobin in Red Cells of Human Blood Smears

THE peroxidatic activity of the red blood pigment can be inhibited by cyanide in the case of methaemoglobin (HbIII), but not in the case of oxyhaemoglobin (HbII) (ref. 1). It seemed promising to use this difference for a differential staining procedure of HbII- and HbIII-containing red cells by first adding cyanide to the heparinized or citrated blood in question and by staining smears prepared from it by a peroxidase reaction.

STANDARDIZED HEAT FIXATION FOR BLOOD SMEARS.

Because a technique was needed for good fixation of blood smears without the use of solvents such as methanol, the effect of heat fixation on human and rat blood smears was studied. Slides were placed in a fused silica dish, covered by a Pyrex petri dish lid, and heated for 5 min in an oven at 150° C. After such fixation, smears stained by Giemsa' s solution showed that the blood elements retained their characteristic morphology. Thus, even though volatile substances may be lost and heat-labile constituents may be modified, this simple method avoids contact of the cells with solvents.

A Possible Explanation of the Formation of Long Platelets from Pulmonary Megakaryocytes

THE purpose of this communication is to suggest a possible explanation of the formation of the elongated sausage-shaped, sometimes bizarre long platelets that are occasionally encountered in human blood smears. Since Bizzozero1 first described the platelets as a distinct entity in 1882, much has been conjectured as to the source and mechanism of their formation. Before Wright2 described the formation of blood platelets from bone marrow megakaryocytes in the marrow sinusoids in 1910, the thrombocytes were assumed to be formed from disintegration of erythrocytes and leucocytes. Howell and Donahue3 unsuccessfully attempted to show by lung per fusion studies in 1937 that the thrombocytes were formed from megakaryocytes. The latter they believed originated in the lungs. More recently, Humphrey4 has added chemical evidence to the hitherto purely morphological evidence that the platelets are derived from the megakaryocytes. But according to Wintrobe5: “The mechanism of production of platelets from the cytoplasm of megakaryocytes is by no means clear.”

Glycogen in basophilic leucocytes in human blood smears.

In a recent study of the occurrence of the periodic acid-Schiff reaction in various normal cells of the blood and connective tissue by Wislocki, Rheingold and Dempsey,1 small clear red dots were observed in the pale pink cytoplasm of the basophilic leucocytes. These investigators were not able to identify the basophiles in control blood smears digested by saliva. They therefore concluded that the red stained material in the undigested smears may have been glycogen but that the “apparent finding needs further verification.” Similarly, Gibb, and Stowell2 decided that “Because myeloid cells which were free from glycogen were not observed in normal peripheral blood and bone marrow films, basophils, although not specifically identified, may also contain glycogen.”To make certain that basophils could be identified in smears digested by saliva before treatment with periodic acid and leucofuchsin, dried smears of human blood were stained in Wright's blood stain in absolute methyl alcohol for 3 minutes. This step ...

Staining of the Processes (Flagella) of Human Erythrocytes

As noted by Kite 1 and Oliver,2 when the red blood cells of various vertebrates, from frogs and fishes to man, are examined in sterile Ringer's solution to which hirudin has been added, certain of the cells are found to possess various types of motile and nonmotile processes. These processes may be seen in petrolatum-sealed preparations with the dark field, as well as on blood cells mounted in a Barber moist chamber freely open to the air, and examined under the oil immersion by ordinary transmitted light, without the aid of special condensers. Within from forty to fifty minutes after the preparation is made some of the erythrocytes are seen to possess one or more processes, varying in length from 2 to 3 microns to as much as 30 microns. Some of these processes, which may appear on crenated as well as on uncrenated cells, exhibit a rapid whiplike motion; others show a slow undulatory motion, while still others are absolutely motionless. The processes are definitely produced by the red cell, possess unquestioned continuity with it, and, under certain conditions, are capable of extremely rapid retraction back into the cell. Erythrocytes with whipping or slowly undulating processes are actually motile, free-swimming cells, and I have followed such motile red cells through as many as fifty oil immersion fields. Not only are these processes occasionally retracted, but much more frequently they break off from the cell, and the motile processes continue to whip or undulate even after being detached. In fresh blood preparations such processes, broken off from red blood corpuscles, may rather easily be mistaken for spirochetes, leptospira and similar organisms. Some of the processes, after whipping for twenty or thirty minutes, may be seen to take on a granular, beaded appearance; sometimes only a single, terminal bead is found near the end of a process. So far as I am aware, the processes have been observed on erythrocytes only in fresh preparations of blood. The purpose of this note is to record their successful staining in fixed smears of normal human blood.

Variations in Monocytic Response to Peroxydase Test.

Naegeli states that almost all, if not all, of the human monocytes are peroxydase reacting. The author found no peroxydase-reacting monocytes in the blood of a rabbit. In a normal adult man there were present 3.4% peroxydase mononuclear phagocytes and 1.1% phagocytes that did not react to the peroxydase test. The conclusion was that the peroxydase cells of human blood are of myeloid origin, which is the view held by Naegeli. The negative phagocytes were thought to be the true monocytes derived from the reticular portion of lymphoid tissue. Recently some unpublished observations have brought me to a consideration of the alternative view, namely, that the peroxydase-positive phagocyte gets its granulation secondarily in a more or less accidental manner. The conclusions arrived at in this paper are based on tissue cultures of blood leukocytes. Methods and Results: Numerous cultures of human blood were made by the coverglass method and examined by the peroxydase test daily from one day to 4 weeks. This was done on the assumption that the peroxydase granulation of the monocytes would disappear, provided the granulation was acquired secondarily and was not an essential part of the cell cytoplasm. After one month in the incubator large peroxydase reacting phagocytes were present in the cultures. Never was the number of phagocytes great and no conclusion was possible in regard to the proportion of positive and negative cells. Rhoads and Parker found at the end of 2 weeks of incubation both positive and negative mononuclear phagocytes in normal human blood. The next procedure was to culture rabbit leukocytes in contact with variable amounts of human blood: Rabbit blood was cultured on sterile cover-glasses covered with smears of human blood made with aseptic precautions. The smears were kept dry in the air for 2 hours. The rabbit blood was obtained from the heart and drops of blood placed directly on the covers from the small caliber needle. The covers which had previously been rimmed with sterile vaseline were inverted over the well and placed in the incubator within 5 minutes after withdrawal of the blood from the heart. After about one hour the covers were sealed permanently with paraffin. At the end of 2 weeks as many as 500 large mononucleated cells were present in some of the cultures while in the human blood cultures mentioned above the cells at the most numbered a few dozen.