Investigation of various structures of DN A molecules (Ⅲ)—Coil-globe transition of λ-DNA induced by cationic surfactant

The structure transition of λ-DNA induced by cationic surfactant cellar media was investigated by using CD, SEM and AFM. The experimental data of CD revealed that λ-DNA can be induced from B-form to a collapsed structure with the addition of the cationic surfactant CTAB to the system. The condensed process of λ-DNA from coil state to small globular state (diameter about 1.25 μm) and finally big globular state (diameter about 5.4 μm) was observed by using SEM and AFM. Project supported by the Ninth Five-Year Great and Important Project of the Chinese Academy of Sciences and National Postdoctoral Science Foundation of China.     

Two X family DNA polymerases, λ and μ, in meiotic tissues of the basidiomycete, Coprinus cinereus

The X family DNA polymerases λ (CcPolλ) and μ (CcPolμ) were shown to be expressed during meiotic prophase in the basidiomycete, Coprinus cinereus. These two polymerases are the only members of the X family in the C. cinereus genome. The open reading frame of CcPolλ encoded a predicted product of 800 amino acid residues and that of CcPolμ of 621 amino acid residues. Both CcPolλ and CcPolμ required Mn²⁺ ions for activity, and both were strongly inhibited by dideoxythymidine triphosphate. Unlike their mammalian counterparts, CcPolλ and CcPolμ had no terminal deoxynucleotidyl transferase activity. Immunostaining analysis revealed that CcPolλ was present at meiotic prophase nuclei in zygotene and pachytene cells, which is the period when homologous chromosomes pair and recombine. CcPolμ was present in a slightly wider range of cell stages, zygotene to diplotene. In analyses using D-loop recombination intermediate substrates, we found that both CcPolλ and CcPolμ could promote primer extension of an invading strand in a D-loop structure. Moreover, both polymerases could fully extend the primer in the D-loop substrate, suggesting that D-loop extension is an activity intrinsic to CcPolλ and CcPolμ. Based on these data, we discuss the possible roles of these polymerases in meiosis.     

Optimizing the crystal size and habit of β-sitosterol in suspension

The aim of this work was to survey how processing parameters affect the crystal growth of β-sitosterol in suspension. The process variables studied were the cooling temperature, stirring time and stirring rate during recrystallization. In addition, we investigated the effect a commonly used surfactant, polysorbate 80, has on crystal size distribution and the polymorphic form. This study describes the optimization of the crystallization process, with the object of preparing crystals as small as possible. Particle size distribution and habit were analyzed using optical microscopy, and the crystal structure was analyzed using X-ray diffractometry. The cooling temperature had a remarkable influence on the crystal size. Crystals with a median crystal length of ≈23 μm were achieved with a low cooling temperature (<10°C); however, a fairly large number of crystals over 50 μm appeared. Higher cooling temperatures (>30°C) caused notable crystal growth both in length and width. Rapid (250 rpm), continuous stirring until the suspensions had cooled to room temperature created small, less than 50 μm long (median <20 μm), needle-shaped crystals. The addition of surfactant slightly reduced the size of the initially large crystals. Both hemihydrate and monohydrate crystal forms occurred throughout, regardless of the processing parameters. By using an optimized process, it was possible to obtain a microcrystalline suspension, with a smooth texture.     

Comparative pollen morphology of Turkish species of <Emphasis Type="Italic">Petrorhagia</Emphasis> (Caryophyllaceae) and its systematic implications

Pollen morphology of 10 taxa from Turkey, four of them endemic, belonging to the genus Petrorhagia (Caryophyllaceae), P. alpina subsp. alpina, P. alpina subsp. olympica, P. cretica, P. dubia, P. hispidula, P. lycica, P. pamphylica, P. peroninii, P. prolifera and P. saxifraga, has been investigated using light (LM) and scanning electron (SEM) microscopy. Pollen morphology differences among these taxa have been determined. The pollen type of investigated taxa is polyporate (12–22-porate), sphaeroidal, the exine exhibits a tectate structure and a microechinate ornamentation. Petrorhagia alpina subsp. alpina has the smallest pollen grain diameter (18.57 μm) and P. dubia the largest one (37.80 μm). The number of pores ranges from 12 (in P. dubia and P. saxifraga) to 22 (in P. alpina subsp. alpina and P. alpina subsp. olympica and P. hispidula) with a minimum pore diameter of 2.37 μm in P. alpina subsp. alpina and a maximum pore diameter of 4.23 μm in P. peroninii. The exine thickness ranges from 1.73 μm in P. saxifraga to 3.78 μm in P. pamphylica. In this study, the systematic implications are discussed in the light of palynological results.     

Aerodynamic particle size analysis of aerosols from pressurized metered-dose inhalers: Comparison of andersen 8-stage cascade impactor,next generation pharmaceutical impactor,and model 3321 aerodynamic particle sizer aerosol spectrometer

The purpose of this research was to compare three different methods for the aerodynamic assessment of (1) chloroflurocarbon (CFC)-fluticasone propionate (Flovent), (2) CFC-sodium cromoglycate (Intal), and (3) hydrofluoroalkane (HFA)-beclomethasone dipropionate (Qvar) delivered by pressurized metered dose inhaler. Particle size distributions were compared determining mass median aerodynamic diameter (MMAD), geometric standard deviation (GSD), and fine particle fraction <4.7 μm aerodynamic diameter (FPF_{<4.7 μm}). Next Generation Pharmaceutical Impactor (NGI)-size distributions for Flovent comprised finer particles than determined by Andersen 8-stage impactor (ACI) (MMAD=2.0±0.05 μm [NGI]; 2.8±0.07 μm [ACI]); however FPF_{<4.7 μm} by both impactors was in the narrow range 88% to 93%. Size distribution agreement for Intal was better (MMAD=4.3±0.19 μm (NGI), 4.2±0.13 μm (ACI), with FPF_{<4.7 μm} ranging from 52% to 60%. The Aerodynamic Particle Sizer (APS) undersized aerosols produced with either formulation (MMAD=1.8±0.07 μm and 3.2±0.02 μm for Flovent and Intal, respectively), but values of FPF_{<4.7 μm} from the single-stage impactor (SSI) located at the inlet to the APS (82.9%±2.1% [Flovent], 46.4%±2.4% [Intal]) were fairly close to corresponding data from the multi-stage impactors. APS-measured size distributions for Qvar (MMAD=1.0±0.03 μm; FPF_{<4.7 μm}=96.4% ±2.5%), were in fair agreement with both NGI (MMAD=0.9±0.03 μm; FPF_{<4.7 μm}=96.7%±0.7%), and ACI (MMAD=1.2±0.02 μm, FPF_{<4.7 μm}=98%±0.5%), but FPF_{<4.7 μm} from the SSI (67.1%±4.1%) was lower than expected, based on equivalent data obtained by the other techniques. Particle bounce, incomplete evaporation of volatile constituents and the presence of surfactant particles are factors that may be responsible for discrepancies between the techniques.     

Inhibitory Effects of Gynostemma Pentaphyllum on the UV Induction of Bacteriophage λ in Lysogenic Escherichia coli

Effects of gynostemma pentaphyllum (GP) on the bacteriophage λ induced by ultraviolet (UV) irradiation have been studied. The results showed that GP could inhibit the UV induction of bacteriophage λ in lysogenic cells. The inhibitory effects were dependent on the concentration and the reaction time of GP, and were efficient at 40∼125 μg ml⁻¹ for 10 min. The inhibitory rate was higher than 70% when the GP concentration was 50 μg ml⁻¹. By electron spin resonance (ESR) and spin-trapping techniques, the signals of free radicals were detected in the suspension of the λ lysogenic bacteria induced by ultraviolet irradiation, but after the addition of GP the signals were decreased. These results indicate that gynostemma pentaphyllum not only is a scavenger of free radicals, but also possesses the biological function of anti-irradiation, and that there is a close relation between the UV irradiation of the bacteriaphage λ and free radicals. Received: 18 December 2000 / Accepted: 9 March 2001     

A simple and rapid method for the detection of contaminants in a bioreactor

Chromatographic detection of particles with 0.05 to 10.0 μm in diameter by using a capillary tube is reported. Retention times of particles depended on their diameter. Complete peak separation was observed in the chromatogram of a mixture of particles with 0.05 and 10.0 μm in diameter. The application of this technique as detecting method for contaminants in a bioreactor is considered.     

Morphological differences among three species of newly settled pocilloporid coral recruits

 Investigation of the life history of corals is hampered by an inability to identify early recruits. In this study, the pattern of formation and morphology of the juvenile skeletons of three laboratory-reared pocilloporids, Seriatopora hystrix, Stylophora pistillata and Pocillopora damicornis, were compared to determine whether they could be reliably distinguished. The pattern of skeleton formation, including the origin and structure of the septa, columella and corallite wall was similar in all species. Following the completion of the primary corallite wall after 4–5 days, these species could be identified by differences in the diameter of the primary corallite. The mean diameter (±SE) of each species differed markedly: S. hystrix 400 ± 2.7 μm, range 325–450 μm; S. pistillata 505 ± 3.5 μm, range 400–550 μm; P. damicornis 697 ± 7.5 μm, range 492–885 μm. Values for the primary corallite diameter overlapped in only 3% of samples, demonstrating the potential utility of this feature as a tool for classifying recruits obtained from the field. Accepted: 4 January 2000     

Tracheids in white spruce seedling's long lateral roots in response to nitrogen availability

This study examined how the availability of inorganic nitrogen (N) modified the anatomical characteristics of white spruce (Picea glauca (Moench) Voss) roots related to their hydraulic properties. Seedlings were grown for one growing season in 4 L capacity pots filled with sand under one of three N levels: low (10 ppm), medium (50 ppm) and high (125 ppm). First order lateral roots with intact tips were sampled from dormant seedlings in October. Root segments were collected from 4, 10, and 14 cm distances above the root tip for fixation and sectioning and for maceration. Additional specimens were collected from the 4 and 14 cm distances for maceration and scanning electron microscopy of xylem pits. Root diameter and surface area occupied by the xylem in root cross sections increased basipetally in all treatments but exceptions were found. Higher N-levels significantly increased root diameter and surface area occupied by the xylem. In the two higher N treatments secondary root development was more advanced near the root tip than in the low N treatment. There was a strong positive correlation between root diameter and cross-sectional root area occupied by the xylem (30–50% of the root cross section) but not in portions with little secondary development. Non-conducting space within the xylem occupied 10–13% of its cross-sectional surface. Tracheids of the primary xylem were larger, had larger lumens but thinner cell walls than those of the secondary xylem. Low N treatment seedling tracheids had smaller total cross-sectional area, less lumen, and less cell wall surface area than the two other N treatments. Tracheid diameter means were between 19–20 μm in the high and medium N treatments, and 15.2 μm in the low N treatment. The range was 4.5–51.3 μm. Tracheid length was not significantly affected by N. The average tracheid was about 1000 μm long, and the range was 110–3530 μm. Pit-border diameters ranged between 4.1–20.6 μm (average 10–11 μm) and were not affected by the N treatment. Pit aperture diameters were within 0.62–10.2 μm range (average between 3–4 μm) and were also not significantly affected by the N treatment, although tracheids from the medium N-treatment roots tended to have larger apertures. The pit border diameter equals that of the margo while the aperture size should be similar to that of the torus of the pit membrane. If the capacity for axial water transport in spruce roots is affected by N, it would be by its impact on conduit diameter and, possibly on the pit-membrane pore sizes but not by changes to conduit length and to the size of the pit membrane surface area. This revised version was published online in June 2006 with corrections to the Cover Date.     

Neuronal nitric oxide synthase immunopositive neurons in cat vestibular complex: a light and electron microscopic study

Nitric oxide is a unique neurotransmitter, which participates in many physiological and pathological processes in the organism. Nevertheless, there are little data about the neuronal nitric oxide synthase immunoreactivity (nNOS-ir) in the vestibular complex of a cat. In this respect, the aims of this study were to: (1) demonstrate nNOS-ir in the neurons and fibers, from all major and accessory vestibular nuclei; (2) describe their light microscopic morphology and distribution; (3) investigate and analyze the ultrastructure of the NOS I-immunopositive neurons, fibers, and synaptic boutons. For demonstration of the nNOS-ir, the peroxidase–antiperoxidase–diaminobenzidin method was applied. Immunopositive for nNOS neurons and fibers were present in all major and accessory vestibular nuclei. On the light microscope level, the immunopositive neurons were different in shape and size. According to the latter, they were divided into four groups—small (with diameter less than 15 μm), medium-sized (with diameter from 15 to 30 μm), large type I (with diameter from 30 to 40 μm), and large type II (with diameter greater than 40 μm). On the electron microscope level, the immunoproduct was observed in neurons, dendrites, and terminal boutons. According to the ultrastructural features, the neurons were divided into three groups—small (with diameter less than 15 μm), medium-sized (with diameter from 15 to 30 μm), and large (with diameter greater than 30 μm). At least two types of nNOS-ir synaptic boutons were easily distinguished. As a conclusion, we hope that this study will contribute to a better understanding of the functioning of the vestibular complex in cat and that some of the data presented could be extrapolated to other mammals, including human.     

Conformational flexibility of alpha-lactalbumin related to its membrane binding capacity

Different folding states of the small, globular milk protein bovine alpha-lactalbumin (BLA) induced by the anionic surfactant sodium dodecylsulphate (SDS) have been examined by fluorescence spectroscopy, CD and NMR. The solution structure of the protein in the absence of SDS was also determined, indicating fluidity even under native conditions. BLA is partly denatured to a molten globule (MG)-like state by micromolar concentrations of SDS, and the transitions from native to MG-like state are dependent on pH, the protein being more sensitive to the surfactant at pH 6.5. As indicated by measurements of the intrinsic emission fluorescence, the tertiary structure disappears at lower concentrations of SDS than most of the secondary structure, as estimated from CD data. The MG-like state induced by low concentrations of SDS is not observable by NMR, and is probably fluctuating and/or aggregating. At higher concentrations of SDS above the critic concentration of micelles, an NMR-observable state reappears. This micelle-associated conformer was partially assigned, and found to bear strong resemblance to the acid-tri-fluoroethanol state, retaining weakened versions of the A and C helix of native BLA. We discuss the results in terms of the inherent flexibility of the protein, and its ability to form multiple folding states and to bind to membranes. Also, we propose that proteins with stable MG-like conformers can have these states stabilized by low levels of compounds with surfactant properties in vivo.     

Induction and development of somatic embryos from spinach (Spinacia oleracea) leaf segments

Somatic embryogenesis was induced in embryogenic calli of Spinacia oleracea L., on a Murashige and Skoog (1962) medium, containing 4.6 μM kinetin as the sole growth regulator. Abscisic acid, gibberellic acid, and indole-3-acetic acid were supplemented to kinetin-containing medium and their effects on the initiation of somatic embryos was studied. Abscisic acid at a particular concentration (4 μM) dramatically increased the number of embryos per g fresh weight of callus, while both gibberellic acid and indole-3-acetic acid suppressed the embryo initiation. It is suggested that the promoting effect of abscisic acid on the embryo initiation may be explained as a stress response of the tissue. The relative number of globular embryos vs. the embryos in heart/torpedo and cotyledonary stages was increased at 4 μM abscisic acid and at all gibberellic acid concentrations (0.3–10 μM). In contrast, the ratio of globular to polar embryos was lower than in controls at 1 μM abscisic acid and indole-3-acetic acid (1 and 10 μM). The effects of growth regulators on the ratio of globular to polar embryos indicate that they interfere with the normal distribution of cell division and cell expansion during early embryogenesis. This revised version was published online in June 2006 with corrections to the Cover Date.     

Synthesis and characterization of the antibacterial potential of ZnO nanoparticles against extended-spectrum β-lactamases-producing <Emphasis Type="BoldItalic">Escherichia coli</Emphasis> and <Emphasis Type="BoldItalic">Klebsiella pneumoniae</Emphasis> isolated from a tertiary care hospital of North India

The reemergence of infectious diseases and the continuous development of multidrug resistance among a variety of disease-causing bacteria in clinical setting pose a serious threat to public health worldwide. Extended-spectrum β-lactamases (ESBLs) that mediate resistance to third-generation cephalosporin are now observed all over the world in all species of Enterobacteriaceae, especially Escherichia coli and Klebsiella pneumoniae. In this work, ZnO nanoparticles (NPs) were synthesized by the sol–gel method and characterized by powder X-ray diffraction, scanning electron microscopy (SEM) and atomic force microscopy (AFM). The image of synthesized ZnO NPs appeared spherical in SEM with a diameter of ≈19 nm and as hexagonal crystal in AFM. Clinical isolates were assessed for ESBL production and shown to be sensitive to ZnO NPs by different methods such as minimal inhibitory concentration (MIC) and minimal bactericidal concentration, time-dependent growth inhibition assay, well diffusion agar methods and estimation of colony forming units (CFU) of bacteria. The lowest MIC value for E. coli and K. pneumoniae was found to be 500 μg/ml. The results showed that ZnO NPs at 1,000 μg/ml completely inhibit the bacterial growth. The antibacterial effect of ZnO nanoparticles was gradual, but time- and concentration-dependent. The maximum inhibition zone at100 μg/ml for E. coli and K. pneumoniae was 22 and 20 mm, respectively. With the increasing ZnO NP loading, there is significant reduction in the numbers of CFU. At the concentration of 1,000 μg/ml, the decline in per cent survival of E. coli and K. pneumoniae was found to be 99.3% and 98.6%, respectively.     

Distribution and morphology of the ampullary organs of the estuarine long-tailed catfish, <Emphasis Type="Italic">Euristhmus lepturus</Emphasis> (Plotosidae,Siluriformes)

Ampullary organs of Euristhmus lepturus occur in high densities along the head and in four parallel pathways along the trunk of the body. Large ampullary pores (125–130 μm) are easily distinguishable from other sensory epithelial pores due to the differences in size and the presence of a collar-like structure. Simple, singular ampullary organs of the head region consist of an ampullary pore connected to a long canal with a diameter of 115–175 μm before terminating as a simple ampulla with an external diameter of 390–480 μm. The ampullary canal is composed of 1–2 layers of flattened squamous epithelial cells, the basement membrane and an interlocking collagen sheath. The innermost cells lining the canal wall are adjoined via tight junctions and numerous desmosomes, as are those of the receptor and supportive cells. Canal wall tissue gives rise to a sensory epithelium containing between 242 and 285 total receptor cells, with an average diameter of 11.7 ± 5.3 μm, intermixed with medially nucleated supportive cells. Each receptor cell (21.38 ± 4.41 μm, height) has an apically positioned nucleus and a luminal surface covered with numerous microvilli. Neural terminals abut the basal region of receptor cells opposite multiple presynaptic bodies and dense mitochondria. Supportive cells extend from the ampullary lumen to the basement membrane, which is adjacent to the complex system of collagen fibres.     

Detection of distinct endocytotic and phagocytotic activities in epithelial cells (pinacocytes) of freshwater sponges (Porifera, Spongillidae)

 Light and electron microscopical investigations using externally applied fluorescent and gold-labeled markers have revealed the existence of distinct endocytotic and phagocytotic activities in basal epithelial cells (pinacocytes) of the freshwater sponges Spongilla lacustris and Ephydatia es) of the f. The total rate of endocytotic membrane uptake, ascertained by the application of the cationic lipid probe TMA-DPH, was found to be 3.2% of the cell surface area/h. A typical fluid-phase endocytosis, demonstrated by the use of the water-soluble membrane-impermeable tracers BCECF-dextran and LY-CH, participates in endocytotic activity at a rate of 0.7% of the cell surface area/h and results in the formation of endosomes measuring 0.8–1 μm in diameter. Moreover, the application of labeled BSA succeeded in the detection of a receptor-mediated endocytosis amounting to a concentration-dependent uptake of 2.3–2.8% of the cell surface area/h. Coated pits and coated vesicles conveying the adsorbed BSA measure 0.3 μm in diameter and are covered on the cytoplasmic face with a clathrin-like protein (HC, 180 kDa; LC, 30 kDa). To facilitate phagocytotic activities, a series of fluorescent–labeled and chemically treated particles such as bacteria or latex beads have been successfully employed. Accordingly, the measured values of phagocytic membrane uptake between 1 and 8% of the cell surface area/h depend on the variety of size as well as the chemical nature of the different bioparticles and clearly point to phagocytosis as a key mechanism for providing freshwater sponges with nourishment. Accepted: 3 April 1997     

Cases of spermiogenesis in young European bison

Development of spermiogenesis and sizes of seminiferous tubules of the testes and epididymal duct were studied in 45 young European bisonBison bonasus (Linnaeus, 1758) males from an enclosed breeding centre and a free-ranging population in the Białowieża Forest. Of 13 males in age class II (up to two years), four showed the presence of elongated late spermatids: one male aged 15 months from the enclosed breeding centre and three from the free-ranging population (2 males aged 18 months and one about 24 months old). Of 7 males from age class III (up to 3 years), four showed spermiogenesis, of which the youngest, 26 and 32 months old, were from enclosed breeding. Spermiogenesis was observed in males with high body weight in a given age class. In males of age class I (up to one year old), the mean diameter of the seminiferous tubules was 49.8 μm, in class II — 110.7 μm, and in class III — 162 μm, the mean diameter of the epididymal duct being 110.8 μm, 187.2 μm and 273.4 μm, respectively. Measurements of seminiferous tubules and epididymal duct were significantly correlated with age and body weight of males and differed significantly between the three age classes.     

Spawning of the colonial coral <Emphasis Type="Italic">Cladocora caespitosa</Emphasis> (Anthozoa,Scleractinia) in the Southern Adriatic Sea

Data on sexual reproduction of scleractinian coral species living in temperate zones, particularly in the Mediterranean Sea, are quite scarce. This paper describes sexual reproduction of the colonial coral Cladocora caespitosa from Veliko jezero (Mljet Island) in the Adriatic Sea. Spawned orange eggs and white sperm bundles were observed on the coral bank of C. caespitosa two nights before the full moon (20 June 2005) coinciding with increasing water temperature and correlated with the lunar cycle. Spawning was observed during five nights, involving about 30% of the colonies from the coral bank. Different colonies on the bank released only one type of gamete during the reproductive period. The diameter of the sperm bundles ranged from 100 to 200 μm (average 163 μm; SD = 47.08), while the female gametes diameter ranged from 300 to 500 μm (average 416 μm; SD = 73.12).     

Mesenterial filaments of the black coral <Emphasis Type="Italic">Cirrhipathes</Emphasis> cfr. <Emphasis Type="Italic">anguina</Emphasis> provide a home to developing nauplii

Inspection of two female colonies of the monopodial black coral Cirrhipathes cfr. anguina from the coral reef of the Marine Park of Bunaken (Indonesia) revealed the occurrence of crustacean developing eggs within the mesenterial filaments of the polyps. Egg diameter, which in the smallest gametes was about 50–60 μm, increased in tandem with embryo development, reaching the value of 170 μm, at the nauplius stage. The attribution to the crustacean taxon was derived from morphological investigations carried out in light and electron microscopy (TEM, SEM) on the eggs and on the embryos removed from them. The final stage of nauplius was characterised by three pairs of appendages: uniramouse antennulae, biramouse antennae and manidibulae. In addition, naupliar eye and caudal setae were also evident. These nauplii were ascribed to the larval stage of an unidentified species. Coral/copepod association could represent a reproductive strategy, put into action by some marine copepods. Incubation within an appropriate host prevents predation by planktotrophic organisms, thus reducing population depletion.     

Vegetative nuclei and differentiation of chromosomes inTrichophyton vanbreuseghemii

The stages of nuclear division were observed inTrichophyton vanbreuseghemii microcultures stained with Giemsa and by the Feulgen reaction. The course of karyokinesis is discussed. It takes the form of mitosis, modified by narrowing of the lumen of the hyphal cell and the movement of the cytoplasm. No spindle or centrioles were found. In vegetative hyphae (1.2–1.5 μm in diameter), the metaphase chromosomes were arranged lineally in a row following the direction of the long axis of the hyphal cell. The distribution of anaphase chromosomes occurs perpendicularly or obliquely to the cell walls. The chromosomes were spherical, with a diameter of about 0.3 μm. A haploid number of chromosomes (five) was found.     

Atypical micromegakaryocytes,promegakaryoblasts and megakaryoblasts: a critical evaluation by immunohistochemistry,cytochemistry and morphometry of bone marrow trephines in chronic myeloid leukemia and myelodysplastic syndromes

A morphometric analysis of bone marrow trephine biopsies has been performed to study the frequency and planimetric characteristics of so-called atypical micromegakaryocytes in chronic myeloid leukemia (CML) and myelodysplastic syndromes (MDS). In addition, an attempt was made to discriminate this particular cell population from small immature elements of megakaryocytopoiesis, such as promegakaryoblasts and megakaryoblasts. The staining reactions employed included periodic acid-Schiff (PAS), alpha-naphthyl acetate esterase (ANAE) and immunohistochemistry with a monoclonal antibody against platelet glycoprotein IIIa (Y₂/ 51-CD61). Comparison of the various staining reactions applied to the different megakaryocytic elements together with morphometric measurements resulted in a clearcut identification of promegakaryoblasts. These were defined as the earliest immature and exclusively CD61-positive precursors. Atypical micromegakaryocytes were characterized by their dysplastic features and strong ANAE reactivity in addition to their positive CD61 staining. When stringent diagnostic criteria (diameter ranging between 10 to 15 μm, mean size about 12 urn) were applied, this abnormal cell population comprised less than 10% of total megakaryocytopoiesis in CML and MDS. It may be assumed that dysmegakaryocytic features in the latter disorders are partially generated by small to medium-sized megakaryocytes (diameter less than 30 μm). In conclusion, the relative frequency of promegakaryoblasts in the normal bone marrow (range 6–8%) is confirmed by evaluation of the immunohistochemical and cytochemical staining methods (CD61 and ANAE). Furthermore, the ANAE reaction facilitates the recognition of atypical micromegakaryocytes as well as small megakaryocytes. Thus cytochemistry provides a better insight into alterations of these cell lineages in various pathological conditions. This work was supported by the Deutsche Forschungsgemeinschaft (DFG-Th 390/1–2)