果胶酶的细胞化学定位：马尾松树脂道发生的细胞化学证据

运用薄切片和电镜细胞化学方法观察了马尾松(Pinus massoniana Lamb.)茎皮层树脂道的发育及发育过程中果胶酶的变化。树脂道的发育过程一般可分为4个阶段,即原始细胞阶段、胞间隙形成阶段、腔道扩大阶段和树脂道成熟阶段。在原始细胞阶段,果胶酶的反应产物首先出现在原始细胞膨胀的细胞壁角隅处,然后沿细胞壁中层分布。胞间隙形成后,果胶酶的反应产物分布在细胞壁和胞间隙的交界面。随着胞间隙的扩大,反应产物的密度逐渐降低。当树脂道成熟后,上皮细胞壁上则没有果胶酶的反应产物出现。细胞化学证据表明: 在树脂道的发育过程中,果胶酶降解树脂道原始细胞细胞壁中层,支持树脂道以裂生方式形成。果胶酶的细胞化学定位技术还可用于其他植物中与果胶水解有关的发育过程。     

果胶酶的细胞化学定位:马尾松树脂道发生的细胞化学证据

运用薄切片和电镜细胞化学方法观察了马尾松(Pinus massoniana Lamb.)茎皮层树脂道的发育及发育过程中果胶酶的变化.树脂道的发育过程一般可分为4个阶段,即原始细胞阶段、胞间隙形成阶段、腔道扩大阶段和树脂道成熟阶段.在原始细胞阶段,果胶酶的反应产物首先出现在原始细胞膨胀的细胞壁角隅处,然后沿细胞壁中层分布.胞间隙形成后,果胶酶的反应产物分布在细胞壁和胞间隙的交界面.随着胞间隙的扩大,反应产物的密度逐渐降低.当树脂道成熟后,上皮细胞壁上则没有果胶酶的反应产物出现.细胞化学证据表明:在树脂道的发育过程中,果胶酶降解树脂道原始细胞细胞壁中层,支持树脂道以裂生方式形成.果胶酶的细胞化学定位技术还可用于其他植物中与果胶水解有关的发育过程.     

油红O染色原代未活化胰腺星状细胞脂肪滴的实验观察

目的观察油红O染色原代培养的未活化胰腺星状细胞脂肪滴,并予鉴定。方法选取接种于6孔板中培养4天的原代未活化胰腺星状细胞,予100g/L甲醛液固定,磷酸盐缓冲液漂洗后,加入5g/L油红O饱和液染色,镜下动态观察。胞浆内的脂肪滴一旦着色,即可洗去油红,苏木素衬染胞核,漂洗分化脱色后镜下成像。结果未活化的胰腺星状细胞脂肪滴被油红0染成鲜艳的红色,大小不一的串珠样"油珠子"分散于胞质中,簇集成"环状",呈戒环样包绕在细胞核周围。结论油红染色原代未活化胰星状细胞脂肪滴是鉴定该细胞的重要方法之一。     

油松茎次生木质部中树脂道的发育过程和组织化学研究

利用组织化学方法对油松茎次生木质部树脂道发育过程中上皮细胞内树脂滴和淀粉粒的动态变化进行了研究。发现在树脂道原始细胞阶段,每个原始细胞含淀粉粒较少,含树脂滴稀少。在树脂道形成阶段,淀粉粒数目较多,体积增大,树脂滴也呈递增趋势。在树脂道成熟阶段,淀粉粒数目变化不大,而体积明显变小,树脂滴的体积增大,数目减少。     

少量贴壁培养细胞电镜原位包埋方法的探讨

该文探讨了对少量贴壁培养细胞较易操作且能保存较好超微结构的透射电镜样品包埋的方法。将Hela细胞分为三组:(1)不使用环氧丙烷,将树脂胶囊直接倒扣包埋于塑料培养皿;(2)不使用环氧丙烷,将细胞爬片倒扣包埋于胶囊;(3)使用环氧丙烷并将细胞爬片倒扣包埋于胶囊。将三组带有细胞的树脂胶囊进行超薄切片,电镜观察后发现,第一种方法包埋简便,超薄切片上无细胞缺失孔洞,且超微结构保存较好。     

白花前胡营养器官结构及其分泌道分布规律的研究

运用石蜡切片方法,观察白花前胡营养器官的显微结构及其分泌道的分布特征,以明确营养器官内分泌结构的分布规律,为揭示白花前胡次生代谢产物的积累提供解剖学依据。结果表明,(1)白花前胡成长根从外到内由周皮、中柱鞘薄壁组织和次生维管组织组成,而且中柱鞘薄壁组织不同于一般双子叶植物根的结构;茎从外到内由表皮、皮层和维管柱组成;叶为异面叶结构。(2)白花前胡根、茎及叶中均有分泌道存在,分泌道在根中分布于中柱鞘薄壁组织和次生韧皮部中,茎中分布于皮层和髓中,叶中分布于维管束上下两侧的薄壁组织中。     

Rubisco和RCA在青菜叶绿体中的分布及病毒侵染对其细胞定位的影响

运用免疫金标电镜术观察了青菜叶细胞中光合作用关键酶Rubisco和Rubisco活化酶(RCA)的细胞化学定位,结果显示Rubisco和RCA免疫金颗粒主要分布于薄壁组织叶绿体的间质中,在基粒片层上很少,表皮的气孔保卫细胞和维管束薄壁细胞叶绿体内也有分布,在细胞质及线粒体等细胞器中无特异性分布。同时比较观察了感染芜菁花叶病毒(TuMV)的青菜叶绿体Rubisco和RCA免疫金标记结果,发现病组织中结构尚完整的叶绿体Rubisco和RCA标记率略有下降,而结构严重破坏的叶绿体中两种酶标记率分别仅为正常叶绿体的58．44％和64．67％,表明病毒侵染可导致Rubisco和RCA含量下降,影响寄主植物的光合作用。     

光(温)敏核不育水稻花药和小孢子发生的细胞化学

利用细胞化学方法,对光（温）敏核不育水稻农垦585和W6154S的花药和小孢子发生过程的观察结果表明,在可育条件下,其花药组织和小孢子发生过程不论形态结构还是细胞化学变化都基本一致。小孢子母细胞时期的药隔薄壁组织、药壁中层及药室内壁中分布了一些多糖颗粒,但到进入减数分裂时多糖颗粒基本消失。绒毡层在解体前一直富含细胞质,从染色反应看,它表现为小孢子母细胞时期的蛋白质向减数分裂开始后的多糖物质的转变过程。在不育条件下,农垦585在小孢子母细胞时期就出现异常,其败有时间比W6154S要稍早一些。两者最后都表现为典败,但W6154S的花药壁解体较为彻底,只剩下干皱的表皮和药室内壁,而农垦585的花药壁还有多层细胞结构。     

人毛乳头细胞组织化学研究

毛乳头细胞是一种高度特殊化的成纤维细胞。本文通过对体外培养的毛乳头细胞进行组织化学染色研究发现,它对阿新蓝、甲苯胺蓝和PAS染色均呈阳性,并对甲苯胺蓝显异染性．与原位时的细胞染色结果相同,表明在体外培养下．毛乳头细胞合成和分泌酸性、中性粘多糖的能力仍能维持较长时间;在细胞聚集区和多层化细胞团中有丰富的细胞外基质,阿新蓝和PAS染色呈强阳性,说明细胞外基质的存在与毛乳头细胞的聚集有很大关系;另外毛囊真皮鞘细胞对阿新蓝、甲苯胺蓝染色呈阳性反应．无甲苯胺蓝的异染性,PAS染色阴性,而真皮成纤维细胞这些染色均阴性,说明它与毛乳头细胞关系密切。     

香蕉束顶病和花叶心腐病的快速测定技术研究

种植无病蕉苗是防治香蕉束顶病和花叶心腐病的根本措施。筛选无病蕉苗的快速测定技术。除可引用2,3,5-氯化三苯基四(氨)唑浸渍徒手切片;在36℃条件下。24小时后:束顶病株叶脉切片的维管束呈现红色,其它组织为红褐色;花叶心腐病株叶脉切片(含维管束)全部呈现黑褐色;而健株叶脉切片由原绿色变成红色,最后褪成无色。此外。也可取叶片主脉的徒手切片,用1％曙红B水溶液染色。健株切片呈桔红色,病株切片呈砖红色,通过镜检:束顶病株叶片切片的表皮下,可观察到围绕在维管束周围成堆的分布有畸形叶绿体细胞团;花叶心腐病的畸形叶绿体细胞团,多分布在薄壁细胞中,且多散生;而健株的切片没有发现这种畸形的叶绿体细胞团,两种方法同样可靠。     

Ultrastructure of the secretory cells of the submucosal glands in the human maxillary sinus

Tissue samples obtained from the lateral wall of the maxillary sinuses of five patients were examined by light microscopical, histochemical, and ultrastructural techniques. Submucosal glands were tubulo-alveolar mixed glands. The acini consisted of either all serous or all mucous cells, or a mixture of both. Serous granules were stained by toluidine blue, or by hematoxylin and eosin (H and E), but showed little or no reaction with periodic acid-Schiff (PAS) or Alcian blue. Mucous granules were pale in toluidine blue or H and E preparations, and consisted primarily of acid mucosubstances, as demonstrated by their staining reaction with PAS and Alcian blue. At the electron microscope level, the serous granules were either homogeneously dense, or showed a substructure consisting of at least two layers of distinctly different electron-opacity. Typical mucous droplets consisted of a fibrillar network dispersed in a translucent matrix. A second secretory product was present in the mucous cells in the form of elongated, membrane-bounded structures containing numerous parallel filaments, which measured about 55 Å in diameter. The mucous droplets and the filamentous bodies appear to arise from the opposite faces of the Golgi complex in the mucous cells. The filamentous bodies showed a pronounced tendency to fuse with the mucous droplets. All acini were surrounded by a well-defined myoepithelial layer and contained intercellular nerve terminals.     

Fine structural and histochemical studies on salivary glands of Peripatoides novae-zealandiae (Onychophora) with special reference to acid phosphatase distribution

The Onychophora feed on small arthropods and produce saliva when ingesting prey. Although saliva undoubtedly helps to liquefy the food its constituents have not yet been fully described. The salivary glands, two long tubes of glandular epithelium, are known to secrete a powerful protease, however, besides other enzymes and mucus. In Peripatoides novae-zealandiae there are protein-secreting cells of three types, referred to here as columnar, cuboidal and modified cells, and mucus cells. The anterior two-thirds of the gland show most cell diversity, while the posterior region consists mainly of columnar cells. These are the most numerous elements overall and they probably secrete salivary protease. In thick resin sections the granules of all protein-secreting cells stain strongly with methylene blue. Those of columnar cells are markedly uneven in size and accumulate distally, eventually filling the cytoplasm. More proximal Golgi regions may be discernible. Mucus cells are all of one type and their secretion droplets are stained lightly by methylene blue. The electron microscope shows that distal microvilli, desmosomes and septate junctions are common to all gland cells. In columnar cells, secretory material is contributed by Golgi complexes and by rough endoplasmic reticulum. Early secretory vacuoles containing dense material are seen in the concavity of Golgi regions. They are precursors to larger condensing vacuoles whose contents have a more flocculent appearance, and which may attain 3–4 μm in diameter. These evolve into secretory granules, usually of uneven texture, which are up to 2–5 μm in diameter. Histochemical tests for acid phosphatase show moderate amounts of enzyme throughout the gland. In whole mounts and sections the strongest reaction is in a band of cuboidal cells along the anterior median border. Columnar cells show a diffuse cytoplasmic reaction towards the base and sometimes distal to the nucleus, and mucus cells may also react strongly round the nucleus. Cytoplasm near the lumen shows little reaction. The secretory granules do not appear to contain active enzyme. Under the electron microscope a positive reaction for acid phosphatase is seen in lysosomal derivatives near the base and lateral periphery of gland cells. These bodies are probably autophagic vacuoles and they may contain membranous whorls and possibly old secretion granules. Acid phosphatase is involved also in the elaboration of new secretory granules in both columnar and mucus cells. Dense reaction product is found in a system of interconnected tubules and cisternae near the innermost face of the Golgi complex, which is interpreted as GERL. Acid phosphatase is present in the peripheral zone of adjacent early secretory vacuoles, and interconnections occur between GERL and secretory vacuoles. It is suggested that GERL tubules containing the enzyme may fuse with early secretory vacuoles and release acid phosphatase at their periphery. The acid phosphatase reaction is negative in large condensing vacuoles and most secretory granules. These findings are consistent with what is known from mammalian cells, including those of salivary glands.     

Open-Face, Epoxy Embedding of Single Cells for Ultrathin Sections

Intact stamens of Tradescantia were fixed, dehydrated, and infiltrated with an epoxy resin. Each stamen was then put into a drop of resin on a microscope slide, which was transferred to the stage of a dissecting microscope so that individual hairs could be detached from the filament with fine tungsten needles. The detached hairs were transferred to drops of resin ca. 2 mm in diameter (6 or 7 in each of two rows) lying on a slide heavily coated with evaporated carbon. Polymerization was carried out in an oven until the resin attained a degree of viscosity that permitted orientation of the isolated hairs (by using a compound microscope) without their subsequent dislocation. When the small drops of resin had hardened after further polymerization, the positions of the hairs were marked by circumscribing the cells with India ink. The block was pried from the slide after rapid cooling with solid CO₂, and was then trimmed and sectioned. Cells suspended in culture medium were embedded in much the same way; they were centrifuged to obtain a pellet, which was fixed, dehydrated, and infiltrated. A small fragment of the pellet with a little resin was placed on a microscope slide, where the cells were dissociated under a dissecting microscope at ca. 100 × magnification. Individual cells were then picked up with tungsten needles and transferred to droplets of resin on a carbon-coated slide. The subsequent steps were similar to those described for the staminate hairs. Pieces of tissue in the 50-500 μ range were also handled by the foregoing technique. However, after infiltration they were put into large drops of resin on a slide coated with silicone mold-release rather than on a surface coated with carbon.     

A polychromatic staining method for epoxy embedded tissue: a new combination of methylene blue and basic fuchsine for light microscopy

A simple and rapid method is described for staining semithin sections of material embedded in epoxy resin for observing tissues prior to transmission electron microscopy. The method is suitable for tissue fixed with a glutaraldehyde-formaldehyde mixture and postfixed in osmium tetroxide. No etching or oxidizing procedures are necessary. Sections 0.5-0.8 µm thick are dried onto a slide and stained with either 0.75% methylene blue and 0.25% azure B or 0.5% methylene blue and 0.5% azure II in 0.5% aqueous borax and heated over a flame for 8-10 sec. The slides are rinsed with water, then stained the same way with 0.1% basic fuchsine in 5% aqueous ethanol. Cytoplasm stains blue; nuclei darker blue; collagen, mucus and elastin pink to red; fat and intracellular lipid droplets gray-green.     

A polychromatic staining method for epoxy embedded tissue: a new combination of methylene blue and basic fuchsine for light microscopy

A simple and rapid method is described for staining semithin sections of material embedded in epoxy resin for observing tissues prior to transmission electron microscopy. The method is suitable for tissue fixed with a glutaraldehyde-formaldehyde mixture and postfixed in osmium tetroxide. No etching or oxidizing procedures are necessary. Sections 0.5–0.8 µm thick are dried onto a slide and stained with either 0.75% methylene blue and 0.25% azure B or 0.5% methylene blue and 0.5% azure II in 0.5% aqueous borax and heated over a flame for 8–10 sec. The slides are rinsed with water, then stained the same way with 0.1% basic fuchsine in 5% aqueous ethanol. Cytoplasm stains blue; nuclei darker blue; collagen, mucus and elastin pink to red; fat and intracellular lipid droplets gray-green.     

A polychromatic staining method for epoxy embedded tissue: a new combination of methylene blue and basic fuchsine for light microscopy.

A simple and rapid method is described for staining semithin sections of material embedded in epoxy resin for observing tissues prior to transmission electron microscopy. The method is suitable for tissue fixed with a glutaraldehyde-formaldehyde mixture and postfixed in osmium tetroxide. No etching or oxidizing procedures are necessary. Sections 0.5-0.8 microm thick are dried onto a slide and stained with either 0.75% methylene blue and 0.25% azure B or 0.5% methylene blue and 0.5% azure II in 0.5% aqueous borax and heated over a flame for 8-10 sec. The slides are rinsed with water, then stained the same way with 0.1% basic fuchsine in 5% aqueous ethanol. Cytoplasm stains blue; nuclei darker blue; collagen, mucus and elastin pink to red; fat and intracellular lipid droplets gray-green.     

Fine structure of the corpus allatum of the female blow-fly Calliphora erythrocephala

Summary An electron microscopical study of the corpus allatum (CA) of the adult female Calliphora was undertaken.The cells have a very irregular shape. Light and dark cells are found. Mitochondria occur in great numbers. Microtubules are frequently observed. Free ribosomes are plenty, but rough-surfaced reticulum is scarce. Golgi complexes are not very conspicuous. Axons, mostly containing neurosecretory granules, are frequently found between the cells.The active corpus allatum is remarkable by the numerous lipid droplets and the abundance of tubular agranular reticulum. The reticulum sometimes forms aggregates from which vacuoles are budded off. The vacuoles lose their membrane, at the same time becoming slightly electron opaque, thus being transformed into lipid droplets.It is tentatively postulated that the hormone (or a precursor) is synthesized in the tubules of the agranular reticulum, collected in the vacuoles, and, when the membrane disintegrates, it is dissolved in lipid. The lipid droplets are thought to be released into the haemolymph through the surface of the gland or via intercellular channels.The inactive corpus allatum of the six days old sugar fed flies is small and more or less shrunken. The agranular reticulum is poorly developed, vacuoles are small, and lipid droplets few. The reticulum tends to form whorls, which eventually may possibly be transformed into myelin figures.We wish to express our gratitude to the Danish Natural Science Research Council for placing a Zeiss electron microscope at our disposal, and to the Carlsberg Foundation for supporting our work with grants. We are grateful to Prof. C. Overgaard Nielsen for laboratory facilities, and we are indebted to Mrs. Eva Jensen for her skilful technical assistance.     

The differentiation of early embryonic stem cells into adipocytes-like cells

An early embryonic stem cell line, EES-6 cells, was established from 2-cell stage embryos of ddY mice. The cells were maintained in an undifferentiated state with D-MEM/F12 medium supplemented with 10% fetal bovine serum (FBS) (GM) and 1 ng of leukemia inhibitory factor (LIF) without any feeder cells. In this study, EES cells were cultured with a medium containing embryotrophic factors (ETFs) which promoted the differentiation of EES cells into white and brown adipocytes-like cells for a period of 5 days. Lipid droplets in brown adipocyte-like cells were stained with Sudan III; however, large lipid-like droplets in white or brown adipocyte-like cells were unstained with either Sudan III or alcian blue. These findings have numerous possibilities for therapeutic use such as regeneration of skin and wound healing.     

The Floral Nectary of Tropaeolum majus L.--The Nature of the Secretory Cells and the Manner of Nectar Secretion

The floral nectary of Tropaeolun majus L. was studied with theaid of a microscope with transmitting and incident light, atransmission electron microscope and a scanning electron microscope.The Gomori method was used for the localization of acid phosphatase.As a result of this investigation the previously accepted viewthat nectar in this plant is secreted only from the hair tipsof the inner epidermis of the calyx spur was found to be inaccurate.The present studies showed that the parenchyma cells locatedbetween the inner epidermis and the region of the vascular bundlesof the lowest third of the spur, are the main nectar-secretingelements of the nectary. These secretory cells release the nectarsolution into intercellular spaces leading to modified stomata,through which it is exuded into the spur cavity. The modifiedstomata occur in the lowest portion of the spur only. At thestage of secretion small droplets of liquid of high viscositywere observed on the epidermal hairs. These droplets presumablycontain polysaccharides and a certain amount of sugar.