易卷曲叶表皮制片技术（NaOCl法）的改进

在利用NaOCl法制备用于光学显微镜观察的叶表皮过程中,一些科/属植物的叶表皮在100%乙醇脱水后遇二甲苯便发生卷曲,增加了叶表皮制片的难度, 甚至无法进行。本文介绍一种简便易行的方法：在系列乙醇脱水后,加盖小块盖玻片,防止叶表皮脱水后卷曲,并使其保持平整,便于显微摄影。这种方法使得叶表皮显微制片技术更加完善。     

一种禾本科植物鲜叶下表皮制片方法

介绍一种禾本科植物鲜叶下表皮制片的快速方法,该方法也能适用于多数单子叶植物鲜叶和经FAA固定液固定的单子叶植物叶片的气孔观察。该方法制片能有效防止叶表皮脱水、透明后发生卷曲,使保持完整的结构,便于准确的显微观察。具操作简单、材料易得、可靠性和稳定性高等优点。     

蕨类植物腊叶标本叶表皮制片技术的改进

对蕨类植物腊叶标本叶表皮的制片技术进行了一些改进,克服了传统叶表皮制片中易出现的不足之处,简化了整个制作过程.这种方法使得蕨类植物叶表皮显微制片技术更加完善,可以为从事植物形态分类学基础研究的工作者提供技术支持.     

简便有效的叶表皮离析方法——过氧化氢-醋酸法

在长期叶表皮制片过程中发现,过氧化氢-醋酸离析法简便实用,对蕨类植物小型叶和大型叶、裸子植物鳞形叶、钻形叶、针形叶、条形或带形叶及被子植物宽大的叶片均能适用。尤其重要的是,过氧化氢-醋酸离析法不会破坏叶表皮细胞的结构,有利于精确观察叶表皮形态特征,是一种比较理想的叶表皮离析方法。     

植物叶表皮永久制片技术的改进

对植物叶表皮永久装片的制作过程,在实践中通过摸索而进行了一些改进,克服了传统叶表皮制片中易出现的几点不足之处,完善并简化了整个制作过程。新的永久的制片技术可以为从事植物形态分类学基础研究的工作者们提供基础的技术支持。     

石蜡切片法中染色技术的改良

石蜡切片是观察动植物组织最直接有效的方法之一。本实验对水稻品种9311的叶片进行切片并用1%的番红—乙醇染液和1%的苯胺蓝—乙醇染液进行染色,结果显示此改良方法得到的组织比传统石蜡切片更完整,染色的效果比传统石蜡切片更清晰,并且利用此方法对水稻叶片进行制片,可以大量缩短制片后期复水及脱水的时间,提高工作效率。     

一种简便快速鲜叶表皮制片技术

用透明胶带夹粘住适当大小的新鲜植物叶片,经轻轻挤刮,撕开两胶带,上、下表皮粘附在两胶带上,经NaOCl溶液处理3～5min,以离析粘附在表皮上的叶肉细胞,在流水下用软刷刷洗,凉干后,放在载玻片上盖上盖玻片,即可在显微镜下观察,摄影.该法简便、快速,更适用于叶片较小的植物叶表皮制片.     

栲树叶片SEM样品制备

栲树叶片下表皮覆盖着一层密集的薄壁盾状毛影响了对其叶表面特征的观察。经FAA固定→二甲苯和丙酮的混合液(1∶1)浸泡→恒温金属浴加温→超声波(50～60Hz,90W)振荡器→系列酒精浓度脱水处理后,叶表面薄壁盾状毛脱落,在SEM扫描电子显微镜下可观察到叶表皮形态结构(气孔器,细胞排列及近圆形的毛基)。此方法操作简便、效果明显,并且安全和无污染。     

洋葱表皮撕取方法的改进

“临时装片的观察”和“质壁分离及复原”实验中,撕取的洋葱表皮是否卷曲往往影响实验进程和效果。如果按照课本上介绍的方法撕取,表皮很难展平。我们在实验教学中发现：用口径为０４ｃｍ的打孔器,在洋葱鳞片叶的外表皮上轻轻旋转扎一下,划出一个小圆形,然后用镊子...     

金线莲与台湾金线莲显微结构比较

通过石蜡切片和表皮制片观察比较金线莲Anoectochilus roxburghii和台湾金线莲A.formosanus根、茎、叶的显微结构特征。结果表明,金线莲与台湾金线莲显微结构的主要区别在于根皮层厚度、维管束数目、叶构造、叶脉颜色、表皮乳头细胞的形状和气孔密度,这些特征可作为金线莲物种的显微鉴别依据。     

In vitro indentation to determine the mechanical properties of epidermis

The lack of understanding of the mechanical behavior of the human skin layers makes the development of drug delivery using microneedles or microjets a challenging task. In particular, the key mechanical properties of the epidermis composed of stratum corneum and viable epidermis should be better understood. Micro-indentation experiments were applied, using a spherical tip with a large diameter to the sample thickness ratio. The Young's moduli were derived via an analytical and a numerical method. The tests showed that the analytical method was not appropriate to assess the Young's moduli. That is why a numerical model was used to obtain the correct stiffness. When loaded perpendicularly, the stiffness of both the epidermis and stratum corneum vary between 1 and 2MPa. No significant differences in stiffness between the stratum corneum and viable epidermis were observed.     

A method for flow cytometric cell cycle analysis of normal and psoriatic human epidermis based on a detergent/citric acid technique for suspension of nuclei

A new method is described for flow cytometric cell cycle analysis of normal and psoriatic human epidermis, based on non-enzymatic tissue disaggregation. The epidermis was isolated by treatment with acetic acid and stored by freezing. After thawing, the epidermis was disintegrated into a nuclear suspension by 3 steps: incubation with dithiotreitol, whirling in a buffer (pH 7.4) with the non-ionic detergent Nonidet P40, EGTA, RNase and spermine, and whirling after addition of citric acid to a final concentration of 1% (pH 2.4). The suspension was stained with propidium iodide and filtered before flow cytometry. The yield of suspended nuclei was approximately 70% of the original number of cells in the tissue. The detergent/citric acid method was found to be preferable to an ultrasonication method previously used on human epidermis. All cell cycle and cell maturation stages were represented in the detergent/citric acid suspension, in contrast to the selection of immature G1, S and G2 stages with enzymatic methods. In the analysis of psoriatic epidermis inadequately matured (parakeratotic) cells were present in the suspension and had to be discriminated by gating on light scattering intensity, as they were not susceptible to lysis and did not stain properly. The fraction of S phase nuclei was on average 1.9% in normal and 7.7% in psoriatic epidermis, thus confirming the results of other investigators using enzymes. The presence of mitotic figures in the suspension was demonstrated by flow sorting. In this way the mitotic fraction was estimated to 0.06% in normal and 0.22% in psoriatic epidermis, confirming histological data of other investigators.     

Use of high pressure liquid chromatography for analysis of deoxyribonucleotides in the epidermis

An analytical method for free deoxyribonucleotide (nucleoside monophosphate) in the epidermis is presented. Free nucleotides were extracted from tissue using a methylal ethanol mixture. The deoxyribonucleotides were separated using DEAE-cellulose and DBAE-cellulose. The analysis of free deoxyribonucleotides was carried out by high pressure liquid chromatography on a column of Lichrosorb-NH₂ with a single buffer of potassium phosphate. The elution order of nucleotides was CMP, AMP, UMP, IMP, GMP, and XMP. This procedure employing high pressure liquid chromatography for the detection of deoxyribonucleotides in the epidermis makes it possible to elucidate the biological characteristics and significance of DNA metabolism in normal epidermis and changes that occur in pathological conditions.     

中国五味子属植物叶表皮研究

用浓硫酸－铬酸离析法,在光学显微镜下观察了中国五味子属植物１４种,１亚种,２变种,共２４个样品成熟叶的叶表皮细胞及气孔器特征,结果表明：五味子属植物叶片的上、下表皮细胞呈多边形或不规则形,垂周壁式样为平直,菜或浅汉浪;少数种类上表皮有气孔器或分泌细胞,所有的种类下表皮具气孔器和分泌细胞;气孔器类型以平列型为主,并伴有侧列型,极少数出现不规则型,气孔极区呈稍角质和厚或棒形角质加厚,稀Ｔ形角质加厚,叶     

A Rapid,Highly Efficient and Economical Method of Agrobacterium-Mediated In planta Transient Transformation in Living Onion Epidermis

Transient transformation is simpler, more efficient and economical in analyzing protein subcellular localization than stable transformation. Fluorescent fusion proteins were often used in transient transformation to follow the in vivo behavior of proteins. Onion epidermis, which has large, living and transparent cells in a monolayer, is suitable to visualize fluorescent fusion proteins. The often used transient transformation methods included particle bombardment, protoplast transfection and Agrobacterium-mediated transformation. Particle bombardment in onion epidermis was successfully established, however, it was expensive, biolistic equipment dependent and with low transformation efficiency. We developed a highly efficient in planta transient transformation method in onion epidermis by using a special agroinfiltration method, which could be fulfilled within 5 days from the pretreatment of onion bulb to the best time-point for analyzing gene expression. The transformation conditions were optimized to achieve 43.87% transformation efficiency in living onion epidermis. The developed method has advantages in cost, time-consuming, equipment dependency and transformation efficiency in contrast with those methods of particle bombardment in onion epidermal cells, protoplast transfection and Agrobacterium-mediated transient transformation in leaf epidermal cells of other plants. It will facilitate the analysis of protein subcellular localization on a large scale.     

A generalized method for transfecting root epidermis uncovers endosomal dynamics in Arabidopsis root hairs

Progress in analysing the cellular functions of many structural proteins has accelerated through the use of confocal microscopy together with transient gene expression. Several methods for transient expression have been developed in the past few years, but their application has seen limited success beyond a few tractable species and tissues. We have developed a simple and efficient method to visualize fluorescent proteins in Arabidopsis root epidermis using co-cultivation of seedlings with Agrobacterium rhizogenes. The method is equally suitable for transient gene expression in other species, including Thellungiella, and can be combined with supporting molecular and biochemical analyses. The method promises significant advantages for study of membrane dynamics, cellular development and polar growth in root hairs without interference in the development of the plant. Since the method targets specifically the root epidermis, it also offers a powerful tool to approach issues of root-rhizosphere interactions, such as ion transport and nutrient acquisition. As a proof of principle, we carried out transfections with fluorescent markers for the plasma membrane (NpPMA2-GFP, Nicotiana plumbaginifolia L. Plasma Membrane H(+)-ATPase 2), the endoplasmic reticulum (YFP-HDEL), and the Golgi apparatus (sialyl transferase-GFP) to trace their distribution in growing Arabidopsis root hairs and epidermis. The results demonstrate that, in Arabidopsis root hairs, movement of the Golgi is faster than previously reported for tobacco leaf epidermal cells, consistent with the high secretory dynamics of the tip growing cell; they show a pattern to the endoplasmic reticulum within the cytoplasm that is more diffuse than found in tobacco leaf epidermis, and they confirm previous findings of a polarized distribution of the endoplasmic reticulum at the tip of growing root hairs.     

沙棘雌雄株叶片解剖结构比较研究

为了研究沙棘雌、雄株叶片的第二性征,本文采用石蜡切片法观察了沙棘雌、雄株叶片结构的差异。结果表明：（1）沙棘雌、雄株叶片均由表皮、叶肉和叶脉3部分组成,表皮均由1层细胞构成,表皮毛发达,上表皮有拟泡状细胞;叶肉栅栏组织与海绵组织分化明显。（2）雌株上表皮具更多的拟泡状细胞,其主脉韧皮部薄壁细胞及其下方的一些薄壁细胞含较多的后含物,下表皮的表皮毛更浓密;而雄株的叶片厚度、叶片上表皮厚度、栅栏组织厚度、栅栏组织厚度/海绵组织厚度均显著大于雌株,且其主脉维管束更发达。结果表明,沙棘雌雄株叶片解剖结构存在明显差异,这些差异是第二性征的表现,也是沙棘长期进化中形成的稳健的适应策略,可能有利于该物种的繁衍。     

Epidermal cell kinetics of the pig: a review

Age-related changes in cell kinetic parameters for the epidermis of pigs have been shown to be small, indicating that young pigs may be used for experimental studies. It was not possible to draw any firm conclusions about any strain-related differences in the cell kinetics of the epidermis of the pig. Lower LI values have been quoted for the miniature pig and the Yorkshire pig than for the Large White pig. However, these differences may be related to variations in experimental technique. The cell kinetic data for the Yorkshire pig are not consistent. Very high values for the mitotic index suggested a high rate of cell turnover, whilst data from single pulse labelling and grain count halving studies indicate a relatively low rate of cell turnover. The results from continuous labelling studies on the epidermis of the Yorkshire pig suggest that the basal cell turnover time (TT) is a factor of two or more shorter (136 h) than the estimates obtained using other methods. In the Large White pig estimates of TT were similar using a variety of techniques and were comparable with the TT estimate for the Yorkshire pig obtained using the continuous labelling method. There is some degree of inconsistency in the literature with regard to possible diurnal variations in the cell kinetic parameters for the epidermis. In the study of Archambeau & Bennet (1984) distinct diurnal variations were found in the LI, although the reliability of this finding is questionable due to the small number of animals used. Later studies by Morris et al. (1987) have suggested that diurnal variations are negligible in the epidermis of the pig. The majority of labelled cells (80%) in pig epidermis are located in the basal layer, although a significant proportion (20%) occurs suprabasally, in the cell layer immediately above the basal layer. Therefore, the epidermis can be regarded as having a bilayered proliferative cell compartment. The results from studies on irradiated pig skin (Morris & Hopewell, 1986, 1988, 1989) are not consistent with the presence of a homogeneous proliferative compartment in the epidermis, and are best explained by the occurrence of an heterogeneous proliferative compartment consisting of a stem cell subpopulation and a much larger population of transit proliferative cells.