激光共热扫描显微镜的光学层析特性研究

本文从理论上研究了激光共焦扫描显微镜的光学层析特性,并给出了探测器的针孔大小,聚焦物镜数值孔径与光学层析的关系,最后还给出了利用光学层技术发现的一种寄生虫新的形态结构的实验结果。     

光学相干层析成像技术用于裸鼠皮肤霉菌感染研究

利用中心波长为850 nm的宽带光源SLD实现了纵向分辨率为8μm的光学相干层析成像系统。系统采用傅里叶域光学延迟线实现了深度扫描速度为160 mm/s,成像深度为3 mm。获得了裸鼠皮肤霉菌感染部位和健康皮肤的光学相干层析(optical coherence tom ography,OCT)图像,皮肤病变前后的内部结构信息清晰可见。     

光学相干层析分子成像方法初探

介绍了分子对比剂在光学相干层析成像(optical coherence tom ography,OCT)技术中的研究现状,概述了迄今出现的几种不同的光学相干层析分子成像方法(m olecu lar contrast OCT,简称为MCOCT),讨论了MCOCT的几个重要的实际问题:对比剂的选择范围、激发光强的限制、各种方法灵敏度比较以及MCOCT应用于临床与生物学领域需要考虑的因素。     

基于光学相干层析图像的离体牙三维重建

利用shear-warp算法对离体牙的光学相干层析图像进行三维重建,通过不透明度传递函数的合理设置及光照模型的引入实现牙齿内部组织结构的可视化,便于医生在早期龋齿诊断中定位病变.介绍了shear-warp算法的原理、用于龋齿检测的全光纤光学相干层析成像系统及其二维层析图,以及利用离体牙牙冠的二维层析图重建获得三维结构图.     

人手部皮下微循环特性随深度变化的研究

本研究使用手持式扫频光学相干层析(SSOCT)系统分别在人左手无名指指甲上方、无名指指腹和手掌大拇指根部选取3 mm×3 mm区域进行了在体成像,使用指数补偿方法还原血流信息,获取不同深度的血流正面光学断层图像(en face图像),从血管灌注密度(PD)、血管直径(VD)两个方面分析了微循环随深度的变化特性。试验结果证明,在上述三个成像位置中,随着成像深度的增加,血管PD与血管VD均增大,血流信息的丰富程度增加,但同时en face图像的清晰度有所下降。其中左掌大拇指根部微血管最为细密,血流信息较为丰富,比较适宜作为微循环疾病诊断研究的成像部位。分层投影结果显示,左手无名指指甲上方部位在成像深度较浅(220～660μm)时,血管PD明显小于其他两个成像部位,说明此处在该范围内血流信息丰富程度不够。本文测量了随深度变化的手部血流几何学特征,验证了光学相干层析(OCT)技术基于手部皮下微循环光学断层图像进行疾病诊断的可行性。许多微循环疾病早期就已经产生了微血管病变,因此对这种微循环变化的探测在疾病的早期诊断方面具有临床指导意义。     

基于全场光学相干层析图像的人体肝组织癌变定量特征分析研究

利用实验室搭建的全场光学相干层析(FFOCT)系统对人体正常与癌变肝组织分别进行成像,获得了高分辨率的层析图像。针对人眼识别FFOCT图像时难以区分正常与早期癌变肝组织的问题,提取两者FFOCT图像的灰度特征并进行统计学分析,以实现在参数特征上区分两者的目的。结果表明,在正常与癌变肝组织初步提取的均值、方差和峰度等五个特征参数中,有四个显示出了明显的数值差异,可用于鉴别肝组织是否癌变。该研究结果为进一步研究如何利用FFOCT图像中所提取的特征参数作为判断实际临床诊断中生物组织癌变与否的诊断标准打下了基础。     

倒置双光子显微镜的使用与维护

双光子激发荧光显微是一种非线性光学显微技术,它结合了激光扫描共聚焦显微镜和双光子激发技术,具有高时空分辨率、高信噪比和固有的三维层析分辨能力等优点。介绍了双光子显微镜的软硬件组成和技术参数,样品制备方法,双光子图像采集的常用操作规程,日常维护和仪器管理等方面。旨在为双光子仪器的使用者与管理者提供参考,使之更好地服务于教学与科研。     

高级脂肪酸的纸层析

关于高级脂肪酸的纸层析,国内外已有很多报道。我们从1972年起陆续作了一些工作。本文准备结合自己的体会,介绍这方面的技术。一、反相层析滤纸的制备纸层析是分配层析。分配层析的原理是利用物质在固定相和流动相之间的分配达到分离的目的。     

层析法纯化破伤风毒素的试验研究

为了获得高纯度的破伤风毒素,用疏水层析和离子交换层析纯化破伤风毒素。破伤风毒素培养滤液经Phenyl Sepharose疏水层析除去大部分杂质,再经DEAE Sephadex离子交换层析进一步纯化。经两步层析纯化后,毒素纯度达到2000Lf/mg PN以上,回收率为52%~73%。用此方法,连续纯化五批毒素,均获得高纯度的破伤风毒素。试验证明破伤风毒素经疏水层析和离子交换层析可得到有效纯化。     

一种改进的羟基磷灰石层析法分离DNA的恒温加热装置

本文设计了一种控温准确,操作简便的加热装置,提高了羟基磷灰石层析的分辨能力,并研究采用此装置在不同温度条件下的层析效果。实验证明,用羟基磷灰石层析法分离单、双链DNA,60℃时分离效果最佳,65℃以上则严重影响层析结果,层析温度应控制在60℃±5℃。     

成熟玉米秆组织结构切片方法的比较研究

于玉米成熟期选择健壮茎秆的基部第三节间为材料,采用徒手切片法、冰冻切片法、石蜡切片法、薄切片法等4种方法,比较不同方法的玉米茎秆组织结构切片质量,为研究玉米茎秆结构与其倒伏的关系奠定技术基础。结果表明:徒手切片法是获得成熟玉米秆组织结构切片较为方便、快速的方法,切片面积较大,适合大范围观察统计;冰冻切片法是获得成熟玉米秆组织结构较快的方法,切片面积较小,适合小范围观察;薄切片是获得高质量成熟玉米秆组织结构切片的最好方法,但切片面积太小,适合高倍数显微观察和小范围电镜扫描观察组织结构;石蜡切片不适合作为成熟玉米秆的组织切片方法。研究认为,徒手切片法是最适合成熟玉米秆组织结构观察研究的制片方法。     

Optical sectioning microscopy with planar or structured illumination

A key requirement for performing three-dimensional (3D) imaging using optical microscopes is that they be capable of optical sectioning by distinguishing in-focus signal from out-of-focus background. Common techniques for fluorescence optical sectioning are confocal laser scanning microscopy and two-photon microscopy. But there is increasing interest in alternative optical sectioning techniques, particularly for applications involving high speeds, large fields of view or long-term imaging. In this Review, I examine two such techniques, based on planar illumination or structured illumination. The goal is to describe the advantages and disadvantages of these techniques.     

The effects of superior ovarian nerve sectioning on ovulation in the guinea pig

The effects on spontaneous ovulation associated with the unilateral or bilateral sectioning of the superior ovarian nerves (SON) were analyzed in guinea pigs at different time intervals of the estrous cycle. Day 1 of the estrous cycle was defined as the day when the animal presents complete loss of the vaginal membrane (open vagina). Subsequent phases of the cycle were determined by counting the days after Day 1. All animals were autopsied on the fifth day of the estrous cycle after surgery. Sectioning the right, left, or both SONs on day 5 (early luteal phase) resulted in a significant increase in the number of fresh corpora lutea. Ovulation increased significantly when the left SON (L-SON) was sectioned during late follicular phase (day 1) and medium luteal phase (day 8). When surgery was performed on days 1 or 8, neither sectioning the right SON (R-SON) nor sectioning the SON bilaterally had an apparent effect on ovulation rates. Similarly, ovulation rates were not affected when unilateral (right or left) or bilateral sectioning of the SON was performed during late luteal phase two (day 12). Unilateral or bilateral sectioning of the SON performed during the early luteal phase (day 5) was associated with a significant decrease in uterine weight. A comparable effect was observed when the L-SON was sectioned during late follicular phase (day 1), or medium luteal phase (day 8). No effects on uterine weight were observed when unilateral or bilateral sectioning of the SON was performed during late luteal phase. Our results suggest that in the guinea pig the SON modulates ovulation, and that the degree of modulation varies along the estrous cycle. The strongest influence of the SONs on ovulation occurs during early luteal phase, and decrease thereafter, being absent by late luteal phase. In addition, sectioning the left or the right SON caused different responses by the ovaries of adult guinea pigs. This paper discusses the mechanisms by which ovulation increased when the SON was surgically cut.     

Activity of caudal mesenteric ganglion in relation to its impulsation (age aspect)

It has been shown in acute experiments on cats and kittens that the pre- and postganglionic nerves of the caudal mesenteric sympathetic ganglion (CMSG) are bioelectrically active from the first day of the animal's life. When the impulsation which comes to the ganglion along the afferent and preganglionic fibers is removed, the impulse activity of the post-ganglionic neurons disappears completely. In newborn kittens the level of activity of the postganglionic fibers is determined to a greater degree by the afferent impulsation of the hypogastric nerves than in adult cats. The effect of the preganglionic impulsation, especially that coming through the intramesenteric nerves, predominates in adult animals. The efferent impulsation of the hypogastric nerves in cats not older than five days disappeared after sectioning of two to three preganglionic branches of the CMSG; in six to nine and 14 to 15-day-old cats, after sectioning of four and in month old cats after sectioning of five; and in two to three month old cats after sectioning of six branches. In adult cats after the first sectioning the postganglionic impulsation frequently intensified, and after sectioning of seven branches the activity completely disappeared. Automatic activity of the ganglia was not observed, postganglionic impulsation developed and changed under the influence of the impulses coming to the ganglion.Scientific Research Institute of Childhood and Adolescent Physiology, Academy of Pedagogical Sciences USSR, Moscow. Translated from Neirofiziologiya, Vol. 3, No. 5, pp. 533–542, September–October, 1971.     

麻疯树种子发育过程中脂类物质变化的显微和超微结构观察

用石蜡切片、半薄切片和超薄切片方法研究了麻疯树种子发育过程中脂类物质的变化。结果表明:脂类物质主要储存于胚乳当中,当种子发育成熟时胚乳中迅速积累了大量的油脂;种皮在发育过程中脂类物质含量较多,成熟时外种皮硬化,内种皮含有大量的油脂;种子成熟时胚中含有少量的脂类物质。细胞内脂类物质含量比较多时,内质网、线粒体、质体和高尔基体数量也会较多。种子完全成熟时进行采收加工是最为合适的。     

Optical sectioning microscopy

Confocal scanning microscopy, a form of optical sectioning microscopy, has radically transformed optical imaging in biology. These devices provide a powerful means to eliminate from images the background caused by out-of-focus light and scatter. Confocal techniques can also improve the resolution of a light microscope image beyond what is achievable with widefield fluorescence microscopy. The quality of the images obtained, however, depends on the user's familiarity with the optical and fluorescence concepts that underlie this approach. We describe the core concepts of confocal microscopes and important variables that adversely affect confocal images. We also discuss data-processing methods for confocal microscopy and computational optical sectioning techniques that can perform optical sectioning without a confocal microscope.     

Precision Sectioning of Wood

In discussing the theories and mechanism of sectioning, it is pointed out that virtually no concrete knowledge exists on the subject. The function of different mechanisms and their role in cutting different types of tissue, and the value of microtome knives versus safety razor blades is discussed. Razor blades failed to cut precise sections; a technic for sharpening a knife to give precise sections is outlined, pointing out errors to avoid in sharpening and sectioning. Various types of knife edges are illustrated by photomicrographs. The procedure of sharpening and sectioning technics for critical results is explained. The mechanism of cutting in wood appeared to be of the crushing and tearing type, indicating the necessity of final polishing in sharpening.     

New embedding medium for sectioning undecalcified bone.

Methylmethacrylate (MMA) is the most commonly used embedding medium for sectioning undecalcified bone; however, a number of problems exist with its use in a research laboratory. MMA requires a long infiltration time and temperature control, and it reacts with many polymers. We used Kleer Set resin as an alternative embedding medium for sectioning undecalcified bone specimens. Fluorochrome labeled bone specimens were sectioned transversely using a ground section technique and longitudinally on a sledge macrotome. The slides were viewed using both transmitted light and epifluorescence microscopy. High quality sections were obtained using Kleer Set resin for both sectioning techniques. We have shown that this new embedding medium is simpler, safer, quicker to use and does not interfere with visualization of fluorochromes.     

Correction of distortion of histologic sections of arteries

Histologic sections of arteries can be used to generate three-dimensional (3D) geometric models and identify structural constituents. However, geometric distortions are introduced by fixation, embedding and sectioning; distortions which can, for example, lead to errors in stresses predicted by finite element models. We developed a method to measure and correct for distortions caused by acrylic processing and applied it to intact, healthy porcine coronary arteries. Micro-computed tomography was used to image arteries in the fresh and embedded states. Tissue blocks were sectioned, stained and imaged using a light microscope. Each section contained four registration marks used to determine strains introduced by sectioning and staining. Using these three image sets, 3D geometric models were generated and distortions were measured. Fixation, processing, and embedding resulted in shrinkage of 6.4+/-2.3% axially and 35.4+/-5.0% in mean cross-sectional area (n=5). Shrinkage in a cross section was well characterized by a uniform, equibiaxial strain. Sectioning and staining resulted in additional compressive strains in the sectioning direction of 0.067+/-0.011 and, in the direction perpendicular to sectioning, of 0.023+/-0.005 (n=5). These strains are assumed uniform and form the basis for correcting section geometry. Reconstructions using corrections for sectioning and shrinkage-related distortions had errors of 1.6+/-0.5% (n=5) and 4.0+/-1.7% (n=5), respectively.     

New embedding medium for sectioning undecalcified bone

Methylmethacrylate (MMA) is the most commonly used embedding medium for sectioning undecalcified bone; however, a number of problems exist with its use in a research laboratory. MMA requires a long infiltration time and temperature control, and it reacts with many polymers. We used Kleer Set resin? as an alternative embedding medium for sectioning undecalcified bone specimens. Fluorochrome labeled bone specimens were sectioned transversely using a ground section technique and longitudinally on a sledge macrotome. The slides were viewed using both transmitted light and epifluorescence microscopy. High quality sections were obtained using Kleer Set resin? for both sectioning techniques. We have shown that this new embedding medium is simpler, safer, quicker to use and does not interfere with visualization of fluorochromes.