• Applied Microscopy
• /
• 제16권2호
• /
• pp.1-13
• /
• 1986

The purpose of this study was to investigate the morphology and intercellular junctions of the odontoblast of dental pulp in the rat incisor by means of the freeze fracture electron microscopy. Twenty male Sprague-Dawley rats weighing $150{\sim}200g$ were used. After being anesthetized by an intraperitoneal injection of 0.5 ml sodium pentobarbital per kg in body weight(60 mg/ml) the animals were perfused with 2.5% glutaraldehyde-2% paraformaldehyde fixative in 0.1 M cacodylate buffer, pH 7.2 through the ascending aorta for one hour. The incisors were carefully extracted from the jaws and demineralized by suspending them in 0.1 M EDTA in 3% glutaraldehyde (pH 7.2) for two weeks. After demineralization, the specimens were obtained from the portion divided into five equal parts. For freeze-fracture replication, demineralized tissues were infiltrated for several hours with 10%, 25% glycerol in 0.1M cacodylate buffer as a cryoprotectant and then frozen in liquid Freon 22 and stored in liquid nitrogen. Fracturing and replication were done in Balzers BAF 400D high-vacuum freeze-fracture apparatus at $-120^{\circ}C$ under routine $5X10^{-7}$ Torr vacuum. The tissue was immediately replicated with platinum unidirectionally at $45^{\circ}$ angle and reinforced with carbon at $90^{\circ}$ angle unidirectionally or by using a rotary stage. The replication process was monitored by a quartz-crystal device. The replicas were immersed in 100% methanol overnight. The tissue was then digested from the replica by clorox (laundry bleach), placed into 5% EDTA, and washed repeatedly with distilled water. The replicas were picked up on 0.3% formvar-coated 75 mesh grids and examined in the JEOL 100B electron microscope. The results were as follows; 1. Both in thin sections and freeze-fracture replicas, three types of intercellular junctions were recognizable in the plasma membrane of odontoblast: gap junction, tight junction and desmosome-like junction. 2. The nuclear pores were evenly distributed over the nuclear envelope. The pore complex formed a ring about 70 nm in diameter. 3. Gap junctions were found between odontoblasts as well as odontoblasts and neighbouring pulp cells (fibroblast, subodontoblastic cell process, nerve-like fibre). Gap junctions, which were round, ellipsoid and pear-shaped and 600 nm in diameter, were observed in the odontoblast. 4. Numerous round and ellipsoid gap junctions could be frequently seen on the plasma membranes in cell body and apical part of the odontoblasts. On the P face, the junctions were recognized as a cluster of closely packed particles, measuring about 9 nm in diameter, and on the E face, the junctions were recognized as a shallow grooves.

• 미생물학회지
• /
• 제27권2호
• /
• pp.117-123
• /
• 1989

Radioactive NBZ arginyl diazomethane으로 액포를 표지 한 뒤에ㅣ 액포내의 단백질과 세포막 겉에 존재하는 단백질을 각각 저농도 완충용액과 높은 농도의 염용액으로 제거시켰다. 액포막 단백질을 Triton X-100으로 녹인 후, molecular sieve column chromatography와 ion exchange column chromatography를 사용하여 anginine transporter를 분리하였다.

• Applied Microscopy
• /
• 제25권2호
• /
• pp.53-64
• /
• 1995

세포막의 결합장치들은 형태학적으로나 기능적으로 분화된 특이한 구조이며 각기 인접한 세포막 사이의 특징적인 부위에 위치한다. 그러나 이들 결합장치가 간세포의 기능상태에 연관하여 미세구조적으로 어떠한 변화를 보이는지에 관해서는 아직 잘 알려져 있지 않다. 본 연구는 간세포의 기능적 변동에 따르는 간세포막 사이 결합장치의 형태학적 변화를 알아보기 위하여 정상군과 8일간 기아 시킨 기아군으로 구분한 휜쥐 ($Wister,\;{\uparrow},\;250{\sim}280g$)에서 절취된 간조직을 박절편과 급속동결할 단복제법에 의거 표본을 제작하여 이들 결합장치의 미세구조상을 비교 고찰하였다. 그 결과 담세관주위의 폐쇄띠는 기아상태에서 현저히 감축되거나 소실되어 있었는데 이러한 소견은 박절편과 급속동결할단복제에서 다 같이 관찰되었으며 기아상태하에서 결합반과 교통반의 현저한 증가를 볼 수 있었다. 또한 정상상태하에서 이들 결합장치가 비교적 일정한 구조를 유지하고 있을 것이라는 일반적인 견해와는 달리 본 연구에서는 박절편이나 급속동결할단복제에서 다 같이 폐쇄띠가 부위에 따라 현저한 구조적 차이를 보였다. 본 연구에서 얻어진 이러한 결과로 미루어 폐쇄띠의 감축이나 소실은 기아로 인한 간세포의 기능저하가 담즙물질의 생성을 저해하게 됨으로서 담세관의 내압이 저하된 데 기인된 현상으로 생각된다. 또한 결합반의 증가는 기아로 인하여 감축된 간세포를 지지하므로서 간조직의 유지를 도우려는데 있었을 것이며 교통반의 증가는 기아로 인한 간세포의 기능적 저하가 간세포상호간의 물질교환을 필요로 하였기 때문이었을 것으로 추정된다. 특히 정상상태하에서도 부위에 따라 폐쇄띠의 미세구조적 변화가 있었던 것은 이 결합장치가 간세포의 기능이나 담세관의 상태에 민감하게 반응하기 때문일 것으로 생각된다.

• Applied Microscopy
• /
• 제42권2호
• /
• pp.111-114
• /
• 2012

The scanning electron microscopy is an ideal technique for examining plant surface at high resolution. Most hydrate samples, however, must be fix and dehydrate for observation in the scanning electron microscope. Because the microscopes operate under high vacuum, most specimens, especially biological samples, cannot withstand water removal by the vacuum system without morphological distortion. Cryo-techniques can observe in their original morphology and structure without various artifacts from conventional sample preparation. Rapid cooling is the method of choice for preparing plant samples for scanning electron microscopy in a defined physiological state. As one of cryo-technique, high-pressure freezing allows for fixation of native non-pretreated samples up to $200{\mu}M$ thick and 2 mm wide with minimal or no ice crystal damage for the freezing procedure. In this study, we could design to optimize structural preservation and imaging by comparing cryo-SEM and convention SEM preparation, and observe a fine, well preserved Arabidopsis stem's inner ultrastructure using HPF and cryo-SEM. These results would suggest a useful method of cryo-preparation and cryo-SEM for plant tissues, especially intratubule and vacuole rich structure.

• 한국물환경학회지
• /
• 제34권1호
• /
• pp.46-56
• /
• 2018

A mcyB-specific Ultra-Rapid quantitative PCR was developed for the quantitative detection of Microcystis aeruginosa, which is often a dominant species in green tide. McyB-specific UR-qPCR was optimized under extremely short times of each step in thermal cycles, based on the specific primers deduced from the mcyB in microcystin synthetase of M. aeruginosa. The M. aeruginosa strain KG07 was used as a standard for quantification, after the microscopic counting and calculation by mcyB-specific UR-qPCR. The water samples from the river water with the Microcystis outbreak were also measured by using both methods. The $1.0{\times}10^8$ molecules of mcyB-specific DNA was recognized inner 4 minutes after beginning of UR-qPCR, while $1.0{\times}10^4$ molecules of mcyB-specific templates was detected inner 7 minutes with quantitative manner. From the range of $1.0{\times}10^2$ to $1.0{\times}10^8$ initial molecules, quantification was well established based on $C_T$ using mcyB-specific UR-qPCR (Regression coefficiency, $R^2=0.9977$). Between the numbers of M. aeruginosa cell counting under microscope and calculated numbers using mcyB-specific UR-qPCR, some differences were often found. The reasons for these differences were discussed; therefore, easy compensation method was proposed that was dependent on the numbers of the cell counting. Additionally, to easily extract the genomic DNA (gDNA) from the samples, a freeze-fracturing of water-sample using liquid nitrogen was tested, by excluding the conventional gDNA extraction method. It was also verified that there were no significant differences using the UR-qPCR with both gDNAs. In conclusion, the mcyB-specific UR-qPCR that we proposed would be expected to be a useful tool for rapid quantification and easy monitoring of M. aeruginosa in environmental water.