• Korean Journal of Microbiology
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• v.37 no.1
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• pp.61-65
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• 2001

Epifluorescence microscopy was used to observe epiphytic bacteria directly on plant leaf surfaces as well as indirectly in the leaf liberating solution by staining with fluorochromes of 4',6-diamidino-2-phenylindole (DAPI) and acridine orange(AO). Epiphytic bacteria could not be well observed on the leaf surface by staining with AO due to an intrusive orange or red background fluorescence. However, DAPI gave us clear epifluorescent images of the bacteria on the leaf. On the contrary, epiphytic bacteria in the liberating leaf solution were well observed on filters stained by both types of fluorochrome, although DAPI showed better fluorescent images than AO and not necessarily required a washing step of the filters stained. The optimum conditions of the DAPI stains were 5 $\mu$g/ml for 5 min both for leaves and for filters of the liberating solution. It was confirmed that a critical step in the epifluorescence microscopy of leaf surfaces was to minimize release of water from the leaf. For this, the stained leaf samples were put on a filter paper, kept in a dry oven at $70^{\circ}C$ for 2 min instead of air-drying, and then immediately observed by epifluorescence microscopy. The established technique was applied to enumerate epiphytic bacteria on oak tree leaf surfaces.

• Journal of Dental Rehabilitation and Applied Science
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• v.31 no.3
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• pp.203-211
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• 2015

Purpose: The purpose of this study was to investigate the reosseointegration periods when the rough surface implants, which had complete bone-implant ankylosis, suddenly losed the osseointegration. Materials and Methods: The implants with RBM surface treatment were inserted into both tibias of 23 rabbits. Two implants were submerged into each side. After six weeks, the primary removal torque was measured by Digital torque gauge, and then the implants were replaced and submerged to estimate the level of reosseointegration. After assigned healing periods for each group, the removal torque was measured again. BIC (Bone-Implant contact, %) ratio was measured through histomorphometric analysis.Paired t-test was processed by SPSS 14.0. One-way ANOVA and Tukey's post-hoc test was processed to analyze statistically significant differences among the groups. Results: In comparison with the primary removal torque, the secondary removal torque was increased after 11 days and significantly increased from 2 weeks. In fluorochrome labeling, the origin of mineralization was observed after 7 days, which showed as fluorescent bands around the bone-implant interfaces. After 11 days, the bone formation was apparent, and it is increased continuously with the passage of the time. Conclusion: In 11 days after the implant replacement, the secondary removal torque was almost as same as the primary value, and was significantly higher from 2 weeks. The mineralized shapes were observed in 7 days after the implant replacement, and then the bone formation appeared visibly in 11 days.

• Clinical and Experimental Reproductive Medicine
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• v.24 no.3
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• pp.335-340
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• 1997

Fluorescence in situ hybridization (FISH) techniques allow the enumeration of chromosome abnormalities and from a great potential for many clinical applications. In order to produce quantitative and reproducible results, expensive tools such as a cooled CCD camera and a computer software are required. We have developed a Chromosome Image Processing System (Chips) using FISH that allows the detection and mapping of the genetic aberrations. The aim of our study, therefore, is to evaluate the capabilities of our original system using a black-and-white video camera. As a model system, three repetitive DNA probes (D18Z1, DXZ1, and DYZ3) were hybridized to variety different clinical samples such as human metaphase spreads and interphase nuclei obtained from uncultured peripheral blood lymphocytes, uncultured amniocytes, and germ cells. The visualization of the FISH signals was performed using our system for image acquisition and pseudocoloring. FISH images were obtained by combining images from each of probes and DAPI counterstain captured separately. Using our original system, the aberrations of single or multiple chromosomes in a single hybridization experiment using chromosomes and interphase nuclei from a variety of cell types, including lymphocytes, amniocytes, sperm, and biopsied blastomeres, were enabled to evaluate. There were no differences in the image quality in accordance with FISH method, fluorochrome types, or different clinical samples. Always bright signals were detected using our system. Our system also yielded constant results. Our Chips would permit a level of performance of FISH analysis on metaphase chromosomes and interphase nuclei with unparalleled capabilities. Thus, it would be useful for clinical purposes.

• Clinical and Experimental Reproductive Medicine
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• v.23 no.1
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• pp.25-32
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• 1996

This work has been carried out to examine the number of Total, ICM and TE cells of F1 mouse blastcysts at day 4 after IVF by differential labelling of the nuclei with polynucleotide-specific fluorochromes and to obtain a fundamental information of preimplantation mouse embryo development. Blastocysts produced by superovulated B6CBA F1(C57BL/${\times}$CBA) eggs were inseminated with $1{\times}10^6$spermatozoa/ml and cultured in M16 medium at $37^{\circ}C$, 5% $CO_2$ incubator for 95hrs. Blastocysts were classified as early, middle, expanded and hatching stage according to the developmental morphology; blastocoel expansion and zona thickness. The results obtained in these experiments were summarized as follows; 1) The development rate of blastocysts at 95hrs after IVF was 86.7% and classified blastocysts to early, middle, expanded and hatching were 16.3%, 18.9%, 10.5% and 40.9%, respectively. 2) The numbers of total blastomere using bisbenzimide in the classified blastocysts to early, middle, expanded and hatching were 35.6${\pm}$1O.4, 49.4${\pm}$8.6, 60.8${\pm}$1O.7 and 62.7${\pm}$13.9, respectively. 3) In ICM and TE cell number by using differential labelling with polynucleotide-specific fluorochrome in the classified blastocysts to early, middle, expanded and hatching; ICM numbers were 9.6${\pm}$3.0, 13.6${\pm}$3.9, 16.0${\pm}$3.3 and 19.5${\pm}$4.6, respectively and TE cell numbers were 30.6${\pm}$5.1, 39.9${\pm}$5.8, 42.2${\pm}$8.1 and 43.7${\pm}$11.1, respectively. These results showed the same increase pattern according to development advance level. Also, when compared with the results of total count were obtained between bisbenzimide only and differential labelling, both of them showed the same increase pattern according to development level and at the same time their cell numbers were almost the same. So, rapid and simple cell count method using differential labelling can be used for the examination of later preimplantation development or as an indicator of embryo quality according to the variables of culture conditions.