• Title/Summary/Keyword: Cell counting

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Culturing the Uncultured in the Ocean

  • Cho, Jang-Cheon
    • Proceedings of the Microbiological Society of Korea Conference
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    • 2005.05a
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    • pp.28-32
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    • 2005
  • Epifluorescence microscopy and direct viable counting methods have shown that only 0.01-0.1% of all the microbial cells from marine environments form colonies on standard agar plates. To culture novel marine microorganisms, high throughput culturing (HTC) techniques were developed to isolate cells in very low nutrient media. This approaches was designed to address microbial metabolic precesses that occur at natural substrate concentrations and cell densities, which are typically about three orders of magnitude less than in common laboratory media. Approximately 5000 cultures of pelagic marine bacteria were examined over the course of 3 years. Up to 14% of cells from coastal seawater were cultured using this method, a number that is 1400 to 140-fold higher than obtained by traditional microbiological culturing techniques. Among the cultured organisms are many unique phylogenetic lineages that have been named as new phyla (7), orders (2, 5, 12), families (3), and genera (1, 4, 6). Over 90% of the cells recovered by this method do not replicate in standard agar plating, the most common method of microbial cell cultivation.

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A Compensation Mechanism of Cell Delay Variation for the Satellite TDMA/ATM Network (위성 TDMA/ATM망에서의 셀 지연 변이 보상기법)

  • 정하재;오창석
    • Journal of the Korea Institute of Information and Communication Engineering
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    • v.4 no.2
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    • pp.323-336
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    • 2000
  • In order to achieve the deployment of B-ISDN services in a short time, B-ISDN is being combined with terrestrial ATM network and satellite network which covers a widespread service range. This paper analyzes and investigates several previously existent CDV compensation methods in order to compensate CDV arising from interfacing satellite TDMA and ATM. Specifically to supplement the problems of timestamp and cell number counting methods, new Partial Timestamp mechanism for CDV compensation is proposed. The CDV compensating efficiency of the proposed mechanism is analyzed and evaluated by simulations. It goes to show that Partial timestamps mechanism has many advantages in the aspect of the CDV compensation.

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The Sister Chromatid Exchange Frequencies and Chromosome Aberrations in Mouse Lymphocyte by X-Ray Irradiation (X-선 조사에 의한 마우스 淋巴球의 SCE 경도와 염색체이상)

  • 황인담;기노석;이정상;김남송;이재형;이준배
    • Journal of Environmental Health Sciences
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    • v.16 no.1
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    • pp.55-65
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    • 1990
  • This study was carried out to investigate the effects on sister chromatid exchanges (SCEs) and chromosome aberrations in PHA or LPS stimulated mouse spleen and bone marrow lymphocytes after an acute whole body irradiation. Frequencies of sister chromatid exchanges were significantly increased with the increased dose(from zero to 400tad) but there was no differences between B-cell and T-cell. By times, the maximum induced SCE levels was observed at 12 hours after irradiation and then returned to base level at one day in 100rad group and three day in 400rad group. There was a significant difference in chromosome aberration with increasing exposure. X-ray irradiated chromosome aberration was long lived relative to SCE. This results show that counting the incidence of SCE may not provide a sensitive system for detecting X-ray exposure.

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White Blood Cell Types Classification Using Deep Learning Models

  • Bagido, Rufaidah Ali;Alzahrani, Manar;Arif, Muhammad
    • International Journal of Computer Science & Network Security
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    • v.21 no.9
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    • pp.223-229
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    • 2021
  • Classification of different blood cell types is an essential task for human's medical treatment. The white blood cells have different types of cells. Counting total White Blood Cells (WBC) and differential of the WBC types are required by the physicians to diagnose the disease correctly. This paper used transfer learning methods to the pre-trained deep learning models to classify different WBCs. The best pre-trained model was Inception ResNetV2 with Adam optimizer that produced classification accuracy of 98.4% for the dataset comprising four types of WBCs.

The Effects of Saengkankunbi-tang on Proliferation, Apoptosis and Cell Signaling Pathways of HepG2 Cells (생간건비탕(生肝健脾湯)이 HepG2 cell의 증식, 세포사멸 및 활성조절 신호전달계에 미치는 영향)

  • Kim, Jae-Yong;Kim, Young-Chul;Lee, Jang-Hoon;Woo, Hong-Jung
    • The Journal of Internal Korean Medicine
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    • v.27 no.1
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    • pp.149-165
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    • 2006
  • Objectives: This study was done to evaluate the effects of Saengkankunbi-tang on cell-viability, proliferation, cell-cycle, apoptosis and DNA replication on HepG2 cell and to find out by which molecular-biological mechanism by which Saengkankunbi-tang operates. Methods : The MTT assay, cell counting assay, [3H]-thymidine incorporation assay, flow cytometric analysis, tryphan blue exclusion assay, western blot analysis, quantative RT-PCR were taken. Results : Saengkankunbi-tang had no effect on proliferation, cell-cycle and DNA replications of HepG2 cells, while it improved cell viability and reduced apoptosis, and it activated Akt and NFKB. But, it did not produce an effect on cell viability and apoptosis when P13K/Akt pathway was blocked by LY294002 nor when $NF{\kapa}B$ activation was blocked by DN-$I{\kapa}B$. Conclusion : These results suggests that Saengkankunbi-tang improves cell viability and reduces apoptosis of HepG2 cells, by activating $NF{\kapa}B$ through PI3K/Akt pathway.

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Effects of IL-3 and SCF on Histamine Production Kinetics and Cell Phenotype in Rat Bone Marrow-derived Mast Cells

  • Lee, Haneul Nari;Kim, Chul Hwan;Song, Gwan Gyu;Cho, Sung-Weon
    • IMMUNE NETWORK
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    • v.10 no.1
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    • pp.15-25
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    • 2010
  • Background: Rat mast cells were regarded as a good model for mast cell function in immune response. Methods: Rat bone marrow mast cells (BMMC) were prepared both by recombinant rat IL-3 (rrIL-3) and by recombinant mouse stem cell factor (rmSCF), and investigated for both proliferation and differentiation in time course. Rat BMMC was induced by culture of rat bone marrow cells (BMCs) in the presence of both rrIL-3 (5 ng/ml) and rmSCF (5 ng/ml). Culture media were changed 2 times per week with the cell number condition of $5{\times}10^4/ml$ in 6 well plate. Proliferation was analyzed by cell number and cell counting kit-8 (CCK-8) and differentiation was by rat mast cell protease (RMCP) II and histamine. Results: Cell proliferation rates reached a maximum at 8 or 11 days of culture and decreased thereafter. However, both RMCP II production and histamine synthesis peaked after 11 days of culture. By real time RT-PCR, the level of histidine decarboxylase mRNA was more than 500 times higher on culture day 11 than on culture day 5. By transmission electron microscopy, the cells were heterogeneous in size and contained cytoplasmic granules. Using gated flow cytometry, we showed that cultured BMCs expressed high levels of $Fc{\varepsilon}RI$ and the mast cell antigen, ganglioside, on culture day 11. Conclusion: These results indicate that rat BMMCs were generated by culturing BMCs in the presence of rrII-3 and rmSCF and that the BMMCs have the characteristics of mucosal mast cells.

Knockdown of UHRF1 by Lentivirus-mediated shRNA Inhibits Ovarian Cancer Cell Growth

  • Yan, Feng;Shao, Li-Jia;Hu, Xiao-Ya
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.4
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    • pp.1343-1348
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    • 2015
  • Human UHRF1 (ubiquitin-like PHD and RING finger domain-containing 1) has been reported to be over-expressed in many cancers, but its role in ovarian cancer remains elusive. Here, we determined whether knockdown of UHRF1 by lentivirus-mediated shRNA could inhibit ovarian cancer cell growth. Lentivirus-mediated short hairpin RNAs (lv-shRNAs-UHRF1) were designed to trigger the gene silencing RNA interference (RNAi) pathway. The efficiency of lentivirus-mediated shRNA infection into HO-8910 and HO-8910 PM cells was determined using fluorescence microscopy to observe lentivirus-mediated GFP expression and was confirmed to be over 80 percent. UHRF1 expression in infected HO-8910 and HO-8910 PM was evaluated by real-time PCR and Western blot analysis. The Cell Counting Kit-8 (CCK-8) assay was used to measure cell viability; flow cytometry and Hoechst 33342 assay was applied to measure cell cycle arrest and apoptosis. Cell invasion was assessed using transwell chambers. Our results demonstrated that the loss of UHRF1 promoted HO-8910 and HO-8910 PM cell apoptosis, while inhibiting cell proliferation. In addition, UHRF1 knockdown significantly inhibited the invasion of human ovarian cancer cells. In the present study, we also showed that depleting HO-8910 cells of UHRF1 caused activation of the DNA damage response pathway, with the cell cycle arrested in G2/M-phase. The DNA damage response in cells depleted of UHRF1 was illustrated by phosphorylation of CHK (checkpoint kinase) 2 on Thr68, phosphorylation of CDC25 (cell division control 25) on Ser 216 and phosphorylation of CDK1 (cyclin-dependent kinase 1) on Tyr 15.

Lactobacillus acidophilus Contributes to a Healthy Environment for Vaginal Epithelial Cells

  • Pi, Woo-Jin;Ryu, Jae-Sook;Roh, Jae-Sook
    • Parasites, Hosts and Diseases
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    • v.49 no.3
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    • pp.295-298
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    • 2011
  • Lactobacillus species in the female genital tract are thought to act as a barrier to infection. Several studies have demonstrated that lactobacilli can adhere to vaginal epithelial cells. However, little is known about how the adherence of lactobacilli to vaginal epithelial cells affects the acidity, cell viability, or proliferation of the lactobacilli themselves or those of vaginal epithelial cells. Lactobacillus acidophilus was co-cultured with immortalized human vaginal epithelial cells (MS74 cell line), and the growth of L. acidophilus and the acidity of the culture medium were measured. MS74 cell density and viability were also assessed by counting cell numbers and observing the cell attachment state. L. acidophilus showed exponential growth for the first 6 hr until 9 hr, and the pH was maintained close to 4.0-5.0 at 24 hr after culture, consistent with previous studies. The growth curve of L. acidophilus or the pH values were relatively unaffected by co-culture with MS74 cells, confirming that L. acidophilus maintains a low pH in the presence of MS74 cells. This co-culture model could therefore potentially be used to mimic vaginal conditions for future in vitro studies. On the other hand, MS74 cells co-cultured with L. acidophilus more firmly attached to the culture plate, and a higher number of cells were present compared to cells cultured in the absence of L. acidophilus. These results indicate that L. acidophilus increases MS74 cell proliferation and viability, suggesting that lactobacilli may contribute to the healthy environment for vaginal epithelial cells.

Development of mcyB-specific Ultra-Rapid Real-time PCR for Quantitative Detection of Microcystis aeruginosa (Microcystis aeruginosa의 정량을 위한 mcyB 특이 초고속 실시간 유전자 증폭법의 개발)

  • Jung, Hyunchul;Yim, Byoungcheol;Lim, Sujin;Kim, Byounghee;Yoon, Byoungsu;Lee, Okmin
    • Journal of Korean Society on Water Environment
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    • v.34 no.1
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    • pp.46-56
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    • 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.

Colonial Cyanobacteria, Microcystis Cell Density Variations using Ultrasonic Treatment (초음파 처리 조건에 따른 집락형 유해남조류 Microcystis 세포수 변화 연구)

  • Lee, Hae-Jin;Park, Hae-Kyung;Heo, Jun;Lee, Hyeon-Je;Hong, Dong-Gyun
    • Journal of Korean Society on Water Environment
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    • v.34 no.2
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    • pp.210-215
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    • 2018
  • It is difficult to count colonial cyanobacteria Microcystis cells since the thickness of colonies is constrained by amorphous mucilage, making it impossible to estimate the number of cells. Disaggregation of Microcystis colonies into single cell is needed to improve the accuracy and precision of cell density estimation of naturally collected samples. Uultrasonic treatment method is commonly used owing to the simplicity and immediacy of the procedure. However, amplitude, frequency, and duration of ultrasonic treatment also cause cell loss during the experiment. Optimal ultrasonic treatment has not been standardized yet. Therefore, the objective of this study was to investigate optimal ultrasonic treatment by analyzing cell density and colony numbers. We collected colonial Microcystis from Changnyeong-Haman weir area in Nakdong River during harmful algal boom period from September to October in 2017. Ultrasonic treatment method was applied to disrupt colonies into single cells to enumerate cell density. Among treatment conditions, results from continuously treated for 100 seconds were found to be the optimum to reduce colonies to a suspension of single cell without cell losses under high and low density of Microcystis cells. Lugol iodine fixed cells followed by sonication showed less negative impact of cell damage within the optimal treatment time (100 seconds). Furthermore, disaggregated cells treated by sonication enables microscopic observation more easily since gas vacuoles were collapsed to facilitate sedimentation of cells under the counting chamber for quantitative enumeration of buoyant Microcystis cells.