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A Novel Polyclonal Antiserum against Toxoplasma gondii Sodium Hydrogen Exchanger 1

  • Xiao, Bin;Kuang, Zhenzhan;Zhan, Yanli;Chen, Daxiang;Gao, Yang;Li, Ming;Luo, Shuhong;Hao, Wenbo
    • Parasites, Hosts and Diseases
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    • 제54권1호
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    • pp.21-29
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    • 2016
  • The sodium hydrogen exchanger 1 (NHE1), which functions in maintaining the ratio of $Na^+$ and $H^+$ ions, is widely distributed in cell plasma membranes. It plays a prominent role in pH balancing, cell proliferation, differentiation, adhesion, and migration. However, its exact subcellular location and biological functions in Toxoplasma gondii are largely unclear. In this study, we cloned the C-terminal sequence of T. gondii NHE1 (TgNHE1) incorporating the C-terminal peptide of NHE1 (C-NHE1) into the pGEX4T-1 expression plasmid. The peptide sequence was predicted to have good antigenicity based on the information obtained from an immune epitope database. After induction of heterologous gene expression with isopropyl-b-D-thiogalactoside, the recombinant C-NHE1 protein successfully expressed in a soluble form was purified by glutathione sepharose beads as an immunogen for production of a rabbit polyclonal antiserum. The specificity of this antiserum was confirmed by western blotting and immunofluorescence. The antiserum could reduce T. gondii invasion into host cells, indicated by the decreased TgNHE1 expression in T. gondii parasites that were pre-incubated with antiserum in the process of cell entry. Furthermore, the antiserum reduced the virulence of T. gondii parasites to host cells in vitro, possibly by blocking the release of $Ca^{2+}$. In this regard, this antiserum has potential to be a valuable tool for further studies of TgNHE1.

Numerical analysis of melt migration and solidification behavior in LBR severe accident with MPS method

  • Wang, Jinshun;Cai, Qinghang;Chen, Ronghua;Xiao, Xinkun;Li, Yonglin;Tian, Wenxi;Qiu, Suizheng;Su, G.H.
    • Nuclear Engineering and Technology
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    • 제54권1호
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    • pp.162-176
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    • 2022
  • In Lead-based reactor (LBR) severe accident, the meltdown and migration inside the reactor core will lead to fuel fragment concentration, which may further cause re-criticality and even core disintegration. Accurately predicting the migration and solidification behavior of melt in LBR severe accidents is of prime importance for safety analysis of LBR. In this study, the Moving Particle Semi-implicit (MPS) method is validated and used to simulate the migration and solidification behavior. Two main surface tension models are validated and compared. Meanwhile, the MPS method is validated by the L-plate solidification test. Based on the improved MPS method, the migration and solidification behavior of melt in LBR severe accident was studied furthermore. In the Pb-Bi coolant, the melt flows upward due to density difference. The migration and solidification behavior are greatly affected by the surface tension and viscous resistance varying with enthalpy. The whole movement process can be divided into three stages depending on the change in velocity. The heat transfer of core melt is determined jointly by two heat transfer modes: flow heat transfer and solid conductivity. Generally, the research results indicate that the MPS method has unique advantage in studying the migration and solidification behavior in LBR severe accident.

Experimental Applications of in situ Liver Perfusion Machinery for the Study of Liver Disease

  • Choi, Won-Mook;Eun, Hyuk Soo;Lee, Young-Sun;Kim, Sun Jun;Kim, Myung-Ho;Lee, Jun-Hee;Shim, Young-Ri;Kim, Hee-Hoon;Kim, Ye Eun;Yi, Hyon-Seung;Jeong, Won-Il
    • Molecules and Cells
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    • 제42권1호
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    • pp.45-55
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    • 2019
  • The liver is involved in a wide range of activities in vertebrates and some other animals, including metabolism, protein synthesis, detoxification, and the immune system. Until now, various methods have been devised to study liver diseases; however, each method has its own limitations. In situ liver perfusion machinery, originally developed in rats, has been successfully adapted to mice, enabling the study of liver diseases. Here we describe the protocol, which is a simple but widely applicable method for investigating the liver diseases. The liver is perfused in situ by cannulation of the portal vein and suprahepatic inferior vena cava (IVC), with antegrade closed circuit circulation completed by clamping the infrahepatic IVC. In situ liver perfusion can be utilized to evaluate immune cell migration and function, hemodynamics and related cellular reactions in each type of hepatic cells, and the metabolism of toxic or other compounds by changing the composition of the circulating media. In situ liver perfusion method maintains liver function and cell viability for up to 2 h. This study also describes an optional protocol using density-gradient centrifugation for the separation of different types of hepatic cells, allowing the determination of changes in each cell type. In summary, this method of in situ liver perfusion will be useful for studying liver diseases as a complement to other established methods.

Macrophage Migration Inhibitory Factor (MIF) Interacts with Bim and Inhibits Bim-mediated Apoptosis

  • Liu, Lingfeng;Chen, Jinzhong;Ji, Chaoneng;Zhang, Jiayi;Sun, Junlei;Li, Yao;Xie, Yi;Gu, Shaohua;Mao, Yumin
    • Molecules and Cells
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    • 제26권2호
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    • pp.193-199
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    • 2008
  • The pro-apoptotic Bcl-2 family member Bim acts as a sensor for apoptotic stimuli and initiates apoptosis through the mitochondrial pathway. To identify novel regulators of Bim, we employed the yeast two-hybrid system and isolated the human gene encoding macrophage migration inhibitory factor (MIF), a ubiquitously expressed proinflammatory mediator that has also been implicated in cell proliferation, the cell cycle and carcinogenesis. The interaction between MIF and Bim was confirmed by both in vitro and in vivo protein interaction assays. Intriguingly, protein complexes between MIF and the three major Bim isoforms (BimEL/BimL/BimS) could be detected in HEK293 and K562 cells, especially in cells undergoing apoptosis. Moreover, exogenous expression of MIF partially inhibited Bim-induced apoptosis in HEK293 cells. SiRNA-mediated knockdown of MIF increased apoptosis in K562 cells exposed to the chemical oxidant diamide. Endogenous MIF may regulate the pro-apoptotic activity of Bim and inhibit the release of cytochrome c from mitochondria.

Mycobacterium tuberculosis의 효과적인 RNA 추출방법 (An Effective Method of RNA Extraction from Mycobacterium tuberculosis)

  • 오태상;강희윤;남유선;김영진;유은경;이민영;조선영;이희주
    • Annals of Clinical Microbiology
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    • 제19권1호
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    • pp.20-23
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    • 2016
  • RNA 기반 연구에서 고품질의 RNA 추출은 상당히 중요하다. Mycobacterium tuberculosis는 세포벽에 mycolic acid가 풍부하여 두껍고 왁스와 같은 성질을 띈다. 이로 인해 용균이 쉽지 않아 RNA 추출이 어렵다. TRIzol 시약과 silica bead를 이용한 균 파쇄를 통하여 높은 수율의 RNA가 추출이 가능하였다.