• Journal of Sensor Science and Technology
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• v.19 no.6
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• pp.421-427
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• 2010

Alizarin Red S modified screen printed carbon electrodes were developed for the electrochemical detection of aluminum ion. The electrodes developed use screen-printed carbon electrodes(SPCEs) coupled with chemical modification with an organic chelator, Alizarin Red S(ARS), for aluminum ion detection in aqueous solution. For sensor fabrication ARS was directly immobilized on the surface of SPCEs using PVA-SbQ(The poly(vinyl alcohol) bearing stryrylpyridinium groups). Aluminum concentrations were indirectly estimated by amperometric determination of the non-complexed ARS immobilized on the electrodes, after its complexation with aluminum. The sensitivity of the sensor developed was $3.8\;nA{\mu}M^{-1}cm^{-2}$ and the detection limit for aluminum was $25\;{\mu}M$.

• Proceedings of the PSK Conference
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• 2003.10b
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• pp.117.2-117.2
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• 2003

The mechanism by which lead enters glial cells was examined. The uptake of lead reached saturation when assays were performed in buffers at pH 5.5 and 7.4. The Vmax and Km was 2.7 pmoles/mg protein/min and 13.4 M in the buffer at pH 7.4, respectively, whereas the Vmax and Km was 329 fmoles/mg and 8.2 M in the buffer at pH 5.5, respectively. Uptake in a buffer at pH 5.5 but not at pH 7.4 was inhibited by iron. Cells treated with the iron chelator desferoxamine displayed higher levels of the divalent metal transporter mRNA and protein. (omitted)

• The Korean Journal of Nuclear Medicine
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• v.37 no.6
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• pp.402-417
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• 2003

Purpose: To evaluate the use of monoclonal antibody (MoAb) as a carrier of the receptor-binding ligand the receptor mediated uptake into liver and subsequent metabolism of $^{111}In-labeled$ galactosylated MoAb-chelator conjugates were investigated and compared with those of $^{111}In$ labeled MoAb. Materials and Methods : T101 MoAb, $IgG_2$ against human lymphocytic leukemic cell, conjugated with cyclic DTPA dianhydride (DTPA) or 2-p-isothiocyanatobenzyl-6-methyl-DTPA (1B4M) was galactosylated with 2-imino-2-methoxyethyl-1-thio-${\beta}$-D-galactose and then radiolabeled with $^{111}In$. Biodistribution and metabolism study was peformed with two $^{111}In-conjugates$ in mice and rats. Results: $^{111}In-labeled$ T101 and its galactosylated conjugates were taken to the liver by the time, mostly within 10 min. However DTPA conjugate was retained longer in the liver than the 1B4M conjugate (55% vs 20% of injected dose at 44 hr). During this time, the radiornetabolite of DTPA conjugate was excreted similarly into urine (24%) and feces (17%). The radiometabolite of 1B4M was excreted primarily into feces (68%) rather than urine (8%). Size exclusion HPLC analysis of the bile and supernatant of liver homogenate showed two peaks the first (35%) with the retention time (Rt) identical to IgG and the second (65%) with Rt similar to free $^{111}In$ at 3 hr post-injection for the 1B4M conjugate, indicating that the metabolite is rapidly excreted through the biliary system. in contrast to DTPA conjugate, the small $^{111}In-DTPA-like$ metabolite was the major radioindium component (90%) in the liver homogenate as early as 3 hour post-injection, but the cumulative radioindium activity in feces was only 17% at 44 hour, indicating that the metabolite from DTPA conjugate does not clear readily through the biliary tract. Conclusion: The galactosylation of the MoAb conjugates resulted in higher hepatocyte uptake and enhanced metabolism, compared to those without galactosylation. Metabolism of the MoAb-conjugates is different between compounds radiolabled with different chelators due to different characteristics of radiometabolites generated in the liver.

• Nuclear Medicine and Molecular Imaging
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• v.43 no.5
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• pp.487-494
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• 2009

Purpose: Small size of recombinant scFv antibody has many advantages such as rapid blood clearances and improved targeting antibodies to tumor region. On the other hand owing to small size, number of amino group is insufficient in conjugation with chelator and fluorescence labeling. This study is to introduce poly lysine tag to the C-terminal end of scFv lym-1 sequence for fluorescence chelator conjugation. Materials and Methods: Poly lysine scFv lym-1 gene, cloned into pET-22b (+) vector, was expressed in E. coli BL21 (DE3) strain. Antibody purification was performed with Ni-NTA column and then size exclusion column chromatography. Expression and purification levels of poly lysine tagged scFv lym-1 antibody were confirmed by western blot analysis. I-124, I-125, I-131 and Tc-99m were used for radiolabeling of purified poly lysine scFv lym-1. Flow cytometry analysis of FIT( conjugated poly lysine scFv lym-1 was performed for confirmation of immunoreactivity of human Burkitt's lymphoma cells. Results: Poly lysine scFv lym-1 antibody was purified through two steps and identified as molecular weight of 48 KDa. Radiolabeling yields of I-124, I-125, I-131 and Tc-99m into poly lysine scFv lym-1 were >99%, >99%, >95% and >99%, respectively. Flow cytometry analysis of poly lysine scFv and scFv lym-1 was showed similar immunoreactivity to human Burkitt's lymphoma cells. Conclusion: Poly lysine tag was useful for the sufficient number of amino groups to scFv lym-1 antibody for chelator conjugation with minimizing loss of immunoreactivity.

• Journal of Korean Medicine Rehabilitation
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• v.20 no.1
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• pp.27-36
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• 2010

Objectives : We investigated the role of intracellular calcium chelator, bis-(2-aminophenoxy)-ethane-N,N,N',N'-tetraacetic acid(BAPTA), on the modulation of phosphorylation of the spinal N-methyl-D-aspartate receptor(NMDAR) NR1 and NR2B subunits following electroacupuncture(EA). Methods : Bilateral 2 Hz EA stimulation with 1.0 mA was delivered at those acupoints corresponding to Zusanli(ST36) and Sanyinjiao(SP6) in man via needles for 30min. Results : EA analgesia was reduced by intra-peritoneal injection at a higher dose of BAPTA from termination of EA stimulation. At 60 min after EA treatment, the total number of c-fos-immunostained neurons in each regions of the dorsal horn in the $L_{4-5}$ segments was decreased by BAPTA injection, especially in nucleus proprius. The mean integrated optical density (IOD) of NR1 and NR2B subunits were increased only in superficial laminae of EA-treated rats when compared with normal rats. However, the mean IOD of pNR1 was significantly decreased by BAPTA injection in both the superficial laminae and neck region and pNR2B in the superficial laminae. Western blot analyses confirmed the decreased expression of pNR1 and pNR2B. Conclusions : We concluded that intracellular calcium may well play an important role in EA analgesia by modulating the phosphorylation state of spinal NMDAR subunits.

• Applied Microscopy
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• v.30 no.2
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• pp.141-152
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• 2000

This study aims to demonstrate the effect of squalene (SQ), one of the natural chelator, on the ultrastructural changes in the mouse liver caused by $CdCl_2$. A total of 40 healthy ICR that weighted 30 gm $({\pm}2gm)$ was used for experiment. The experimental group was divided into two groups; group A and B. The group A administrated $CdCl_2$ (4.0 mg/kg) to the intraperitoneal. The group B administrated $CdCl_2$ (4.0 mg/kg) to the intraperitoneal treated with SQ (180 mg/kg, 2 time/day). Each group was observed at 24, 48, 72, 96 hours after injected $CdCl_2$. The results were as follows: 1. Group A Nuclear membrane was observed very irregular at the 24 hours and keep rounded-shape from 48 hours. Mitochondria were observed destruction of inner cavity to the 72 hours and some showed inner cavity destruction at 96 hours. RER (rough endoplasmic reticulum) showed the dilation of inner cavity all the around and reformed typical lamellae from 72 hours. Lysosome were observed in the cytoplasm from 24 hours. From 72 hours, glycogen showed over cytoplasm. 2. Group B Nuclear membrane was observed regular at overall the time. Mitochondria showed normal shapes (round, rod) at overall the time. To 48 hours, inner cavity of rER dilated and destructed lamellae. But from 72 hours, observed typical lamellae of rER Lysosome were observed from 24 hours. And glycogen showed from 24 hours. These results suggest that squalene attenuates the toxic effect of the $CdCl_2$ in the mouse liver.

• Biomolecules & Therapeutics
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• v.13 no.2
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• pp.78-83
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• 2005

2,3,7,8-Tetrachlorodibenzo-p-dioxin(TCDD) has previously shown to induce neurotoxicity through intracellular $Ca^{2+}$ increase in rat neurons. In this study we investigated the role and signaling pathway of intracellular $Ca^{2+}$ in TCDD-induced inhibition of neuronal cell proliferation in SK-N-SH human neuronal cells. We found that TCDD(10nM) rapidly increased the level of intracellular $Ca^{2+}$, which was completely blocked by the extracellular $Ca^{2+}$ chelation with EGTA (1 mM) or by pretreatment of the cells with the non-selective cation channel blocker. flufenamic acid (200 ${\mu}M$). However, pretreatment of the cells with dantrolene (25 ${\mu}M$) and TMB-8(10 ${\mu}M$), intracellular $Ca^{2+}$-release blockers, or a voltage-sensitive $Ca^{2+}$ channel blocker, varapamil (100 ${\mu}M$), failed to block the TCDD-induced $Ca^{2+}$ increase in the cells. In addition, TCDD induced a rapid and transient activation of phatidvlinositol 3-kinase (PI3K) and extracellular signal-regulated kinase 1/2(ERK1/2), which was ingnificantly blocked by the pretreatment with BAPTA, an intracellular $Ca^{2+}$ chelator, and LY294002, a PI3K inhibitor. Furthermore, inhibitors of PI3K, ERK, or an intracellular $Ca^{2+}$ chelator further potentiated the anti-proliferative effect of TCDD in the cells. Collectively, the results suggest that intracellular $Ca^{2+}$ and PI3K-dependent activation of ERK 1/2 may be involved in the TCDD-induced inhibition of cell proliferation in SK-N-SH human neuronal cells.

• Biomolecules & Therapeutics
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• v.26 no.6
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• pp.546-552
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• 2018

A comprehensive collection of proteins senses local changes in intracellular $Ca^{2+}$ concentrations ($[Ca^{2+}]_i$) and transduces these signals into responses to agonists. In the present study, we examined the effect of sphingosine-1-phosphate (S1P) on modulation of intracellular $Ca^{2+}$ concentrations in cat esophageal smooth muscle cells. To measure $[Ca^{2+}]_i$ levels in cat esophageal smooth muscle cells, we used a fluorescence microscopy with the Fura-2 loading method. S1P produced a concentration-dependent increase in $[Ca^{2+}]_i$ in the cells. Pretreatment with EGTA, an extracellular $Ca^{2+}$ chelator, decreased the S1P-induced increase in $[Ca^{2+}]_i$, and an L-type $Ca^{2+}$-channel blocker, nimodipine, decreased the effect of S1P. This indicates that $Ca^{2+}$ influx may be required for muscle contraction by S1P. When stimulated with thapsigargin, an intracellular calcium chelator, or 2-Aminoethoxydiphenyl borate (2-APB), an $InsP_3$ receptor blocker, the S1P-evoked increase in $[Ca^{2+}]_i$ was significantly decreased. Treatment with pertussis toxin (PTX), an inhibitor of $G_i$-protein, suppressed the increase in $[Ca^{2+}]_i$ evoked by S1P. These results suggest that the S1P-induced increase in $[Ca^{2+}]_i$ in cat esophageal smooth muscle cells occurs upon the activation of phospholipase C and subsequent release of $Ca^{2+}$ from the $InsP_3$-sensitive $Ca^{2+}$ pool in the sarcoplasmic reticulum. These results suggest that S1P utilized extracellular $Ca^{2+}$ via the L type $Ca^{2+}$ channel, which was dependent on activation of the $S1P_4$ receptor coupled to PTX-sensitive $G_i$ protein, via phospholipase C-mediated $Ca^{2+}$ release from the $InsP_3$-sensitive $Ca^{2+}$ pool in cat esophageal smooth muscle cells.

• The Korean Journal of Pharmacology
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• v.32 no.3
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• pp.347-355
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• 1996

We examined the effects of EGTA and verapamil on phorbol ester-and forskolin-stimulated LH releases and $LH{\beta}$ subunit mRNA levels in order to verify the role of extracellular $Ca^{++}$ on PKC- or cAMP-induced increases in LH release and $LH{\beta}$ subunit mRNA levels in cultured anterior pituitary cells of rat. Forskolin-stimulated $LH{\beta}$ subunit mRNA levels as well as LH release were all suppressed by prevention of $Ca^{++}$ mobilization from extracellular environment, after the treatment of EGTA as a $Ca^{++}$ chelator or verapamil as a $Ca^{++}$ channel blocker. PMA-stimulated $LH{\beta}$ subunit mRNA levels were also suppressed by the treatment of EGTA and verapamil, while PMA-induced LH release was not affected. From the present study, it is, therefore, suggested that PKC activation and cAMP elevation all stimulate $LH{\beta}$ subunit mRNA levels and these are extracellular $Ca^{++}$-dependent. However, LH releases by PKC activation and cAMP increase seem to be different each other. LH release by PKC activation is thought to be independent of extracellular $Ca^{++}$. On the other hand, cAMP stimulated-LH release is thought to be dependent on the entry of extracellular $Ca^{++}$.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.21 no.2
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• pp.71-79
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• 1988

For the purpose of producing low salt fermented anchovy by accelerated method with a strong proteolytic bacteria, in this study, a strong proteolytic bacterium was isolated from low salt fermented anchovy and its bacteriological characteristics and properties of protease were experimented. The results obtained were as fellows : three proteolytic bacteria, Aeromonas anaerogenes Barillus subtilis and Staphylococcus saprophyticus were isolated from low salt fermented anchovy($4\%\;of\;salt,\;4\%\;of\;KCl,\;0.5\%\;of\;lactic\;acid,\;6\%$of sorbitol and $4\%$ of alcohol extract of red pepper) after 40 days fermentation. Among these strains, which grow best at $30^{\circ}C$, pH 7.0, B. subtilis was found the best proteolytic strain and benefit for industrial use as shown $0.95\;hr^{-1}$ of specific growth rate, $89{\mu}g-Tyr/hr.ml$ of maximum activity after 12 hrs culture in TPY broth. The protease produced by by B. subtilis showed maximum activity at $35^{\circ}C$, pH 7.0, and molecular weight was estimated to be 23,000 by Sephadex G-100 filtration, and it was supposed to be a kind of metal chelator sensitive neutral protease from the results of strong sensitivity against EDTA, o-phenanthroline and metal ions such as $Cu^{2+},\;Ni^{2+},\;Fe^{2+}.Km$ value of that by method of Lineweaver-Burk was determinded to be $0.73\%$ for casein as a substrate.