Shao-min SHUANG(双少敏)1, Xiao-na QIAO(乔晓娜)1, Dan-dan WANG( 王丹丹)2, Li FAN(樊丽)1,Dong-tao LU(芦冬涛)1, Li-xiao MU(慕丽晓)1, Ye-hong ZHOU(周叶红)1
1. School of Chemistry and Chemical Engineering, Shanxi Universit y, Taiyuan 030006, China;2. China Research Institute of Daily Chemical Industry, Taiyuan 030006, China
Abstract-The interaction of Fura-2 with Ca2+ is studied using steady fluorescence technique. The effect of pH on the spectra behavior o f Fura-2 in the presence of Ca2+ is investigated, the excitation maxima ( 340 nm) and the isobestic point (360 nm) for the fluorescence spectra of Fura-2 depend on pH. At different temperatures the apparent dissociation constants ( Kd) of Fura-2-Ca2+ complex are examined, Kd is found to decrease with increasing temperatures (20 ℃, 37 ℃, 50 ℃) and Δ H is calculated to be 21.16 kJ/mol by using the Van’t Hoff equation at pH 7 .4 for all the temperatures tested. The determination of intracellular Ca2+
concentration ([Ca2+]i) in lymphocyte is developed by using Fura-2 as a fluorescence probe in the presence of Cefotaxime at 37 ℃ and pH 7 .4.
Key words-Fura-2; pH; temperature; [Ca2+ ]i; cefotaxime
Manuscript Number: 1674-8042(2010)04-0395-06
dio: 10.3969/j.issn.1674-8042.2010.04.21
References
[1]S. Feske, 2007. Calcium signaling in lymphocyte activation and disea se. Nat. Rev. Immunol., 7: 690-702.
[2]M. J. Berridge, M. D. Bootman, P. Lipp, 1998. Calcium-a life and de ath signal. Nature, 395: 645-648.
[3]R. Huang, J. Cioffi, R. Kimberly, J., et al, 1995. B cell differenti ation factor-induced human B cell maturation: stimulation of intracellular calc ium release. Cell. Immunol., 164: 227-33.
[4]J. L. Ambrus, L. Chesky, P. McFarland, et al, 1991. Induction of pro liferation by high molecular weight B cell growth factor or low molecular weight B cell growth factor is associated with increases in intracellular calcium in d ifferent subpopulations of human B lymphocytes. Cell. Immunol., 134: 314-324.
[5]O. A. Sukocheva, A. Y. Abramor, J. O. Leritskaya, et al, 2001. Modul ation of intracellular Ca2+ concentration by vitamin B12 in rat thymocytes . Blood Cell, Molecules and Diseases, 27: 812-824.
[6]B. Lynn, Dustin Ph. D, 2000. Ratiometric analysis of calcium mobil ization. Clin. Appl. Immunol. Rev. 1: 5-15.
[7]G. Grynkiewicz, M. Poenie, R. Y. Tsien, 1985. A new generation of Ca 2+ indicator with greatly improved fluorescence properties. J. B iol. Chem., 260: 3440-3450.
[8]C. N. Topala, J. P. H. Schoeber, Lydia E. Searchfield, et al, 2009. Activation of the Ca2+-sensing receptor stimulates the activity of the ep ithelial Ca2+ channel TRPV5. Cell Calcium, 45: 331-339 .
[9]Jun Li, Jun-jie Xiao, Dan-dan Liang, et al, 2010. Inhibition of mi tochondrial translocator protein prevents atrial fibrillation. Europea n Journal of Pharmacology, 632: 60-64.
[10]L. Archibald, L. Phillips, D. Monnet, et al, 1997. Antimicrobial re sistance in isolates from inpatients and outpatients in the United States: incre asing importance of the intensive care unit. Clin. Infect. Dis., 24: 211-15.
[11]R. N. Jones, 1996. Impact of changing pathogens and antimicrobial s usceptibility patterns in the treatment of serious infections in hospitalized pa tients. Am. J. Med., 100: 3S-12S.
[12]P. A. Todd, R. N. Brogden, 1990. Cefotaxime. An update of its pharm acology and therapeutic use. Drugs, 40: 608-651.
[13]C. H. Kuo, K. Maita, S. D. Sleight, et al, 1983. Lipid peroxidation : a possible mechanism of cephaloridine-induced nephrotoxicity. Toxicol. Appl. Pharmacol, 67: 78-88.
[14]C. Cojocel, K. H. Laeschke, G. Inselmann, et al, 1985. Inhibition o f cephaloridine-induced lipid peroxidation. Toxicology, 35: 295-305.
[15]H. U. Weltzien, C. Moulon, S. Martin, et al, 1996. T cell immune re sponses to haptens. Structural models for allergic and autoimmune reactions. Toxicology, 107: 141-151.
[16]F. A. Lattanzio, D. K. Bartschat, 1991. The effect of pH on rate co nstants, ion selectivity and thermodynamic properties of fluorescent calcium and magnesium indicators. Biochem. Biophys. Res. Commun, 177: 18 4-191.
[17]T. J. Shuttleworth, J. L. Thompson, 1991. Effect of temperature on receptor-activated changes in [Ca2+]i and their determination using fluorescent probes. J. Biol. Chem., 266:1 410-1414.
[18]J. Paltauf-Doburzynska, W. F. Graier, 1997. Temperature dependenc e of agonist-stimulated Ca2+ signaling in cultured endothelial cells. Cell Calcium, 21: 43-51.
[19]D. Larsson, B. Larsson, T. Lundgren, et al, 1999. The effect of pH and temperature on the dissociation constant for fura-and their effects on Ca 2+ in enterocytes from a poikilothermic animal. Atlantic Cod (Gadu s morhua). Anal. Biochem, 273: 60-65.
[full text view]
