Synthesis and evaluation of Ru-complexes as anisotropy probes for protein hydrodynamics and immunoassays of high-molecular-weight antigens

Biophysical Chemistry
H SzmacinskiJ R Lakowicz

Abstract

We investigated three unsymmetrical Ru-complexes, namely [Ru(bpy)2 (phen-ITC)]2+, [Ru(bpy)2(dcbpy)] and [Ru(bpy)2(mcbpy)]+ for use as probes for rotational diffusion and in immunoassays of high-molecular-weight antigens. For this purpose we synthesized reactive forms of these metal-ligand complexes and conjugated them to human serum albumin (HSA). The maximal anisotropies (r0) for the HSA-bound forms in frozen solution are 0.23, 0.17 and 0.14 for the (dcbpy), (mcbpy) and (phen-ITC) derivatives, respectively. The activated Ru metal-ligand complexes have either one or two NHS-esters or an isothiocyanate group as the reactive moiety. The usefulness of these complexes in immunoassays was determined by titration of the labeled HSA with polyclonal anti-HSA. The highest steady state anisotropy (r) values (0.190) were observed for the [Ru(bpy)2(dcbpy)]-labeled HSA on titration with polyclonal antibody. However, a relative increase in the steady state anisotropy (r/r0) on titration with polyclonal antibody was found for the phen-ITC probe (96%), as compared to the dcbpy (83%) or mcbpy (79%) derivatives. These findings were confirmed by time-resolved frequency-domain measurements. In particular the higher mean correlation times calculate...Continue Reading

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Citations

Jan 6, 2001·Analytical Biochemistry·J R LakowiczJ S Kang
Mar 3, 2010·Journal of Fluorescence·Myung Sup KimJung Sook Kang
Jun 5, 2002·Biochimica Et Biophysica Acta·Jung Sook KangJoseph R Lakowicz
Apr 18, 1998·Biochimica Et Biophysica Acta·H SzmacinskiG J Meyer
Dec 4, 2008·Journal of the American Chemical Society·Igor L MedintzHedi Mattoussi
May 20, 1998·Biophysical Journal·G VerebT M Jovin
Aug 14, 2008·Chirality·Chunxia JiangFrederick M Macdonnell
Jul 27, 2000·Photochemistry and Photobiology·M E Jiménez-HernándezM T Portolés
Jan 14, 2012·Dalton Transactions : an International Journal of Inorganic Chemistry·Kenneth Kam-Wing LoWendell Ho-Tin Law
Jun 1, 1997·Journal of Fluorescence·Henryk MalakFelix N Castellano
Sep 23, 1997·Analytical Biochemistry·E TerpetschnigJ R Lakowicz
Jul 15, 1997·Analytical Biochemistry·L TolosaJ R Lakowicz
Jun 4, 2014·Organometallics·Geoffrey AbbottOgar O L Ichire
Sep 6, 2007·Analytical Chemistry·María Lourdes Sánchez-MartínezAgustina Gómez-Hens
Mar 15, 2009·Journal of Organometallic Chemistry·Ayesha SharminJ B Alexander Ross
Sep 25, 2001·Analytical Chemistry·B P MaliwalJ R Lakowicz
Apr 16, 1999·Analytical Chemistry·I GryczynskiJ R Lakowicz

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