Fluorescence polarization detection for affinity capillary electrophoresis

Electrophoresis
X Chris LeMichael T Lam

Abstract

Affinity capillary electrophoresis (ACE) with laser-induced fluorescence polarization (LIFP) detection is described, with examples of affinity interaction studies. Because fluorescence polarization is sensitive to changes in the rotational motion arising from molecular association or dissociation, ACE-LIFP is capable of providing information on the formation of affinity complexes prior to or during CE separation. Unbound, small fluorescent probes generally have little fluorescence polarization because of rapid rotation of the molecule in solution. When the small fluorescent probe is bound to a larger affinity agent, such as an antibody, the fluorescence polarization (and anisotropy) increases due to slower motion of the much larger complex molecule in the solution. Fluorescence polarization results are obtained by simultaneously measuring fluorescence intensities of vertical and horizontal polarization planes. Applications of CE-LIFP to both strong and weak binding systems are discussed with antibody-antigen and DNA-protein binding as examples. For strong affinity binding, such as between cyclosporine and its antibody, complexes are formed prior to CE-LIFP analysis. For weaker binding, such as between single-stranded DNA and it...Continue Reading

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Citations

Nov 25, 2003·Journal of Chromatography. B, Analytical Technologies in the Biomedical and Life Sciences·C BertucciV Andrisano
Feb 5, 2003·Current Opinion in Biotechnology·Chiara Galbusera, David D Y Chen
Jul 24, 2012·Analytica Chimica Acta·Ryan A PicouS Douglass Gilman
Jun 12, 2009·Electrophoresis·Niels H H Heegaard
May 24, 2006·Electrophoresis·Rukhsana JabeenJohn Petersen
May 30, 2006·Electrophoresis·Jesper Ostergaard, Niels H H Heegaard
Aug 29, 2007·Analytica Chimica Acta·Jung Hwan KimEun Gyeong Yang
Jan 18, 2011·Analytica Chimica Acta·Meng Jing, Michael T Bowser
Jul 9, 2014·Analytica Chimica Acta·Bin DengX Chris Le
Jul 28, 2015·Food Chemistry·Anna PennacchioSabato D'Auria
Feb 5, 2003·Current Opinion in Biotechnology·Victor Pavski, X Chris Le

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