Using Measurements of Anchoring Energies of Liquid Crystals on Surfaces To Quantify Proteins Captured by Immobilized Ligands

摘要

We describe a simple optical method that employs measurement of the interaction energy of a liquid crystal(LC)with a surface(the so-called anchoring energy)to report proteins captured on surfaces through specific interactions with immobilized binding groups.To define the sensitivity and dynamic range of the response of the LC,we covalently immobilized a tyrosine-containing,13-residue peptide sequence(Y1173)from the epidermal growth factor receptor/kinase(EGFR)at which autophosphorylation takes place and to which the adapter protein She binds.We determined that,on peptide-decorated(Y1173 or pY1173,where pY1173 is the corresponding phosphopeptide)surfaces incubated against anti-phosphotyrosine antibody,the anchoring energy of the LC decreased systematically from 4.4 to 1.4 mu J/m2(with SEM=0.3 mu J/m2 for n=5)as the antibody concentration increased from 10 pM to 100 nM.Over the same range of antibody concentrations in solution and densities of immobilized peptides,independent ellipsometric measurements were not sufficiently sensitive to report the captured antibody(ellipsometric thicknesses were<0.1 nm).These results,when combined with control experiments reported in this article,provide the first demonstration of the use of anchoring energy measurements of LCs to report proteins captured by immobilized ligands on surfaces.The sensitivity and dynamic range of the methodology suggest that it may offer the basis of a simple yet broadly useful principle for reporting the interactions between proteins and other biomolecules that underlie complex and poorly understood chemical and biological events.