Suitable Labels for Molecular Imaging – Influence of Dye Structure and Hydrophilicity on the Spectroscopic Properties of IgG Conjugates
- 21 June 2011
- journal article
- Published by American Chemical Society (ACS) in Bioconjugate Chemistry
- Vol. 22 (7), 1298-1308
- https://doi.org/10.1021/bc1004763
Abstract
Aiming at the design of highly brilliant NIR emissive optical probes, e.g., for in vivo near-infrared fluorescence imaging (NIRF), we studied the absorption and fluorescence properties of the asymmetric cyanines Dy678, Dy681, Dy682, and Dy676 conjugated to the model antibody IgG. The ultimate goal was here to derive general structure-property relationships for suitable NIR fluorescent labels. These Dy dyes that spectrally match Cy5 and Cy5.5, respectively, were chosen to differ in chromophore structure, i.e., in the substitution pattern of the benzopyrylium end group and in the number of sulfonic acid groups. Spectroscopic studies of the free and IgG-bound fluorophores revealed a dependence of the obtained dye-to-protein ratios on dye hydrophilicity and control of the fluorescence quantum yields (Φ(f)) of the IgG conjugates by the interplay of different fluorescence reduction pathways like dye aggregation and fluorescence resonance energy transfer (FRET). Based upon aggregation studies with these dyes, the amount of dye dimers in the IgG conjugates was determined pointing to dye hydrophilicity as major parameter controlling aggregation. To gain further insight into the exact mechanism of dye dimerization at the protein, labeling experiments at different reaction conditions but constant dye-to-protein ratios in the reaction solution were performed. With Dy682 that displays a Φ(f) of 0.20 in PBS and 0.10 for moderate dye-to-protein ratio of 2.5, a low aggregation tendency, and a superior reactivity in IgG labeling, we identified a promising diagnostic tool for the design of NIR fluorescent probes and protein conjugates.This publication has 60 references indexed in Scilit:
- Improved Speciation Characteristics of PEGylated Indocyanine Green-Labeled Panitumumab: Revisiting the Solution and Spectroscopic Properties of a Near-Infrared Emitting anti-HER1 Antibody for Optical Imaging of CancerBioconjugate Chemistry, 2010
- Multimodality Imaging Probes: Design and ChallengesChemical Reviews, 2010
- NIR dyes for bioimaging applicationsCurrent Opinion in Chemical Biology, 2010
- New Strategies for Fluorescent Probe Design in Medical Diagnostic ImagingChemical Reviews, 2009
- H-Type Dimer Formation of Fluorophores: A Mechanism for Activatable, in Vivo Optical Molecular ImagingACS Chemical Biology, 2009
- Near-infrared squaraine dyes for fluorescence enhanced surface assayDyes and Pigments, 2009
- Fluorophore−Quencher Based Activatable Targeted Optical Probes for Detecting in Vivo Cancer MetastasesMolecular Pharmaceutics, 2009
- Determination of Optimal Rhodamine Fluorophore for in Vivo Optical ImagingBioconjugate Chemistry, 2008
- Multimodal Nanoprobes for Radionuclide and Five-Color Near-Infrared Optical Lymphatic ImagingACS Nano, 2007
- A Comparison of the Emission Efficiency of Four Common Green Fluorescence Dyes after Internalization into Cancer CellsBioconjugate Chemistry, 2006