Open AccessUltrasensitive Confocal Fluorescence Microscopy of C-Reactive Protein Interacting With FcγRIIa
Author(s) -
D Manolov,
Carlheinz Röcker,
Vinzenz Hombach,
G. Ulrich Nienhaus,
Jan Torzewski
Publication year - 2004
Publication title -
arteriosclerosis, thrombosis, and vascular biology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.007
H-Index - 270
eISSN - 1524-4636
pISSN - 1079-5642
DOI - 10.1161/01.atv.0000147407.17137.02
Subject(s) - dissociation (chemistry) , chemistry , antibody , receptor , fluorophore , confocal microscopy , fluorescence , c reactive protein , immunolabeling , biophysics , biology , biochemistry , immunology , microbiology and biotechnology , immunohistochemistry , inflammation , physics , quantum mechanics
Background— C-Reactive protein (CRP) is an acute phase protein with a suggested pathogenic role in cardiovascular disease. Previous reports proposed that the low-affinity IgG receptor FcγRIIa is the major receptor for CRP. However, these reports were met with criticism because the use of anti-CRP antibodies in the detection of CRP binding to FcγRIIa may have caused false-positive results.Methods and Results— To resolve this controversy, we used ultrasensitive fluorescence microscopy to study the association, dissociation, and equilibrium of CRP binding to FcγRIIa. CRP indeed binds to FcγRIIa, with low association rates and dissociation rates. Anti-CRP antibodies markedly enhance binding, as is evident from the decrease of the equilibrium dissociation coefficient by 2 orders of magnitude.Conclusions— Our study demonstrates the virtues of single fluorophore labeling and highlights the pitfalls of immunolabeling in investigating CRP/Fc receptor interactions. Importantly, this article provides the first quantitative characterization of CRP binding to FcγRIIa and explains and reconciles the diverse and conflicting data presented in the literature.