Journal
ANALYTICAL CHEMISTRY
Volume 84, Issue 8, Pages 3523-3530Publisher
AMER CHEMICAL SOC
DOI: 10.1021/ac202923j
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Funding
- NanoSystems Initiative Munich (NIM)
- Ludwig Maximilians Universitat Munchen (LMU) Initiative Functional Nanosystems
- Center for Nanoscience
- Deutsche Forschungsgemeinschaft
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The direct quantification of both the binding affinity and absolute concentration of disease-related biomarkers in biological fluids is particularly beneficial for differential diagnosis and therapy monitoring. Here, we extend microscale thermophoresis to target immunological questions. Optically generated thermal gradients were used to deplete fluorescently marked antigens in 2- and 10-fold-diluted human serum. We devised and validated an autocompetitive strategy to independently fit the concentration and dissociation constant of autoimmune antibodies against the cardiac beta 1-adrenergic receptor related to dilated cardiomyopathy. As an artificial antigen, the peptide CORI was designed to mimic the second extracellular receptor loop. Thermophoresis resolved antibody concentrations from 2 to 200 nM and measured the dissociation constant as 75 nM. The approach quantifies antibody binding in its native serum environment within microliter volumes and without any surface attachments. The simplicity of the mix and probe protocol minimizes systematic errors, making thermophoresis a promising detection method for personalized medicine.
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