Reliable and rapid separation of proteins is both a fundamental and challenging problem for bioanalytical and biomedical research. Conventional separation techniques reach their limits at extremes, for example, when increased sample complexity demands for the analysis of relevant disease markers in extremely small concentration and within a huge background. Problems further arise for time critical samples, i.e. when rapid answers are required, such as for samples with temporally degrading or altering composition or when rapid diagnosis is essential, such as during surgery. These limitations can be more drastic for smaller sample volumes and concomitantly low amount of proteins, such as in the case of minimal invasive diagnosis or single cell analysis. The latter is specifically important for understanding specific cellular pathways and malignant progressions, which would otherwise be averaged in the ensemble measurement. Another extremely relevant example represents the diagnosis of Alzheimer's Disease, which is particularly challenging due to the transient nature of the involved peptide species and their extremely low abundance in body fluids. This exploratory proposal aims to develop dielectrophoretic devices for the efficient, rapid and gel-free separation, purification and pre-concentration of proteins on microfluidic platforms. This project applies a new principle for the gel-free separation of proteins in microfluidic systems. It exploits dielectrophoresis (DEP) of proteins, which as polarizable objects respond to a non-uniform electric field with a migrational motion. As the polarizability depends on various parameters such as shape, charge, charge density, permittivity or deformability it thus allows probing the DEP response of proteins in a broad range. The dielectrophoretic response of proteins is provoked in tailored microstructure designs on a lab-on- a-chip platform in which the necessary inhomogeneous electric field gradient can be optimally generated. A fundamental investigation of the DEP behavior of selected proteins will give insight into the necessary electrical driving parameters and reveal optimized conditions for more complex separation problems as well as the purification and pre-concentration of proteins and peptides. This novel device is capable of analyzing protein samples in time scales of a few minutes and reduces sample volumes to the pL-nL range. In particular, we develop a combined microfluidic immunoaffinity and DEP based separation assay for beta-amyloid (A2) oligomers in cerebrospinal fluid. This novel DEP separation method thus represents a development of outstanding importance for biomedical research and point of care diagnostics.

Public Health Relevance

(provided by the applicant): The goal of this project is to design and test devices for fast, reliable and gel-free protein separation and pre-concentration based on dielectrophoresis on a microfluidic platform. The development of these devices extends current separation techniques at their limits, i.e. for low sample concentration, time critical analyzes, miniature sample amounts and complex samples. It will facilitate biomedical research and eventually clinical laboratory practice, in particular, rapid and on-site point of care diagnostics.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Exploratory/Developmental Grants (R21)
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Special Emphasis Panel (ZRR1-BT-7 (01))
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Friedman, Fred K
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Arizona State University-Tempe Campus
Schools of Arts and Sciences
United States
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Nakano, Asuka; Camacho-Alanis, Fernanda; Ros, Alexandra (2015) Insulator-based dielectrophoresis with ?-galactosidase in nanostructured devices. Analyst 140:860-8
Nakano, Asuka; Luo, Jinghui; Ros, Alexandra (2014) Temporal and spatial temperature measurement in insulator-based dielectrophoretic devices. Anal Chem 86:6516-24
Nakano, Asuka; Ros, Alexandra (2013) Protein dielectrophoresis: advances, challenges, and applications. Electrophoresis 34:1085-96