Synthesis and Evaluation of anti-Nonspecific Binding Coating in Microfluidic Devices for ELISA Bioassays

Gelwicks, Melissa J.
A common ELISA (Enzyme Linked Immunosorbant Assay) method for detection of biomolecules utilizes antibody coatings on surfaces that bind specific antigens. An enzyme-antibody conjugate that also binds to the antigen is then added, followed by a substrate that is converted by the enzyme to a detectable product. The sensitivity and reliability of these assays may be significantly reduced by the phenomenon of non-specific binding (NSB), in which the enzyme-antibody conjugate binds not only to the target antigen (in a specific fashion), but also binds by hydrophobic and/or hydrogen-bonding interactions to other species on the surface. The signal produced by such non-specifically bound enzyme-antibody conjugates cannot be distinguished from that produced from the antigen-antibody complex, and a misleadingly high level of signal results. My current research involves development of a microfluidic version of ELISA, with a specific focus on methods for decreasing signal from NSB. The semester began with fabrication of microfluidic devices. I then successfully synthesized long hydrophilic molecules to bind with ELISA surfaces in order to reduce NSB by steric exclusion. I have also tested molecules equipped with zwitterion head groups, which are highly resistant to non-specific protein adsorption, with the intended result of reducing NSB.
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