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  • Enhancing Protein Capture Using a Combination of Nanoyeast Single-Chain Fragment Affinity Reagents and Alternating Current Electrohydrodynamic Forces

    Author(s)
    Vaidyanathan, Ramanathan
    Rauf, Sakandar
    Grewal, Yadveer S
    Spadafora, Lauren J
    Shiddiky, Muhammad JA
    Cangelosi, Gerard A
    Trau, Matt
    Griffith University Author(s)
    Shiddiky, Muhammad J.
    Year published
    2015
    Metadata
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    Abstract
    New high-performance detection technologies and more robust protein capture agents can be combined to both rapidly and specifically capture and detect protein biomarkers associated with disease in complex biological samples. Here we demonstrate the use of recently developed recombinant affinity reagents, namely nanoyeast-scFv, in combination with alternating current electrohydrodynamic (ac-EHD)-induced shear forces, to enhance capture performance during protein biomarker analysis. The use of ac-EHD significantly improves fluid transport across the capture domain, resulting in enhanced sensor-target interaction and simultaneous ...
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    New high-performance detection technologies and more robust protein capture agents can be combined to both rapidly and specifically capture and detect protein biomarkers associated with disease in complex biological samples. Here we demonstrate the use of recently developed recombinant affinity reagents, namely nanoyeast-scFv, in combination with alternating current electrohydrodynamic (ac-EHD)-induced shear forces, to enhance capture performance during protein biomarker analysis. The use of ac-EHD significantly improves fluid transport across the capture domain, resulting in enhanced sensor-target interaction and simultaneous displacement of nonspecific molecules from the electrode surface. We demonstrate this simple proof-of-concept approach for the capture and detection of Entamoeba histolytica antigens from disinfected stool, within a span of 5 min using an ac-EHD microfluidic device. Under an ac-EHD field, antigens were captured on a nanoyeast-scFv immobilized device and subsequently detected using a quantum dot conjugated antibody. This immunosensor specifically detected antigen in disinfected stool with low background noise at concentrations down to 58.8 fM with an interassay reproducibility (%RSD of n = 3) < 17.2%, and in buffer down to 5.88 fM with an interassay reproducibility (% RSD, n = 3) of 8.4%. Furthermore, antigen detection using this immunosensor was 10 times more sensitive than previously obtained with the same nanoyeast-scFv reagents in a microfluidic device employing surface-enhanced Raman scattering (SERS) detection in buffer and at least 200 times more sensitive than methods using screen printed gold electrodes in disinfected stool. We predict this rapid and sensitive approach using these stable affinity reagents may offer a new methodology to detect protein disease biomarkers from biological matrices.
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    Journal Title
    Analytical Chemistry
    Volume
    87
    Issue
    23
    DOI
    https://doi.org/10.1021/acs.analchem.5b02490
    Subject
    Analytical chemistry
    Analytical chemistry not elsewhere classified
    Other chemical sciences
    Publication URI
    http://hdl.handle.net/10072/172422
    Collection
    • Journal articles

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