A fluidic fiber platform modified for the selective extraction and on fiber fluorescence detection of proteins and nucleic acids

Solid phase extraction has become a popular tool for analytical method development as a sample preparation technique. Solid phase sorbents can generally extract analytes based on polarity or selectively based on highly specific interactions between the analyte and an immobilized surface molecule. Biomolecular recognition based solid phases utilize antibodies, nucleic acids, and proteins as a secondary affinity layer on a solid phase support for selective analyte extractions and have been used for sample preparation, purification, separations, bioassays, and molecular diagnostics. Polymer supports for solid phase extraction are desirable for their low cost, abundance, chemical inertness, and pH stability. This thesis reports the use of capillary-channel polymer (C-CP) fibers as a solid support modified for selective extraction and on fiber fluorescence detection of nucleic acids and proteins. The polymer fibers are low cost and their unique shape allows them to be easily adapted into a fluidic fiber based micro spin column configuration for a small scale, inexpensive, low-tech, and easy to use device. Platform performance was successful as demonstrated by modifying the polymer surface with neutravidin via adsorption to tether specific biotinylated analyte recognition moieties. Fluorescence resulted from either hybridization of a complementary fluorophore labeled probe to nucleic acids or conjugation of a fluorophore to the analyte protein. On fiber fluorescence detection of nucleic acids and proteins was successful with minimal non-specific binding of proteins and nucleic acids to the polymer surface. Two sequence specific oligonucleotides were added and probed either separately or simultaneously on the fibers; the fluorescence intensity increasing with the amount of oligonucleotide added, leveling off at 100 pmol. Fluorescence signal never saturated for small volume additions (10ul) up to 100 pmol of added oligonucleotide. The LOD was calculated to be 100s of fmol for all experiments. For selective extraction of proteins, protein recovery was similar for both C-CP fibers and streptavidin conjugated magnetic microbeads. Adsorbed neutravidin did not irreversibly bind to the C-CP fiber surface and was partially removed in the presence of Tween-20. C-CP fibers and spin columns are inexpensive, easy to use, and can be integrated as a solid phase extraction sample preparation step for small scale bioassays or molecular diagnostics