Masters Theses

Date of Award

12-1991

Degree Type

Thesis

Degree Name

Master of Science

Major

Chemistry

Major Professor

Michael J. Sepaniak

Committee Members

Michael Keene, Earl Wehry

Abstract

Certain complex samples require repetitive in situ analysis; however, those samples might be toxic or inaccessible. Fiber optic sensors can analyze remote samples because the flexible fibers are small, and able to transport information over substantial distances. Fiber optic-based biosensors combine the sensitivity of laser-based fluorescence detection with the selectivity of affinityreagents. Our group developed a novel biosensor that delivers fluoroimmunoreagents (solid/liquid phase) to and from a remote sample site through capillary columns, and retains the reacted complex near the fiber with a porous frit. I examined a prototype of this sensor, the Microscale Regenerable Biosensor (MRB), regarding its operational characteristics as they relate to performing competitive-binding fluoroimmunoassay (FIA). I employed a "model" assay of protein A as an affinity reagent for rabbit IgG.

I evaluated the MRB's physical capacities and its response to the model FIA. I found the MRB able to: 1) collect sample, 2) deliver appropriate amounts of liquid/solid reagent phase to the sample reproducibly, 3) rinse away possible interferents, and 4) measure the signal from the reacted complex--all without removing the sensing tip from the sample. I also found the MRB capable of measuring assay parameters such as: amount of total antibody-antigen binding, various analyte:labeled analyte ratios, and specific/nonspecific antibody binding. However, when liquid reagent flows over solid reagent (during MRB operation), I found that the interaction between the model reagents was inadequate to allow a demonstration of a sequential competitive-binding FIA.

These findings suggest that with further development, the MRB could allow 1) repetitive competitive-binding assays for near-continuous monitoring, and 2) multiple analyses within a single remote sample by employing alternate reagents.

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