Development and preliminary evaluation of a fiber optic-based microscale biosensor for performing remote competitive-binding fluoroimmunoassays

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.