sepharose and benzoylecgonine

Storage and quantitative analysis of small volumes of biofluids are challenging, especially when low concentrations of analytes are to be detected in the presence of complex matrices. In this study, we describe an integrated thread-based approach for stabilizing small blood volumes in the dry-state at room temperature, while also offering direct analysis capabilities via thread spray mass spectrometry. The analytical merits of this novel microsampling platform was demonstrated via the direct analysis of diazepam and cocaine in dried blood samples stored for 42 days. In-situ in-capillary blood processing from hydrophobic threads enabled limits of detection as low as parts-per-quadrillion to be reached. We validated this ultra-sensitivity by analyzing small tissue-like residues collected after pushing a thread through the sample once. The implications of this sample collection, storage, and analysis platform can be extensive with direct applications in forensics and clinical studies.

Topics: Amphetamine; Cocaine; Cotton Fiber; Diazepam; Dried Blood Spot Testing; Gossypium; Humans; Limit of Detection; Mass Spectrometry; Methamphetamine; Sepharose; Solid Phase Extraction; Specimen Handling

This study investigates the effect of antibody density on the kinetics of a solid-phase displacement immunoassay. Conducted in flow under nonequilibrium conditions, the assay utilizes a monoclonal antibody to the cocaine metabolite benzoylecgonine, which has been immobilized onto Sepharose beads and saturated with fluorophore labeled antigen. Displacement of antibody-bound labeled antigen by non-labeled antigen occurs when sample is introduced in the buffer flow. Comparison of matrices coated with two different antibody densities revealed that the displacement efficiency is a function of the density of antibody-bound labeled antigen. A higher density of antibody provides a higher amount of displaced labeled antigen, but the displacement efficiency of the assay is decreased. The effect of antibody density on the immunoassay kinetics was analyzed using a mathematical formulation developed to characterize antibody-antigen interactions at solid-liquid interfaces. Higher antibody density proved to be associated with a lower apparent dissociation rate constant. The implications of these results on the design of immunoassays in flow are discussed.

Topics: Animals; Antibodies, Monoclonal; Antigens; Cocaine; Immunoassay; Kinetics; Mice; Sepharose