Digoxin immunoassay with monoclonal and polyclonal antibodies using time-resolved fluorometry

Area of Science:

• Biochemistry
• Immunology
• Analytical Chemistry

Background:

• Accurate digoxin level monitoring is crucial for therapeutic drug management.
• Existing immunoassays may have limitations in sensitivity or specificity.
• Development of novel, reliable digoxin assays is an ongoing area of research.

Purpose of the Study:

• To describe a novel nonisotopic heterogeneous competitive immunoassay for serum digoxin.
• To evaluate the performance of both polyclonal antibody Fab fragments and high-affinity monoclonal antibodies in this assay.
• To establish a simple and effective method for routine digoxin determination.

Main Methods:

• A competitive immunoassay format was developed using immobilized digoxin and biotinylated immunoreactants (Fab or monoclonal antibody).
• Quantification involved streptavidin labeled with a europium chelator (BCPDA) and time-resolved fluorometry.
• The assay was performed in white polystyrene microtiter wells with direct measurement on the solid phase.

Main Results:

• The assay demonstrated an inverse relationship between bound biotinylated moiety and serum digoxin concentration.
• Both polyclonal Fab fragments and monoclonal antibodies yielded comparable and effective assay performance.
• The developed immunoassay is simple to perform with characteristics similar to existing techniques.

Conclusions:

• High-affinity monoclonal antibodies are suitable for routine serum digoxin determination.
• The described nonisotopic immunoassay offers a viable alternative for clinical laboratories.
• The assay's simplicity and comparable performance make it a practical option for digoxin monitoring.