Gentamicin substrate-labeled fluorescent immunoassay containing monoclonal antibody

Area of Science:

• Biochemistry
• Immunology
• Analytical Chemistry

Background:

• Gentamicin therapy requires precise therapeutic drug monitoring to maximize efficacy and minimize toxicity.
• Conventional immunoassays for gentamicin often rely on polyclonal antisera, which can exhibit variability.

Purpose of the Study:

• To develop and validate a modified substrate-labeled fluorescent immunoassay (SLFIA) for gentamicin using monoclonal antibodies.
• To assess the performance characteristics of the new assay, including linearity, sensitivity, precision, and reproducibility.

Main Methods:

• Substitution of conventional antiserum with monoclonal antibodies against gentamicin in a substrate-labeled fluorescent immunoassay.
• Generation and analysis of standard curves to determine assay linearity and sensitivity.
• Evaluation of intra- and interassay precision using coefficients of variation.
• Comparison of results from the developed assay with existing methods (commercial SLFIA, EIA, RIA, FIA) using clinical specimens.

Main Results:

• The gentamicin monoclonal antibody-based SLFIA demonstrated excellent linearity from 0 to 12 µg/ml.
• The assay achieved a sensitivity of 0.4 µg/ml for gentamicin detection.
• High precision was observed with intra- and interassay coefficients of variation of 1.9% and 5.0%, respectively.
• Results from the new assay showed high correlation (r = 0.991) with established gentamicin measurement techniques.

Conclusions:

• The modified SLFIA utilizing monoclonal antibodies provides a precise, sensitive, and reproducible method for gentamicin quantification.
• This nonisotopic immunoassay offers a technologically advanced alternative for therapeutic drug monitoring of gentamicin.
• The assay exhibits excellent lot-to-lot reproducibility, linearity, and reagent stability, making it suitable for clinical use.