Determination of Amitriptyline by Fluorescence Polarization Immunoassay

We developed a procedure for determining tricyclic antidepressant amitriptyline using fluorescence polarization immunoassay against the background of a buffer solution, urine, and breast milk with a relative standard deviation of no more than 5%. Tracers were synthesized by the noncovalent adsorption of amitriptyline on the surface of silicate nanoparticles doped with ruthenium(II) complexes. For the practical application of the proposed method, we selected the optimal concentrations of the tracer and antibodies by dilution. The amounts of reagents were selected based on fluorescence intensity and fluorescence polarization. According to the sensitivity requirements of the tracer, a dilution of 1 : 2 was selected, the fluorescence intensity of which was ten times higher than the background value, while the optimal fluorescence polarization value was fixed at 32. The optimal dilution of antibodies, corresponding to 70% of the fluorescence polarization value from the maximum signal, was 1 : 200. The range of working concentrations of amitriptyline was 1 × 10 –10 –1 × 10 –5 M; the lower limits of determination were at a level of (7–8) × 10 –11 M. The results are promising for monitoring the concentration of amitriptyline both in the pharmaceutical preparation and biological fluids to determine the effectiveness of ongoing therapeutic treatments and for medical and forensic purposes.