Determination of Morphine in Human Urine by the Novel Competitive Fluorescence Immunoassay

Abstract

A competitive fluorescence immunoassay for the identification and quantification of morphine has been developed on the basis of hapten-coated plate format. Hapten was prepared through covalent conjugating a morphine derivative with albumin bovine. In the immunoassay, the hapten was inoculated on a 96-well plate and then bound with monoclonal antibodies labeled with a signal indicating dye, fluorescein isothiocyanate (FITC). Unbound FITC-antibodies were rinsed off from the plate. The fluorescein intensity decreases in the presence of morphine molecules due to the competitively binding to antibodies against hapten. The intensity is inversely correlated with the concentration of morphine. In quantitative analysis for urine samples, we obtained a linearity range of 0.2 μg/mL∼2.5 μg/mL, along with a detection limit of c.a. 1 ng/mL. The fluorescence immunoassay shows low cross-reactivity (below 10%) to 6-acetylmorphine, 3-acetylmorphine, and heroine. The developed method produced comparable results to the standard GC-MS/MS method. In conclusion, a rapid and efficient screening tool for morphine in clinical human urine has been established.

Emerging Technologies for Fluorescence-Based Optical Test Strip Readers

“…There are many different types of fluorescent dyes. Rhodamine dyes (Rhodamine B, excitation/emission wavelength: 554/570 nm) which are known for their excessive illumination [34,35] and photostability [36,37] are preferred for medical imaging including fluorescence microscopy [38], flow cytometry [39] as well as diagnostic tests [40]. Cyanine dyes (Cy5, excitation/emission wavelength: 649/670 nm) are highly sensitive to changes in environmental conditions, such as changes in pH value or the presence of specific molecules, and therefore are often used in biological imaging, including live-cell imaging, and fluorescence resonance energy transfer assays [41][42][43].…”

Monoclonal Antibodies Application in Lateral Flow Immunochromatographic Assays for Drugs of Abuse Detection

“…They are used for the determination of ABO and rhesus blood groups, for HLA tissue grouping, for the immunolabeling of acute leukemia and for the development of immunological tests (enzyme-linked immunosorbent assay (ELISA), lateral flow immunoassay (LFIA), radioimmunoassay (RIA), etc.) [ 18 , 19 , 20 , 21 , 27 , 31 , 55 , 56 , 57 , 58 , 59 , 60 ]. Other Abs issued from animals conjugated to markers or enzymes are used for diagnostic kits manufacturing, immunocytochemical analysis, and research [ 18 , 19 , 20 , 21 , 27 , 31 , 55 , 56 , 57 , 58 , 60 , 61 , 62 ].…”

“…Moreover, a panel of mAbs is usually used to map and study the role of epitopes in certain cellular functions and mechanisms. They have also an important role in proteomics and mass biological screening tests [ 18 , 19 , 20 , 21 , 31 , 43 , 55 , 56 , 58 , 59 ]. In general, mAbs are used in diagnostic, agri-food, veterinary, microbiological, and toxicological tests [ 20 , 21 , 31 , 56 , 58 , 63 ].…”

Europium Nanoparticles-Based Fluorescence Immunochromatographic Detection of Three Abused Drugs in Hair

“…In order to meet the requirements of a more accurate and sensitive analysis, the combination of fluorescent microspheres as labelling probes and portable strip readers has become a research hotspot in recent years. Quantum dots (QDs), carbon nanodots-silica [ 33 ], upconverting nanoparticles [ 34 ], fluorescein isothiocyanate (FITC) [ 35 ] and lanthanide-labeled fluorescent nanoparticles [ 36 ], as substitutes for colloidal gold, have been widely used in applications in clinical detection [ 37 ], molecular biology [ 38 ] and other related fields. Europium nanoparticle (EuNP) luminous material has been widely employed because of its benefits of ultra-long emission half-life, massive Stokes shift, longer measurement duration, removal of background fluorescence and better detection sensitivity [ 39 ].…”