Quantification of Opioids in Urine Using an Aptamer-Based Free-Solution Assay

Area of Science:

• Clinical Chemistry
• Biomedical Diagnostics
• Forensic Toxicology

Background:

• The ongoing opioid epidemic significantly impacts neonates, leading to Neonatal Abstinence Syndrome (NAS).
• Current diagnostic methods for neonatal opioid exposure are limited by poor assay performance, false-negatives, and insufficient sample volumes.
• Rapid and sensitive testing is crucial for timely NAS diagnosis and management.

Purpose of the Study:

• To develop and validate a novel, small-volume urine assay for quantifying opioids and their metabolites.
• To assess the assay's sensitivity, specificity, and accuracy compared to existing methods like LC-MS/MS and immunoassays.
• To explore the potential of this assay for noninvasive opioid screening in neonates and pharmacokinetic tracking.

Main Methods:

• Development of a free-solution assay (FSA) utilizing high-affinity DNA aptamer probes.
• Integration of a compensated interferometric reader (CIR) for sensitive detection.
• Quantification of oxycodone, hydrocodone, fentanyl, and their urinary metabolites in small urine volumes (10 μL).

Main Results:

• The FSA demonstrated excellent limits of detection (LODs) and quantification (LOQs) for target opioids and metabolites.
• The assay exhibited 5- to 275-fold higher sensitivity than LC-MS/MS and 50- to 1250-fold higher sensitivity than immunoassays.
• Accurate quantification (>95% specificity) was achieved in approximately 1 hour, with concentrations ranging from 28-81 pg/mL.

Conclusions:

• The developed FSA offers a highly sensitive, rapid, and noninvasive method for opioid quantification using minimal urine samples.
• This assay holds significant potential for improving the diagnosis and management of Neonatal Abstinence Syndrome (NAS).
• The technology could facilitate widespread opioid screening and enable precise pharmacokinetic tracking in clinical settings.