Geoffrey S. Rule

High-throughput sample preparation and analysis using 96-well membrane solid-phase extraction and liquid chromatography-tandem mass spectrometry for the determination of steroids in human urine

A 96-well solid-phase extraction (SPE) system is used to rapidly prepare human urine samples for high-throughput quantitative analysis of two steroids, equilenin and progesterone, by liquid chromatography-tandem mass spectrometry using deuterated estrone as the internal standard. We define high-throughput here as analysis of 384 samples in a 24 h period. A total of 384 samples and standards were extracted by an individual in one day and subsequently analyzed within a 24 h period. The inter- and intratray accuracy and precision obtained over the course of these injections was within 8% coefficient of variation when analyzed by atmospheric pressure chemical ionization mass spectrometry using positive ion detection. A semiautomated sample processing workstation was used to add internal standard and then process 96 samples at a time. The recovery of the analytes from the SPE was approximately 85%. The accuracy and precision obtained was comparable to that ordinarily obtained using manual sample preparation techniques.

Determination of drugs from urine by on-line immunoaffinity chromatography—high-performance liquid chromatography—mass spectrometry

A method for rapid extraction and identification of drugs in urine is described. The system utilizes a high-performance protein G immunoaffinity column coupled to a reversed-phase analytical column by use of a trapping column and switching valve. A small amount of antibody (5 micrograms drug-specific) is used for each analysis to extract either propranolol or lysergic acid diethylamide (LSD) from human urine. Urine diluted with phosphate-buffered saline is pumped directly through the protein G column thus eliminating time- and solvent-consuming sample preparation procedures. On-line ultraviolet or mass spectral analysis provides the means of drug detection and identification. With ultraviolet detection propranolol may be detected in spiked urine at the 250 pg/ml level. A Hewlett-Packard mass spectrometer modified for atmospheric pressure ionization and equipped with an ion spray source allows detection of propranolol in urine at 2.5 ng/ml and LSD at 500 pg/ml using single ion monitoring. The potential applicability of the technique for drug confirmations is discussed.

Flow-injection liposome immunoanalysis (FILIA) for alachlor.

An automated Flow-Injection Liposome ImmunoAnalysis (FILIA) system has been modified from a previous design, using a specific environmental contaminant, the herbicide alachlor, as a model analyte. Signal amplification by means of fluorescent marker-loaded, analyte-tagged liposomes provides high sensitivity. The computer controlled system is composed of commercially available components, with the exception of the column packing material, which has to be prepared for each specific analyte to be determined. The use of such components means that the system is easily modified. The relationships between antibody concentration and assay speed and sensitivity are explored, and the possibilities of using the system for determination of multiple analytes is discussed.

Use of protein A in a liposome-enhanced flow-injection immunoassay

A flow-injection immunoassay for the herbicide alachlor has been developed in a sequential format using both a protein A matrix and liposomes. The protein A matrix is a commercially available controlled-pore glass that serves as the support for antibodies. The liposomes are tagged with a phospholipid–analyte conjugate and encapsulate a fluorescent dye that generates the signal. This report is the first demonstrated combination of these two technologies.