Environmental nicotine contamination in latent fingermarks from smoker contacts and passive smoking.

Hydrophobic silica nanopowder has been used as an effective latent fingermark development agent and subsequently as an enhancement agent in the surface-assisted laser desorption/ionisation-time of flight (SALDI-TOF) mass spectrometry for analysis of fingermark components. The technique has been used in the detection of nicotine and cotinine in the fingermarks of smokers. In order to have confidence in concluding that the nicotine in such samples is indicative of cigarette usage, it is necessary to establish that contamination by environmental contact or from hand to hand contact with smokers or from passive smoking does not lead to false identification of non-smokers as smokers. To investigate this possibility, the background level of nicotine in fingermark material from a number of commonly used places was determined. In addition, a series of experiments was carried out to assess the extent to which nicotine can be transferred through handshakes and finger transfer as well as touching of door handles. The rate of loss of nicotine from latent fingermarks was also assessed over a 24-h period under ambient laboratory conditions. Finally, a laboratory-based model system was evaluated to ascertain the possible transport of nicotine in cigarette smoke from a source to adjacent areas to simulate cross-contamination of a non-smoker by passive exposure. It was observed that person-to-person transfer from a smoker to a non-smoker can occur following handshakes but at low levels and that passive cross-contamination from contact with surfaces is possible under simulated conditions. However, levels of nicotine in the wider environment were found to be too low for detection using this technique which may reflect the half-life of nicotine in latent fingermarks which was about 11h. Likewise, transfer via smoke is possible to objects within about 0.1m of the cigarette but it is unlikely that significant secondary nicotine contamination will occur on the faces and hands of adjacent non-smokers.

Direct on-filter assay of subtilisin-type proteolytic enzymes for rapid analysis of analyte captured from the workplace atmosphere.

A simple assay for some proteolytic enzymes has been developed which can be performed directly on the surface of a cellulose nitrate filter used to capture the analyte during workplace monitoring for health and safety purposes. Following air sampling the analysis is performed on the filter which is retained within the air sampler. This involves two steps: first, a 15 min incubation in which the captured enzyme is dissolved and then digests an alkaline-phosphatase-labelled antibody immobilised as a small dot on the surface of the filter; and second, is a 10 min incubation with substrate solution, which follows an in situ wash under a vacuum. During the incubation colour develops on the spot at the location of the immobilised enzyme antibody conjugate. The intensity of the spot can be assessed visually within the sampler to ascertain the presence or absence of captured enzyme, or alternatively quantitative results can be obtained using an optical scanner. The limit of detection is 5 ng per filter for subtilisin (20 ng for visual discrimination between this standard and the zero). The assay is stable to the effects of ambient air sampling at 31 min(-1) for 18 h.

Mathematical analysis for impedance change with proteolytic digestion of gelatin.

A mathematical analysis is proposed to demonstrate an inter-relationship between the proteolytic digestion of gelatin on the surface of an interdigitated gold electrode and the resulting rate of impedance change, at different collagenase concentrations, in a biosensor used to detect protease in solution. The impedance change due to digestion of the gelatin layer by collagenase for the overall digestion process was expressed in two different stages: an initial exponential period where the rate of impedance change with enzymic digestion was slow, leading to a critical thickness; after which there was a greater change in impedance associated with subsequent dissolution of the layer and partial or complete uncoating of the digits on the electrode surface. An inter-relationship between the rate of impedance change and collagenase concentration within the range 0.2-0.6 mg ml-1 was predicted for the early stages of the digestion process. A kinetic theory for the rapid rate of impedance change with collagenase concentrations could not be developed owing to the rate remaining almost constant for all concentrations of collagenase, after the critical thickness had been reached. An inter-relationship between the rate of impedance change and stirrer speed was also demonstrated.

Detection of protease activity in the wetted surface of gelatin-coated electrodes in air by AC impedance spectroscopy.

As a step towards developing a biosensor which can detect airborne protease droplets, a biosensor which had previously been developed to detect protease in solution is shown to be capable of detecting different concentrations of protease in liquid films on the sensor surface in air. The biosensor measured impedance change due to proteolytic digestion of its gelatin coating. In saturated air there was a rise in impedance, with a loss in weight of the gelatin, in proportion to collagenase concentration. The addition of glycerol to the gelatin caused a lower impedance response and smaller loss in weight. A critical thickness of the gelatin layer prior to a more rapid change in the rate of impedance was noted, with and without the addition of glycerol. In low air humidity (40%), with gelatin, all collagenase concentrations produced a very similar rapid increase in impedance. However, with glycerol-enhanced gelatin, there was a clear distinction between the extent of impedance change with different collagenase concentrations. The application of these findings for use in the field of bioaerosol sampling is discussed.

Enzyme-linked immunosorbent assay for ondansetron captured from airborne samples.

A simple and relatively rapid enzyme-linked immunosorbant assay method for the anti-emetic drug ondansetron has been developed for its quantitation in solution. This has been optimised for use with samples that have been obtained following extraction of filters after the drug's capture from air samples in the workplace. The assay has the sample throughput (40 duplicate samples in 3 h), specificity, sensitivity (LOD of 10.5 ng drug ml-1) and precision (RSD < 11%) necessary for its use in determining airborne concentrations of ondansetron in such samples as part of an occupational health and hygiene monitoring programme.

Recovery of some common solvents from an adhesive commercial skin adsorption pad by thermal desorption-gas chromatography.

A thermal desorption-gas chromatography (GC) system was developed for use with commercial adhesive plasters used for monitoring exposure of hands to common solvents. The efficiency of solvent adsorption on the activated carbon pads located on the plasters was determined for acetone, trichloroethylene, D-limonene, methanol, ethyl methyl ketone, toluene, tetrachloroethylene and m-xylene. The degree of solvent recovery for the system was also investigated for each solvent, as was its sensitivity and reproducibility. All solvents exhibited > 90% adsorption on the pads at spiking levels of 100-200 ng for each solvent, except for m-xylene and d-limonene. Solvent recovery was dependent on the volatility of the solvent at spiking volumes of about 1 microliter per pad with solvents with boiling points above 110 degrees C showing recoveries of < 75%. Increasing primary desorption times and temperatures increased these values. The precision was good with RSD < 5% for all solvents over the range 0.5-5.0 microliters of applied solvent. It was possible to detect 15-60 ng of each solvent component within solvent mixtures on the pads with the exception of D-limonene. It is concluded that all solvents tested except D-limonene can be determined on the pads under the conditions for thermal desorption-GC analysis described. The pads were used under protective gloves with six workers using xylene isomers as solvent in the workplace, when apparent solvent breakthrough through their gloves was observed.

Direct on-filter immunoassay of some beta-lactam antibiotics for rapid analysis of drug captured from the workplace atmosphere.

A simple competitive enzyme-linked immunoassay for the antibiotic ceftazidime and structurally similar beta-lactam antibiotics has been developed which can be performed directly on the surface of a cellulose nitrate filter used to capture the airborne drug during workplace monitoring for health and safety purposes. Post sampling analysis is performed on the filter retained within the air sampler. It involves two steps; the first a 10 min incubation in which the captured drug is dissolved and competes with drug immobilised within a protein conjugate on the surface of the filter for an enzyme-labelled antibody reagent, and the second, following washing under vacuum in situ, a 5 min incubation of substrate solution when colour develops on the spot at the location of the immobilised drug-protein conjugate. The intensity of the spot can be assessed visually within the sampler to ascertain the presence or absence of captured drug, or quantitative results can be obtained using an optical scanner. The intensity of the spots in linear from 10 ng to 1 microgram (r2 = 0.9996, n = 3) and the limit of detection is 1.9 ng of captured drug (10 ng for visual discrimination between this standard and the zero). The assay is precise with between-assay RSD values of < 4% over the linear range of the assay.

Enzyme linked immunosorbent assay for detecting benzodiazepines in urine.

A relatively simple ELISA technique was developed for the detection of a range of benzodiazepines (BZs) in urine. The assay employs a mouse anti-oxazepam antibody that is highly specific for the BZs. The limit of detection using 10 microliters samples of urine was 0.3 microgram ml-1 oxazepam. N-Desmethyldiazepam showed equal cross-reactivity to oxazepam, 11 BZs cross-reacted weakly and flurazepam and chlordiazepoxide did not cross-react at levels reported to be found in urine. No cross-reactivity was observed with drugs of abuse and a range of therapeutic drugs commonly found in urine. The assay was used as a screen to detect the presence of BZs in urine from 88 addicts that had been screened by the EMIT technique and a radioreceptor assay (RRA) for BZs. The ELISA produced two false negatives that were EMIT and RRA positive whereas the EMIT produced four different false negatives that were positive by both ELISA and RRA. Thirty-three positives were common to all three assays. The ELISA was also used to monitor nitrazepam-like activity in the urine of a greyhound receiving 5 mg oral medication and the results were compared with those obtained by RRA. Both assays were able to detect nitrazepam-like activity for up to 10 h post-administration.

Direct determination of some phenothiazine sedatives in greyhound urine by fluoroimmunoassay.

Antibodies have been raised in rabbits to a chlorpromazine sulfoxide-bovine serum albumin immunogen and the resulting antiserum used to develop a magnetizable solid-phase antibody fluoroimmunoassay for the detection of sulfoxide metabolites of commonly used phenothiazine and thioxanthine neuroleptics. These assays were used to screen metabolite levels in the urine of a greyhound following oral medication with chlorpromazine in order to assess the potential of these assays as simple screens for detecting exposure of racing greyhounds to such sedatives. The urine samples were also screened for neuroleptic content using an established radioreceptor assay and by TLC. The immunoassay described represents a relatively simple, sensitive and group-specific alternative method for screening for medication with phenothiazine and structurally similar sedatives in urine samples.

Enzyme linked immunosorbent assay for ceftazidime in airborne samples.

A simple and rapid enzyme linked immunosorbent assay (ELISA) method for the cephalosporin, ceftazidime, has been developed for the quantification of this antibiotic in solutions that have been eluted from filters following the capture of air samples in the workplace. The assay has the specificity, sensitivity and precision necessary for its use in determining airborne concentrations of ceftazidime in the workplace as part of an occupational health and hygiene programme.

Development of an enzyme-linked immunosorbent assay for the detection and measurement of the trypanocidal drug isometamidium chloride in cattle.

An enzyme-linked immunosorbent assay (ELISA) was developed to measure accurately levels of the trypanocidal drug isometamidium in the serum of treated cattle. The assay requires only 5 microliters of test serum, is sensitive to a level of 0.5 pg ml-1 and is highly specific. Cross reactivity does not occur with the two other widely used trypanocidal drugs diminazene aceturate and homidium bromide. Serum drug levels are detectable for up to six months in cattle after a single dose of 1 mg kg-1 intramuscularly, the maximum period under field conditions for which effective prophylaxis can be maintained against tsetse challenge. Application of the assay will aid the rationalisation of treatment campaigns and assist in assessing the occurrence of drug-resistant trypanosome populations.

Urinary chromatographic profiles in schizophrenia.

In a series of articles, Trygstad et al. (1980) suggested that specific patterns of peptide excretion and increased levels of peptide material are found by chromatographic analysis of the urine of patients with a variety of psychiatric illness including schizophrenia. As these results have not been independently replicated in another laboratory, we conducted an investigation of urine samples from 5 DSM-III classified schizophrenic patients and 4 normal subjects using the techniques described by the Norwegian group, and with a series of modifications. The chromatographic profiles obtained differ widely from those reported by Trygstad et al. No significant differences were detected between patients with schizophrenia and controls. The methods used by Trygstad et al. are complex and we have defined several parts of the methods which are subject to variability. Our findings lend no support to the view that patients with psychiatric illness can be readily distinguished from normal subjects by the amount or profile of peptide excretion in urine.

Development of a magnetisable solid-phase fluoroimmunoassay for primaquine and carboxyprimaquine.

Primaquine coupled to keyhole limpet hemocyanin was used as an immunogen to produce antiprimaquine antibodies in three sheep. The antisera obtained were characterised by the increase in fluorescence polarisation found upon binding to fluorescein-labelled primaquine prepared via same route. All sheep showed a good antibody response and one antiserum was coupled to magnetisable solid-phase particles to facilitate the separation of the antibody bound from free labelled antigen and the removal of interfering components which may be present in the sample. The fluoroimmunoassay requires addition of 100 microliters of standard or sample (urine or serum) to 100 microliters tracer (150 nmol/l) followed by 100 microliters of magnetisable solid-phase particles (12.5 g/l). After one hour incubation followed by the usual washing and eluting procedures, using a magnetic rack, the fluorescence of the supernatant was measured directly in a fluorimeter. Sodium salicylate was incorporated in the tracer solution to block the non-specific binding of tracer to the protein in serum samples. Cross-reactivity studies showed that the antibodies have high specificity for the 8-aminoquinoline nucleus but not to the 8-N-aminobutyl side chain. Thus carboxyprimaquine cross-reacted equally with primaquine and the assay can be used to measure their combined level. After extraction of primaquine from a basified sample with methylene chloride, the assay may be applied for the quantitation of either primaquine (in the organic phase) or its acidic metabolites including carboxyprimaquine (in the aqueous phase) separately. This approach was applied for the determination of total primaquine (primaquine and its metabolites) and extracted primaquine in urine samples following a single oral dose of 45 mg primaquine.(ABSTRACT TRUNCATED AT 250 WORDS)

A rapid, qualitative ELISA test for the specific detection of morphine in serum or urine.

We describe a competitive inhibition ELISA technique, with a visual end-point, to detect free morphine in blood or urine. It has a sensitivity of 2 X 10(-7) mol/l using 5 microliter samples. No significant cross-reactivity was observed with other opiate derivatives. The assay has applications as a specific screen for morphine in drug abusers, or to study the metabolism of the drug in the body (as the metabolite, morphine-3-glucuronide, does not cross-react significantly with morphine in the assay).

A competitive enzyme-linked immunosorbent assay for alpha 1-acid glycoprotein in serum and urine samples.

In this solid-phase competitive enzyme-linked immunosorbent assay for alpha 1-acid glycoprotein in serum or urine, antiserum to human alpha 1-acid glycoprotein is incubated with solid-phase-bound alpha 1-acid glycoprotein in the presence of standard or sample. Incubation with second antibody labeled with alkaline phosphatase then follows, before development with substrate. Results obtained correlate well with a fluorescent assay involving the dye Auramine O (r = 0.953) and with radial immunodiffusion (r = 0.921). The present assay covers the range 0.2 to 5 mg/L and 16 samples take 2.5 h to complete. This assay is useful for measuring concentrations of alpha 1-acid glycoprotein in serum and also in urine, for which other assay methods are not sufficiently sensitive.

A specific ELISA method for determining chloroquine in urine or dried blood spots.

Reported is an enzyme-linked immunosorbent assay (ELISA) that has been optimized and validated for the determination of chloroquine in urine or dried blood spots. The assay employs antisera raised in sheep to a chloroquine derivative conjugated to keyhole limpet haemocyanin and chloroquine conjugated to porcine thyroglobulin adsorbed onto the wells of a microtitration plate. The competitive binding of the antiserum to the wells was monitored using an alkaline-phosphatase-conjugated second antibody and a specific substrate. The assay exhibits no cross-reactivity with known chloroquine metabolites, other antimalarials, and commonly used drugs. The method was used to determine chloroquine in dried blood spot extracts and urine from a patient who was receiving a prescribed prophylactic chloroquine regimen. The drug was detected in the urine for 17 weeks and in the dried blood spots for 4 weeks after termination of the therapy.

Reduction of prostaglandin E2 concentrations in synovial fluid of patients suffering from rheumatoid arthritis following tiaprofenic acid or indomethacin treatment.

Prostaglandin E2 (PGE2) is a potent stimulator of inflammation, and the inhibition of its synthesis is one possible mechanism of action of non-steroidal anti-inflammatory drugs (NSAIDs). We have investigated patients suffering from rheumatoid arthritis to determine how synovial fluid levels of PGE2 are affected by tiaprofenic acid or indomethacin medication. Ten patients suffering from rheumatoid arthritis were studied, with 5 patients receiving tiaprofenic acid and 5 indomethacin for a 1-week period. Synovial fluid and serum samples were collected over an 8-hour period on days 1 and 8; these were then assayed for PGE2 and active drug concentrations. The concentration of PGE2 in the synovial fluid fell consistently as the concentration of each drug rose, and low levels of PGE2 persisted on continuation of the medication. Tiaprofenic acid appeared to cause a faster onset of inhibition of PGE2 synthesis than indomethacin.