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PubMed1982

Behaviour of immune complexes and the complement system in normal pregnancy and pre-eclampsia.

A quantitative study of the circulating immune complexes (IC) was carried out on women during normal pregnancy (286) and the post-partum period (20) and women with pre-eclampsia (30). Furthermore, the behaviour of the complement (C) system was followed. Results showed that IC were low in the first trimester of normal pregnancy (25.3%) and decreased in the following trimesters, whereas they were always present in pre-eclampsia. A very significant difference (p less than 0.0001) was seen when we compared the incidence of IC in normal pregnancy at the third trimester and the pre-eclamptic patients. The follow-up study of the IC, carried out on 4 pre-eclamptic women, showed an increase in the IC levels associated with the exacerbation of the pre-eclamptic picture and a decrease after delivery. The study of complement in normal pregnancy showed a decrease in C1-INH, C1s and C1q, whereas C3, C5, C9 and the properdin factor B increased during the following weeks of gestation; CH50 did not vary excepting during the 1st trimester. In the puerperium all values increased. There was no significant difference between the serum levels of the C components in the 3rd trimester of normal pregnancy and pre-eclampsia. High levels of C3d were observed in normal pregnancy at the 3rd trimester and in pre-eclampsia. The study of this split product of C3 showed that there is activation of the C system, but, since the synthesis of the C components is increased, activation could be masked. Alloantibodies and circulating IC could be the factors responsible for this activation in normal pregnancy and in pre-eclampsia, respectively.

Adult↗

PubMed1989

Failure of Bacteroides gingivalis W83 to accumulate bound C3 following opsonization with serum.

Our previous studies have demonstrated that strains of Bacteroides gingivalis are capable of proteolytic degradation and inactivation of complement proteins including the third component of complement C3. Since a crucial step in the ability of complement to control bacterial infections is the binding of C3 fragments to the bacterial surface with subsequent enhancement of phagocytosis, further examination of the importance of the proteolytic capacity of Bacteroides in interactions with complement proteins was carried out by quantitating the amount of C3 bound to two proteolytic Bacteroides gingivalis strains. Pooled normal human serum (NHS) containing 125I-C3 was incubated with strains of B. gingivalis (W83 and ATCC 33277) and the non-proteolytic pathogen A. actinomycetemcomitans strain Y4, and samples of the reaction mixtures were removed at various time intervals for determination of bound C3. B. gingivalis 33277 bound only half the number of C3 molecules as did A. actinomycetemcomitans, while B. gingivalis W83 bound very little C3. A large increase in the number of C3 molecules bound to B. gingivalis W83 was noted in assays carried out in the presence of the protease inhibitor TLCK, indicating that bacterial proteases may be responsible for the lack of binding of C3 to strain W83. TLCK treatment modestly increased the accumulation of C3 on strain 33277, but had no effect on A. actinomycetemcomitans. Analysis of 125I-C3 in supernatants from reaction mixtures of strain 33277, W83, or a proteolytic strain of B. intermedius demonstrated no qualitative differences in the C3 fragments amongst the tested strains or in the presence or absence of TLCK.(ABSTRACT TRUNCATED AT 250 WORDS)

Actinobacillus↗

PubMed1972

Indirect fluorescent-antibody and quantitative agar-gel immunodiffusion tests for the serological diagnosis of paracoccidioidomycosis.

The value of various serological tests in the diagnosis of paracoccidioidomycosis was studied. Quantitative agar-gel immunodiffusion and indirect immunofluorescent tests were performed, and the results were compared with those of complement fixation and qualitative agar-gel procedures. The quantitative immunodiffusion procedure was found to serve as the simplest and safest quantitative test that could be performed for evaluation purposes, whereas the indirect fluorescent-antibody test gave nonspecific reactions and, as such, proved unsuitable.

Agar↗

PubMed1986

Assembly of the membrane attack complex of complement on small unilamellar phospholipid vesicles.

Light-scattering intensity was shown to be a reliable, direct, and quantitative technique for monitoring the assembly of the membrane attack complex of complement (proteins C5b-6, C7, C8, and C9) on small unilamellar phosphatidylcholine vesicles. The assembly on vesicles occurred in a simple fashion; complexes of C5b-7 bound noncooperatively to the vesicles, and final assembly of C5b-9 did not induce vesicle aggregation or fragmentation. When C5b-6 and C7 were mixed in the presence of vesicles but at molar protein/vesicle ratios of less than 1, there was quantitative binding of C5b-7 to the vesicles with no concomitant aggregation of C5b-7. If C7 was added at a slower rate, quantitative binding was obtained at molar C5b-7/vesicle ratios of up to 5. The latter observations (a) were consistent with the proposal that C5b-7 aggregation and membrane binding were competitive events and (b) defined conditions under which light-scattering intensity measurements could monitor C5b-9 assembly on vesicles without contribution from the fluid-phase assembly. The C8/C5b-7 ratio in the phospholipid-C5b-8 complex was 0.97 +/- 0.12, and the maximum ratio of C9/C5b-8 in the final complex was 16.2 +/- 2.0. One C9 molecule associated rapidly with each phospholipid-C5b-8, followed by slower incorporation of the remaining C9 molecules. The initial velocity of the slow phase of C9 addition was easily saturated with C9 and gave an activation energy of 37 kcal/mol. This was identical with the value measured for the analogous process in the fluid-phase assembly.(ABSTRACT TRUNCATED AT 250 WORDS)

Complement C7↗

PubMed1993

Assessment of ram sperm mitochondrial function by quantitative determination of sperm rhodamine 123 accumulation.

A simple procedure is described for determining the functional state of ram sperm mitochondria by quantitative measurement of sperm rhodamine 123 (R 123) accumulation. Sperm were incubated with 1 microgram/ml R 123, and the accumulated R 123 was measured fluorimetrically after release from washed sperm by detergent lysis. Ram sperm R 123 uptake was maximal after 30 min of incubation and responded to changes in both sperm (P < 0.01) and R 123 (P < 0.01) concentration. There was a linear relationship (r = 0.98) between R 123 uptake and the proportion of cold-shocked sperm present in a sperm sample. R 123 uptake was unaffected by 20 mM 2-deoxyglucose or by 10 mM malonate (the latter being sufficient to reduce O2 uptake; P < 0.01). R 123 accumulation in ram sperm was reduced by 6 mg/ml sodium pentobarbitone (P < 0.05), by 1 microM 2,4-dinitrophenol (P < 0.01), and by 0.05% Triton X-100 (P < 0.01). It is concluded that quantitative estimation of R 123 uptake complements oxygen uptake in detecting mitochondrial dysfunction in ram sperm. While it is largely unaffected by inhibition of glycolysis, and is less sensitive than oxygen uptake to trichloroacetic acid cycle inhibition, R 123 uptake is sensitive to factors directly reducing the mitochondrial membrane potential of ram sperm. It may therefore by useful in the evaluation of the effects of such membrane-mediated injuries as cold shock and freezing damage on ram sperm mitochondria.

2,4-Dinitrophenol↗

PubMed2004

Quantitative profiling of plasma peptides in asthmatic mice using liquid chromatography and mass spectrometry.

Asthma is increasing in prevalence worldwide as a result of factors associated with a Western lifestyle. However, simple and reliable diagnostic and prognostic markers are yet to be found. In an attempt to identify protein biomarker profiles among small molecular weight ranges, we employed an approach combining liquid chromatography with mass spectrometry, instead of two-dimensional gel electrophoresis (2-DE), which has previously been used to analyze protein expression patterns. Here we described its application to compare plasma peptides from control and chronic asthma mice. Peptides were quantitatively profiled as a multidimensional peptide mass fingerprint by a combination of reverse-phase high-performance liquid chromatography and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. They were identified by peptide mass fingerprinting using matrix-assisted laser desorption/ionization time-of-flight/time-of-flight tandem mass spectrometry. In this study, we quantitatively identified the fragment f of complement 3 (C3f), which is important in inflammation. C3f was significantly higher in controls than chronic asthma mice. Our strategy allowed the detection and identification of different plasma peptides between control and chronic asthma mice on a proteomic scale. Therefore, these results suggest that native small peptides detected by non-2-DE techniques may be useful and specific biomarkers of disease.

Animals↗

PubMed1994

Using a computer model to explore impairments of acquisition processes following ingestion of diazepam.

Hypotheses about the information processes impaired in diazepam-induced amnesia were tested by fitting the output from a computer simulation of list learning to observed serial position curves and to overt rehearsal protocols. Twenty-four subjects received an average weight-relative dosage of 0.18 mg/kg oral diazepam; 24 subjects received placebo. Immediate free recall of 16-word lists was examined at 2- and 8-s presentation times. Subjects receiving diazepam recalled significantly fewer words than placebo subjects (diazepam = 6.77 +/- 2.39 words; placebo = 9.29 +/- 1.42 words); their memory impairment was greater at the 8-s than 2-s presentation time. Tests of nonlinear regression models based on computer simulations of list learning performance were consistent with the hypothesis that diazepam reduces rehearsal capacity and disrupts the formation or utilization of contextual and inter-item associations. Among these causes of diazepam-induced amnesia, the disruption of contextual associations appears most important. The results further suggest that quantitative modeling of memory data may complement traditional methods of inferring relationships between brain processes and cognitive dysfunction in amnesic states.

Adult↗

PubMed2001

[Quantification of retinal nerve fiber thickness. A comparison of laser scanning ophthalmoscopy, polarimetry and optical coherence tomography in healthy and glaucomatous eyes].

PURPOSE: To evaluate the thickness of the retinal nerve fiber layer (RNFL) in healthy eyes and in eyes of patients with primary open angle glaucoma using the Heidelberg retina tomograph (HRT), the nerve fibre analyser (NFA) and the optical coherence tomograph (OCT). METHODS: In this prospective cohort study, 40 normal eyes and 86 eyes of age-matched glaucoma patients were compared by confocal scanning laser tomography using the HRT, scanning laser polarimetry (NFA) and optical coherence tomography (OCT). The RNFL thickness was measured in the superior, inferior, nasal and temporal regions as well as the total circumference. RESULTS: All three methods revealed a statistically significant difference between normal and glaucomatous eyes with respect to the mean RNFL thickness in the inferior and superior regions (p < 0.001). The mean RNFL thickness in the superior region was 329 microns (HRT), 87 microns (NFA) and 94 microns (OCT) in healthy volunteers compared to 275 microns (HRT), 72 microns (NFA) and 82 microns (OCT) in the patient group. In the inferior region, it was 323 microns (HRT), 87 microns (NFA) and 93 microns (OCT) in healthy subjects versus 240 microns (HRT), 74 microns (NFA) and 83 microns (OCT) in glaucoma patients. Cut-off points to differentiate between normal and glaucomatous eyes could not be defined. There was no difference in the RNFL thickness of right and left eyes. CONCLUSIONS: In RNFL thickness measurements using HRT, NFA and OCT, glaucoma patients showed a significantly thinner RNFL in the superior and inferior areas compared to healthy volunteers. These results confirm the known histological and fundus photographic findings of RNFL thinning near the optic disc in glaucoma patients. Although RNFL thickness cannot be used to diagnose glaucoma in individual patients due to the high interindividual differences, the quantitative assessment of RNFL thickness may complement the diagnostic armamentarium as a sensitive parameter for diagnosing and monitoring glaucomas.

Adult↗

PubMed1987

Sensitive ELISA for quantitating the terminal membrane C5b-9 and fluid-phase SC5b-9 complex of human complement.

Activation of the complement system to completion results either in the generation of a pore-forming, cytolytic C5b-9(m) complex, or of a cytolytically inactive, fluid-phase SC5b-9 complex. In this paper, we describe a sensitive and reliable, sandwich ELISA for C5b-9(m) and SC5b-9, which is based on the use of a monoclonal antibody to a neoantigen of C5b-9 in combination with affinity-purified, polyclonal rabbit antibodies. The ELISA has been calibrated with purified C5b-9(m) and SC5b-9, and can detect 3 ng/ml C5b-9(m) and 20 ng/ml SC5b-9. We show that maximal conversion of C5-C9 in pooled human serum by insulin or zymosan activation generates 220 +/- 40 micrograms/ml SC5b-9. 65 of 100 normal human EDTA plasma samples analyzed in this study contained 100-600 ng/ml SC5b-9, corresponding to 0.04-0.24% of maximal conversion. Levels of circulating SC5b-9 in other donors were below the limit of detection. Incubation of serum at 37 degrees C always led to spontaneous generation of SC5b-9; concentrations ranged from 490-4725 ng/ml after 60 min, 37 degrees C, with a mean of 1848 +/- 1031 (SD) ng/ml amongst 25 donors studied. The terminal complement complex present in EDTA plasma was partially purified by PEG precipitation, DEAE-ion exchange chromatography and sucrose density gradient centrifugation, and was found to contain C8, C9 and S-protein as demonstrable by SDS-PAGE immunoblotting. Thus, the material most probably represented genuine SC5b-9. No significant age- or sex-dependent variations in SC5b-9 levels were noted. The present data call for a critical re-appraisal of several previously published methods for the determination of SC5b-9 levels in human plasma and serum.

Antibodies, Monoclonal↗

PubMed1984

Polychlorinated dibenzofurans (PCDFs): effects of structure on binding to the 2,3,7,8-TCDD cytosolic receptor protein, AHH induction and toxicity.

The effects of structure on the activity of 26 polychlorinated dibenzofurans (PCDFs) as competitive ligands for the 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) rat hepatic cytosolic receptor protein were determined in a dose-response fashion. The ED50 values for these compounds varied 100 000-fold and the most active PCDFs were substituted in the 2,3,7 and 8 lateral positions; the ED50 for the most active PCDF, 2,3,4,7,8-pentachlorodibenzofuran was 1.5 X 10(-8) M which was only slightly less active than 2,3,7,8-TCDD (1.0 X 10(-8) M). A comparison of the binding affinities of several isomer pairs also indicated the relative importance of chlorine substitution at C-4 (or C-6) compared to C-1 (or C-9). Moreover, for some isomers it is apparent that C-4 (or C-6) substituents are more active than lateral substituents for facilitating ligand binding to the receptor protein. This is illustrated by the relative binding potencies of the following isomer pairs: 1,2,4,6,7-/1,2,4,7,8 = 19.2; 2,6,7-/2,3,8- = 2.2; 1,3,6-/1,3,8- = 19. Most of the PCDF structure-activity effects noted above were also observed for the induction of aryl hydrocarbon hydroxylase (AHH) and ethoxyresorufin O-deethylase (EROD) in rat hepatoma H-4-II-E cells in culture. The most active compounds were also substituted in the lateral 2,3,7 and 8 positions and a comparison of C-4 (or C-6) vs. C-1 (or C-9) substituted PCDFs confirmed the higher induction potencies for most of the former group of compounds. The in vitro quantitative structure-activity data were complemented by in vivo studies which determined the relative activities of selected PCDFs as inducers of hepatic microsomal cytochrome P-448 dependent monooxygenases and their effects on body weight gain and thymus weights in immature male Wistar rats. The results indicated that for 2 series of isomers, namely the 2,3,4,7,8-, 1,2,4,7,8- and 1,2,4,7,9-pentachlorodibenzofurans and the 2,3,7,8-, 2,3,4,8- and 1,2,4,8-tetrachlorodibenzofurans, their biologic and toxic potencies were dependent on one major structural factor, the number of lateral chloro substituents. These results support the proposed role of the cytosolic receptor protein in mediating the biologic and toxic effects of the PCDFs.

Animals↗

PubMed2026

Development of a cell-based nanoluciferase reporter system for high-throughput screening of HBV cccDNA inhibitors.

Hepatitis B virus (HBV) persistence is sustained by the viral covalently closed circular DNA (cccDNA) minichromosome, which remains a major barrier to curative antiviral therapies. The lack of reliable quantitative cccDNA detection methods and surrogate markers has hindered efforts to target cccDNA in antiviral high-throughput screening (HTS). Here, we established a novel inducible cccDNA-dependent nanoluciferase (NLuc) reporter cell line, designated HepBLE12, by inserting an in-frame 11-amino acid split-NLuc HiBiT tag into the precore (pC) coding region of an HBV transgene. The resulting 1.3-kDa HiBiT tag on pC serves as the detection module of the split NLuc system, generating quantitative luminescence upon high-affinity complementation with the cognate 18-kDa LgBiT subunit in cell lysates. Notably, the HiBiT assay enables direct detection of intracellular HiBiT-pC protein rather than secreted HBeAg, providing a reporter signal more closely linked to cccDNA activity. HepBLE12 cells exhibited inducible and robust viral DNA replication, and the cccDNA-dependent HiBiT signal was validated under diverse experimental conditions that modulate cccDNA formation or transcription. We further miniaturized the assay to a 384-well format and optimized key parameters following standard HTS assay development practices. The assay was successfully automated and demonstrated excellent performance in a multi-day variability study and a pilot screen, with signal-to-background (S/B) ≈ 9, coefficient of variance (CV) < 10%, and average Z-factor value of 0.74, exceeding canonical HTS quality benchmarks. Together, the HepBLE12 cell-based HTS platform provides a robust and practical tool for identifying inhibitors targeting HBV cccDNA.

Hepatitis B virus↗

PubMed2006

Ultrasound lung comets: a clinically useful sign of extravascular lung water.

Assessment of extravascular lung water is a challenging task for the clinical cardiologist and an elusive target for the echocardiographer. Today chest x-ray is considered the best way to assess extravascular lung water objectively, but this requires radiology facilities and specific reading expertise, uses ionizing energy, and poses a significant logistic burden. Recently, a new method was developed using echocardiography (with cardiac probes) of the lung. An increase in extravascular lung water-as assessed independently by chest computed tomography, chest x-ray, and thermodilution techniques-is mirrored by appearance of ultrasound lung comets (ULCs). ULCs consist of multiple comet tails originating from water-thickened interlobular septa and fanning out from the lung surface. The technique requires ultrasound scanning of the anterior right and left chest, from the second to the fifth intercostal space. It is simple (with a learning curve of < 10 examinations) and fast to perform (requiring < 3 minutes). ULC assessment is independent of the cardiac acoustic window, because the lung on the anterior chest is scanned. It requires very basic 2-D technology imaging, even without a second harmonic or Doppler. ULCs probably represent an ultrasonic equivalent of radiologic Kerley B-lines. On still-frame assessment, cardiogenic watery comets can be difficult to distinguish from pneumogenic fibrotic comets, although the latter are usually more localized and are not dissolved by an acute diuretic challenge. Functionally, ULCs are a sign of distress of the alveolar-capillary membrane, often associated with reduced ejection fraction and increased pulmonary wedge pressure. The ULC sign is quantitative, reproducible, and ideally suited to complement conventional echocardiography in the evaluation of heart failure patients in the emergency department (for the differential diagnosis of dyspnea), in-hospital evaluation (for tailoring diuretic therapy), home care (with portable ultrasound), and stress echocardiography lab (as a sign of acute pulmonary congestion during stress). In conclusion, ULCs represent a useful, practical, and appealingly simple way to image directly extravascular lung water.

Artifacts↗

PubMed2005

Relative activity of the feline immunodeficiency virus promoter in feline and primate cell lines.

The feline immunodeficiency virus (FIV) long terminal repeat (LTR), compared with some primate lentiviral LTRs, is quite a strong basal promoter. However, it seems to be highly species-specific in function and generally not very efficient in cells of non-feline origin. This study systematically explored the function of the FIV LTR in simian Cos cells compared with its activity in feline and human cells. Our studies, using biologically relevant two- and three-plasmid trans complementation assays followed by semi-quantitative reverse transcriptase PCR, show that the FIV LTR is functional in Cos cells. The results of the Cos experiment are different from previously reported literature and suggest that the strain specificity of the FIV LTR is an important determinant of whether the LTR will be functional in a particular cell type.

Animals↗

PubMed2002

Quantitative femto- to attomole immunodetection of regulated secretory vesicle proteins critical to exocytosis.

Although immunoblotting (Western blotting) is widely used for the detection of specific proteins, it is often thought to be an inadequate technique for accurate and precise measurements of protein concentration. However, an accurate and precise technique is essential for quantitative testing of hypotheses, and thus for the analysis and understanding of proposed molecular mechanisms. The analysis of Ca(2+)-triggered exocytosis, the ubiquitous eukaryotic process by which vesicles fuse to the plasma membrane and release their contents, requires such an unambiguous identification and a quantitative assessment of the membrane surface density of specific molecules. Newly refined immunoblotting and analysis approaches permit a quantitative analysis of the SNARE protein complement (VAMP, SNAP-25, and syntaxin) of functional secretory vesicles. The method illustrates the feasibility of the routine quantification of femtomole to attomole amounts of known proteins by immunoblotting. The results indicate that sea urchin egg secretory vesicles and synaptic vesicles have markedly similar SNARE densities.

Animals↗

PubMed1991

Separation of DNA restriction fragments by capillary electrophoresis using coated fused silica capillaries.

The abilities of several different capillary electrophoresis techniques to separate DNA restriction fragments up to 23,000 bp were investigated. Methods employing electroosmotic flow in an untreated silica capillary were found to provide, at best, only partial resolution of the 23 fragments in a 1-kbp DNA ladder. By coating the inner walls of a silica capillary with poly(acrylamide) and filling these capillaries with buffers containing methylcellulose as a sleving medium, all fragments in the 1-kbp DNA ladder were separated. In addition, this technique facilitated the separation of the very large fragments in a lambda DNA-HindIII digest. Optimum resolution was obtained at low separation potentials using buffers containing at least 0.5% methylcellulose. The performance of this technique, i.e., resolution and quantitation, make capillary electrophoresis a powerful complement to slab gel electrophoresis and may make it a preferred alternative to both agarose gel electrophoresis and HPLC for applications such as the confirmation of plasmid integrity.

DNA↗

PubMed2003

Cowpea bruchid Callosobruchus maculatus uses a three-component strategy to overcome a plant defensive cysteine protease inhibitor.

The soybean cysteine protease inhibitor, soyacystatin N (scN), negatively impacts growth and development of the cowpea bruchid, Callosobruchus maculatus[Koiwa et al. (1998) Plant J 14: 371-379]. However, the developmental delay and feeding inhibition caused by dietary scN occurred only during the early developmental stages (the 1st, 2nd and 3rd instars) of the cowpea bruchid. The 4th instar larvae reared on scN diet (adapted) exhibited rates of feeding and development which were comparable to those feeding on an scN-free diet (unadapted) prior to pupation. Total gut proteolytic capacity at this larval stage significantly increased in the scN-adapted insects. The elevated enzymatic activity was attributed to a differential expression of insect gut cysteine proteases (representing the major digestive enzymes), and of aspartic proteases. scN degradation by the gut extract was observed only in adapted bruchids, and this activity appeared to be a combined effect of scN-induced cysteine and aspartic proteases. Thirty cDNAs encoding cathepsin L-like cysteine proteases were isolated from insect guts, and they were differentially regulated by dietary scN. Our results suggest that the cowpea bruchid adapts to the challenge of scN by qualitative and quantitative remodelling of its digestive protease complement, and by activating scN-degrading protease activity.

Amino Acid Sequence↗

PubMed2000

Unnecessary water supplementation for babies: grandmothers blame clinics.

UNLABELLED: This study was designed to obtain policy-and programme-relevant data on exclusive breastfeeding in Lesotho at a time when global recommendations for exclusive breastfeeding were new. During both phases of field research, conducted in 1991 and 1992, qualitative information was obtained through focus groups and individual interviews with mothers, grandmothers and nurses. This was complemented during the second phase with quantitative data collected through a clinic-based survey of mothers. The qualitative and the quantitative findings consistently converged, illustrating a culture of infant feeding in which breastfeeding was central, but exclusive breastfeeding was an unknown concept and not practised. Grandmothers seemed to be more in tune with the ideal of exclusive breastfeeding as they had given their young infants thin gruel only occasionally. Contemporary mothers, in contrast, were regularly giving their young infants water. Mothers and grandmothers frequently cited nurses as the source of advice for giving water. Grandmothers were adamant in pointing out that they had never given water to their own young infants and asserted that they avoided giving it to their grandchildren as they considered it unnecessary and harmful. According to the grandmothers, water supplementation was a new practice that had been introduced through the clinics. CONCLUSION: Efforts to discourage water supplementation and encourage exclusive breastfeeding in this setting need to be directed both at mothers and health providers.

Adult↗

PubMed2004

Qualitative research: contributions to the study of drug use, drug abuse, and drug use(r)-related interventions.

This article describes how qualitative social science research has and can contribute to the emerging field of drug and alcohol studies. An eight-stage model of formative-reformative research is presented as a heuristic to outline the different ways in which qualitative research may be used to better understand micro and macro dimensions of drug use and distribution; more effectively design, monitor and evaluate drug use(r)-related interventions; and address the politics of drug/drug program representation. Tobacco is used as an exemplar to introduce the reader to the range of research issues that a qualitative researcher may focus upon during the initial stage of formative research. Ethnographic research on alcohol use among Native Americans is highlighted to illustrate the importance of closely examining ethnicity as well as class when investigating patterns of drug use. To familiarize the reader with qualitative research, we describe the range of methods commonly employed and the ways in which qualitative research may complement as well as contribute to quantitative research. In describing the later stages of the formative-reformative process, we consider both the use of qualitative research in the evaluation and critical assessment of drug use(r)-intervention programs, and the role of qualitative research in critically assessing the politics of prevention programs. Finally, we discuss the challenges faced by qualitative researchers when engaging in transdisciplinary research.

Anthropology, Cultural↗