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Phlebotomists at the Jackson-Madison County General Hospital perform more than 18,000 venipunctures a month to collect blood specimens for laboratory tests. Because each of the blood draws involves handling five or six labels for the tubes that the blood is collected in, it means the hospital prints about 90,000 labels a month, which makesfor a lot of label sorting.
We monitored the development of hepatocellular carcinoma due to chronic infection with woodchuck hepatitis virus by using monthly serum samples, physical examination, and magnetic resonance imaging. The same woodchucks can be imaged repeatedly over a 1-y period by allowing the animals to recover after each experiment, thus reducing the number of animals required without compromising the quality of the data obtained. Age- and sex-matched uninfected control (n = 5) and chronically infected (n = 5) woodchucks were group-housed according to sex and infection status. Woodchucks were anaesthetized using an inhalation anesthetic (isoflurane) without premedication. During imaging, we regularly monitored heart rate, body temperature, and respiration. Tumor growth was observed using MRI, whereas the extent of hepatocyte injury was followed using serum liver enzymes. Elevated serum gamma glutamyltransferase and aspartate aminotransferase levels indicated hepatocyte injury due to tumor growth. On magnetic resonance images, the liver should appear as a well-defined, homogenous organ with defined regions of hyperintensities from larger blood vessels. Within tumor nodules, the liver appeared irregularly shaped, having heterogeneous intensity from unregulated cellular proliferation. Changes in tumor size can be monitored by imaging infected woodchucks on a regular basis. Using the imaging techniques we describe, the development of hepatocellular carcinoma can be visualized using magnetic resonance imaging, correlated to serum tests, and compared with the results from uninfected control woodchucks, thereby improving the understanding of the disease progress.
We examined a number of large proficiency and control databases supporting the values reported for pesticide residues in agricultural commodities at fractions of a part per million (mg/kg). The average recovery from >100,000 recovery records in 13 databases was 94%. The overall average single-value relative standard deviation (RSD) of the reported recoveries was 17% at a mean concentration (C) of about 10(-7) (0.1 mg/kg). The average apparent HORRAT value (RSD found/RSDR predicted from the Horwitz formula [2*C(-0.1505)]) was 0.8. Analysis of variance indicated that about 60-70% of the variance could not be associated with any particular factor or combination of factors-analyte, commodity, method, laboratory, concentration, database, or their interactions. The most predominant factor, analyte, and its third-order interaction with laboratory and concentration contributed most of the assignable variance. These findings suggested that most of the variability of trace analysis for pesticide residues is "random" in the sense of being inherent and not assignable to specific factor fluctuations.
The antiphospholipid syndrome is a disorder characterized by recurrent thrombosis and the presence of antibodies specific to phospholipids. However, the diagnosis of this syndrome is hampered by the lack of a specific laboratory test. In this study an ELISA for the measurement of antibodies to solid-phase beta2-glycoprotein I (beta2-GPI) was established and compared with anticardiolipin antibodies for diagnosis of antiphospholipid syndrome. Significantly elevated levels of antibodies to beta2-GPI were found in all patients with definite antiphospholipid syndrome (median = 91 AU). Marginally elevated levels of antibodies to beta2-GPI were observed in 5% of patients with systemic lupus erythematosus (SLE; median = 4 AU), 1% with stroke (median = 3 AU), 13% with infectious mononucleosis (median = 3 AU), 10% with HIV infection (median = 3 AU) and 8% with VDRL false-positive serology for syphilis (median = 4 AU), but not in patients with rheumatoid factor, syphilis or carotid artery stenosis. In contrast, significantly raised levels of anticardiolipin antibodies were observed in 100% of patients with definite antiphospholipid syndrome, 30% with SLE, 88% with HIV infection, 94% with syphilis, 62% with infectious mononucleosis, 9% with rheumatoid factor-positive sera, 74% VDRL false-positive serology for syphilis, 47% with stroke and 0% with carotid artery stenosis. This solid-phase assay for antibodies to beta2-GPI is highly specific for the antiphospholipid syndrome and represents an advance in the laboratory diagnosis of this disorder.
A combination guaiac and immunological test Fecatwin/Feca EIA for the detection of faecal occult blood was used in the investigation of 535 symptomatic patients in order to assess the value of this combined test in the detection of colorectal neoplasia. The addition of the immunological part of the test led to an improvement in specificity for colorectal cancer from 80% to 91% and an increase in the positive predictive value of the test for colorectal cancer from 14% to 24%. However, there was a fall in sensitivity from 75% to 67%. There was no substantial advantage in the addition of the immunological part of the test to justify the extra laboratory workload incurred.
The evaluation of myocardial metabolic changes in ischemic heart disease remains centered on the coronary sinus pacing and sampling techniques established over the last 25 years. Lactate remains the marker of choice for most institutions, though centers with more sophisticated laboratories will always be trying to improve on the sensitivity and specificity for ischemia, perhaps using ATP catabolites and nucleotides and measuring coronary sinus flow. The diagnostic value of lactate changes is limited and probably not superior to a well-conducted 12-lead treadmill exercise electrocardiogram test, but it does provide an objective marker for reliable further study and evaluation of interventions. It is almost certainly in the research context that metabolic studies have their place--evaluating drugs, surgery, or angioplasty and perhaps shedding light on obscure entities, such as chest pain with normal coronary arteries and cardiomyopathies. Attention to detail and simplicity of study are more likely to lead to valuable results rather than concentrating on the complexities. Each laboratory should establish the reproducibility of its results before commencing any procedures.
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Contamination in a trace element laboratory can come from a variety of sources, including laboratory gloves. Therefore, vinyl and latex gloves were obtained from as many manufacturers as would supply gloves. These gloves were either prepared for acid-washing and subsequent soaking in an acid solution, or immersed in an acid solution for a duration of either 1 min or 1 h. Incubation washes were analyzed for a variety of trace elements by flame atomic absorption spectroscopy (AAS) or inductively coupled mass spectrometry (ICP-MS). Results indicated that only three brands of vinyl gloves were acceptable for use in a trace element laboratory, whereas others had contamination of different elements. Latex gloves contained such high levels of biologically important elements that they were not considered suitable for routine trace element work. Vinyl gloves of choice should be routinely acid-washed before use in a trace element laboratory.
The effectiveness and safety of a protocol for transcatheter patent ductus arteriosus (PDA) closure was assessed. Our goal is complete mechanical occlusion of the PDA in the catheterization laboratory by adding coils until it is no longer possible to cross the PDA with a guidewire. Detachable coil closure of a PDA with a narrowest diameter of 2.4 +/- 0.1 mm was attempted in 83 patients with a median age of 2.8 years (0.7 to 27.8 years) and whose median weight was 14.5 kg (6 to 61.6 kg). Coils were successfully implanted in 82 of 83 patients, and in 1 patient a large Rashkind double umbrella was used instead. Complete closure was obtained in 80 (97.6%) patients, 48 of those (59%) received more than one coil. Reintervention for residual shunting was required in only 1 patient and another patient has a trivial residual shunt. Device embolization occurred in three cases. Despite the use of multiple coils there was no evidence of significant left pulmonary artery stenosis. The fluoroscopy time increased from 14.0 +/- 2.0 minutes for a single coil to 25.3 +/- 2.9 minutes for multiple coils (p < 0.01). Attempting to obtain complete mechanical occlusion of the PDA during the implant procedure by adding extra coils reduces the need for reintervention for residual or recurrent shunting.
L-929 cells (mouse fibroblasts) permeabilized with dextran sulfate (DSP cells) carry out vigorous and linear rates of glycolysis when supplied with a suitable incubation medium. Glycolysis in DSP cells is pH dependent, being strongly inhibited at pH 6.5. Compared to their nonpermeabilized counterparts, DSP cells exhibit faster glycolytic rates, but tend to convert a smaller proportion of the glucose utilized to lactate. [14C]Glucose is converted to lactate by DSP cells without dilution from endogenous substrates. When exogenous 12C-labeled glycolytic intermediates (12C-I) are added to glycolyzing DSP cells the [14C]lactate produced from [14C]glucose is diluted to varying extents, depending on the intermediate. However, the extent of that dilution (reduced specific activity) is not that expected from the complete mixing of exogenous 12C-I with their corresponding 14C-labeled intermediates coming from [14C]-glucose. DSP cells also respire and convert glucose to CO2. The amount of 14CO2 produced from [14C]glucose is also reduced by addition of most 12C-I, an interesting exception being pyruvate, which had no measurable effect on 14CO2 production and caused only a modest stimulation of respiration in glycolyzing DSP cells. These results suggest that channeling, or some other form of coupling, takes place between the glycolytic production of pyruvate and its further oxidation. These observations confirm previously published data and add further support to the proposition that channeling of glycolytic intermediates occurs in DSP cells but is of the "leaky" type. Although abundant evidence in the literature indicates that various glycolytic enzymes associate with F-actin, as well as other elements of the cytomatrix, we observed no effect of cytochalasin D on lactate production even at very high concentrations of this compound. Our results are compared with those from other laboratories and discussed in the context of metabolic organization.
A general procedure is proposed for the rapid development of a reversed-phase liquid chromatographic (RP-LC) separation that is "orthogonal" to a pre-existing ("primary") method for the RP-LC separation of a given sample. The procedure involves a change of the mobile-phase organic solvent (B-solvent), the replacement of the primary column by one of very different selectivity, and (only if necessary) a change in mobile phase pH or the use of a third column. Following the selection of the "orthogonal" B-solvent, column and mobile phase pH, further optimization of peak spacing and resolution can be achieved by varying separation temperature and either isocratic %B or gradient time. The relative "orthogonality" of the primary and "orthogonal" RP-LC methods is then evaluated from plots of retention for one method versus the other. The present procedure was used to develop "orthogonal" methods for nine routine RP-LC methods from six pharmaceutical analysis laboratories. The relative success of this approach can be judged from the results reported here.
Ability to detect pigmented and nonpigmented Pasteurella pestis is essential in plague research, and is currently dependent on use of the synthetic hemin-agar of Jackson and Burrows. We have devised a new differential medium for this purpose, containing Congo red dye and common, commercially available laboratory media. The ease and simplicity of preparation make the Congo red-agar a practical routine laboratory tool in plague research. These findings, possibly indicating a common binding site for hematin and Congo red, should be useful in efforts to determine the chemical nature of a bacterial component associated with high virulence in P. pestis.
To evaluate effects of anaesthesia delivery systems on respiratory work in infants, we measured end-tidal PCO2 (PETCO2), transcutaneous PCO2 (PTcCO2) respiratory rate and arterial blood gases in infants (ages 0.2 to 23.8 months) anaesthetised with either a Jackson-Rees (n = 11) or paediatric circle absorber system (n = 15). Two hours after induction, with assisted ventilation there was no difference in PETCO2, PTcCO2, respiratory rate, pH or PaCO2 in infants anaesthetised with either system. In the laboratory, we measured inspiratory and expiratory resistances to breathing through each system at various tidal volumes and total gas flows into the circuit. These values were compared to resistances created by breathing through an appropriately sized endotracheal tube. Although there was lower resistance to ventilation through Jackson-Rees system, both systems had lower resistances for almost all flows tested than breathing through an endotracheal tube alone (p less than 0.05). The paediatric circle absorber system is an efficient apparatus for anaesthetising spontaneously breathing infants.
To assess whether deficiencies exist in the processing of contaminated flexible fiberoptic sigmoidoscopes in family practice and internal medicine offices and whether training of office personnel results in a correction of identified deficiencies, we conducted a prospective review of sigmoidoscope processing in family practice and internal medicine offices before and after a training course. Participants were questioned on their current endoscope processing for 17 standards before and 2 months after receiving training. The 19 offices had between 4 and 11 deficiencies per office before training, with an average of 6.8 deficiencies per office. After training, deficiencies ranged from 0 to 8, with an average of 0.9 deficiencies per office (P < or = .001; Student t test). Personnel responsible for processing flexible sigmoidoscopes in family practice and internal medicine offices are insufficiently trained for this function. Endoscopes are not being processed according to current standards. After a 2-hour training period, these persons maintain their equipment close to or according to standards.
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Sixty-four IgG Rh monoclonal antibodies (Mabs) submitted to the Fourth International Workshop on Monoclonal Antibodies Against Human Red Blood Cells and Related Antigens were characterised and tested in quantitative functional assays at five laboratories. The biological assays measured the ability of anti-D to mediate phagocytosis or extracellular lysis of RBC by IgG Fc receptor (Fc gamma R)-bearing effector cells. Interactions of RBC pre-sensitised with anti-D (EA-IgG) with monocytes in chemiluminescence (CL) assays were found proportional to the amount of IgG anti-D on the RBC. Using antibodies to inhibit Fc gamma RI, Fc gamma RII or Fc gamma RIII, the only receptor utilised in the monocyte CL and ADCC assays for interactions with EA-IgG1 was found to be Fc gamma RI. In these assays, enhanced interactions were promoted by EA-IgG3 and additional Fc gamma receptors may have contributed. IgG2 anti-D was not reactive in these assays and EA-IgG4 promoted weak reactions through Fc gamma RI. A macrophage ADCC assay showed that haemolysis of EA-IgG3 was greater than that of EA-IgG1, mediated mainly through Fc gamma RIII. In ADCC assays using lymphocytes (NK cells) as effector cells and papainised RBC target cells, only a minority of IgG1 anti-D Mabs were shown to be able to mediate haemolysis in the presence of monomeric IgG (AB serum or IVIg). These interactions were mediated solely through Fc gamma RIII. Haemolysis via Fc gamma RIII may depend on the presence of certain sugars on the oligosaccharide moiety of IgG. Most Mabs (IgG1, IgG2, IgG3 and IgG4) elicited intermediate, low or no haemolysis in these assays. Blocking studies indicated that low activity IgG1 and IgG4 anti-D utilised only Fc gamma RI. Other IgG1 and IgG3 Mabs appeared to promote haemolysis through Fc gamma RI and Fc gamma RIII while IgG2 was inhibited by Mabs to both Fc gamma RII and Fc gamma RIII, suggesting a variety of Fc gamma R are utilised for anti-D of low haemolytic activity. Excellent agreement between the results of the lymphocyte ADCC assays and antibody quantitation was observed between the participating laboratories.
Science education in the United States at all academic levels is widely perceived to need direct assistance from professional scientists. The current dearth of quality applicants from this country to medical and graduate schools suggests that our existing undergraduate and high school science curriculum is failing to provide the necessary stimulus for gifted students to seek careers in the health sciences. Recognizing the need to become more directly helpful to high school and college science teachers, members of the faculty of the Department of Physiology and Biophysics at the University of Kentucky College of Medicine held a 5.5-day Physiology Summer Workshop during June, 1989. Participants included 25 college teachers from Kentucky and 5 other states plus 22 Kentucky high school teachers. The presence of the two levels of educators provided communication about curricular concerns that would be best addressed by mutual action and/or interaction. Each day's activities included morning lectures on selected aspects of organ system and cellular physiology, a series on integrative physiology, and afternoon laboratory sessions. The laboratory setting allowed the instructor to expand on principles covered in lecture as well as provided the opportunity for in-depth discussion. A selection of evening sessions was presented on 1) grants available for research projects, 2) obtaining funds for laboratory equipment, and 3) graduate education in physiology.