The effect on researchers of handling human fetal tissue.
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Aborted Fetus↗
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Investigation of the effects of solution composition and container material type on the loss of 11-nor-delta 9-THC-9-carboxylic acid.
The loss of 11-nor-delta 9-tetrahydrocannabinol-9-carboxylic acid (THC-COOH) from solution was studied using fluorescence polarization immunoassay (FPIA) technology and x-ray photoelectron spectroscopy (XPS). Several materials (glass, silylated glass, high density polyethylene, polypropylene, polystyrene, polymethylmethacrylate, Teflon, and Kynar) were studied along with three solvents (water, urine, and Abbott cannabinoids diluent). THC-COOH losses ranging from 0 to 9.7 ng/cm2 and concentration reductions to 46% of starting values were measured. XPS indicated the presence of fluorine-labeled THC-COOH at materials surfaces. A half-life of 10 min was calculated for THC-COOH loss from urine stored in high density polyethylene at room temperature. Sample handling losses during pipetting were determined and ranged from 1.1 to 7.9 ng per aliquot. The effects of sample volume and sample handling on the THC-COOH concentrations of controls were also investigated.
Dronabinol↗
[Sample distribution of inner ear fluids in the submicroliter range].
Biochemical analysis of inner ear fluids is fraught with numerous problems. After developing accurate techniques to measure inorganic and organic substances in fluid amounts of 0.1 microliter the problem of measuring and handling series of samples arose. It became necessary to develop methods to handle inner ear fluid taken without sophisticated techniques, i.e. to measure, to store and to dispatch inner ear fluid samples. The fluid volume is measured with a "nanocap" technique and is then transferred to cellulose acetate membranes. A 3 mm long glas capillary tube held by a special device is dipped into the inner ear fluid sample. The capillary tube is then placed on a membrane piece that adheres to the tube. The higher capillary forces of the membrane transfer the fluid out of the glass tube into the membrane which is put into a plastic reaction tube and stored. The reproducibility of the method was tested by flame photometric assays of standard sodium solutions and of perilymph from the scala tympani of guinea pigs. The variation coefficient is below 3% for a standard solution and below 5% for perilymph. The method developed to measure and handle inner ear fluids with the aid of "nanocap" and membrane pieces is easier and faster than the conventional techniques.
Animals↗
[Comparison of 4 processing methods for the diagnosis of peritonitis in patients undergoing chronic ambulatory peritoneal dialysis].
BACKGROUND: Four methods for the diagnosis of peritonitis in patients submitted to chronic ambulatory peritoneal dialysis were evaluated. METHODS: Fluid from eighty-five patients undergoing peritoneal dialysis processed by the following techniques conventional, centrifugation, injection into conventional BACTEC bottles and in double volume BACTEC bottles. In addition, qualitative and quantitative counts of the number of leukocytes/mm3 of peritoneal dialysis, as well as the gram staining of the sediment obtained following centrifugation were carried out. RESULTS: Sixty-eight percent of the peritoneal dialysis fluid processed fulfilled criteria of peritonitis. Global sensitivity of the diagnostic techniques used was 62%. The sensitivities of the different methods were individually: conventional, 46%, centrifugation, 52%, injection into conventional BACTEC bottles, 53%, and injection into double volume BACTEC bottles 58%. Gram staining sensitivity was 9%. CONCLUSIONS: Direct injection of peritoneal dialysis in double volume BACTEC bottles is a useful technique for the etiologic diagnosis of peritonitis in patients submitted to chronic ambulatory peritoneal dialysis. In this study, its sensitivity was greater to that of the other methods evaluated allowing minimal handling and delay in the processing of the sample.
Ascitic Fluid↗
Accurate high-speed liquid handling of very small biological samples.
Molecular biology techniques require the accurate pipetting of buffers and solutions with volumes in the microliter range. Traditionally, hand-held pipetting devices are used to fulfill these requirements, but many laboratories have also introduced robotic workstations for the handling of liquids. Piston-operated pumps are commonly used in manually as well as automatically operated pipettors. These devices cannot meet the demands for extremely accurate pipetting of very small volumes at the high speed that would be necessary for certain applications (e.g., in sequencing projects with high throughput). In this paper we describe a technique for the accurate microdispensation of biochemically relevant solutions and suspensions with the aid of a piezoelectric transducer. It is suitable for liquids of a viscosity between 0.5 and 500 milliPascals. The obtainable drop sizes range from 5 picoliters to a few nanoliters with up to 10,000 drops per second. Liquids can be dispensed in single or accumulated drops to handle a wide volume range. The system proved to be excellently suitable for the handling of biological samples. It did not show any detectable negative impact on the biological function of dissolved or suspended molecules or particles.
Base Sequence↗
Detection and measurement of sulfur mustard offgassing from the weanling pig following exposure to saturated sulfur mustard vapor.
Animal models are employed to investigate mechanisms of injury and to evaluate protective measures against sulfur mustard (HD) exposure. The ability to detect and quantify HD enables the researcher to follow safe procedures in handling skin samples. We designed an experimental procedure to measure HD offgassing from animal models. A Minicams--a portable gas chromatograph equipped with a flame photometric detector and on-line sorbent collection and desorption--was used to monitor the HD concentration. Confirming measurements were made using a two-step process that trapped HD on a Tenax sorbent off-line and then transferred the sample by means of an ACEM 900 to a gas chromatograph equipped with either a flame photometric detector or a mass spectrometer. Sulfur mustard offgassing data are presented from three experiments in which weanling pigs were exposed to saturated HD vapor via vapor caps containing 10 microl of HD. The HD concentration was measured in time-weighted-average (TWA) units at a specific HD application site. The current 8-h maximum exposure limit for HD is 3-ng l(-1), (1 TWA unit). The largest TWA value measured near a 3 h time point was a Minicams measurement of 0.48 TWA at 2 h and 53 min after removal of a vapor cap containing HD from a single exposure site on an animal that had 24 concurrent dorsal exposure sites. Gas chromatography/flame photometric detection and gas chromatography/mass spectrometry were used to confirm the Minicams data and to provide greater sensitivity and selectivity down to 0.1 TWA. The gas chromatography/mass spectrometry data confirmed that HD concentrations fell below 0.1 TWA in <5 h for a specific site. These measurements of HD concentrations provided information on the expeditious and safe handling of HD-exposed tissue.
Animals↗
A validation of extraction methods for noninvasive sampling of glucocorticoids in free-living ground squirrels.
Fecal hormone assays provide a powerful tool for noninvasive monitoring of endocrine status in wild animals. In this study we validated a protocol for extracting and measuring glucocorticoids in free-living and captive Belding's ground squirrels (Spermophilus beldingi). We first compared two commonly used extraction protocols to determine which performed better with commercially available antibodies. We next verified the preferred extraction method by correlating circulating and fecal glucocorticoid measures from a group of individuals over time. For this comparison, we used both a cortisol and a corticosterone antibody to determine which had greater affinity to the fecal metabolites. Cortisol was the primary circulating glucocorticoid, but both hormones were present in well above detectable concentrations in the blood, which does not occur in other sciurids. In addition, the cortisol antibody showed greater binding with the fecal extracts than did the corticosterone antibody. Finally, we used adrenocorticotropic hormone and dexamethasone challenges to demonstrate that changes in adrenal functioning are reflected in changing fecal corticoid levels. These results suggest that our extraction protocol provides a fast, reliable assay of stress hormones in free-living ground squirrels without the confounding influence of short-term rises in glucocorticoid concentrations caused by handling and restraint stress and that it can facilitate ecological and evolutionary studies of stress in wild species.
Adrenocorticotropic Hormone↗
Radiation safety precautions for sentinel lymph node procedures.
Radiation safety professionals need guidelines for controlling occupational radiation doses and handling generated contaminated tissues from sentinel lymph node procedures involving 99mTc. Many healthcare facilities do not have formal controls in place, or they adopt policies that may be too restrictive or impractical. During accreditation inspections at healthcare facilities, agencies such as the College of American Pathology and the Joint Commission on the Accreditation of Healthcare Organizations occasionally raise questions about these issues. Inspectors expect to find formal policies to ensure these procedures are performed safely. The objective of this article is to review the basic radiation safety controls needed during sentinel lymph node procedures and to propose guidelines that may be used to control occupational radiation doses and handling of SLN contaminated tissues.
Decontamination↗
An efficient staining method for ultrathin sections.
A staining method to handle simultaneously as many as 20 electron microscope grids is described. The devices used are easily constructed of readily obtained inexpensive materials. The volumes of stain and wash water required are very small and drying grids is simplified.
Specimen Handling↗
Analytical considerations for free protein S assays in protein S deficiency.
Protein S is an anticoagulant protein that circulates in plasma in complex with C4b-binding protein (C4BP) or in free form. Deficiency of protein S increases the risk of venous thrombosis. Measurement of free protein S, as compared to total levels, has been shown to be superior for prediction of protein S deficiency. We studied the effects of different handling protocols for an immuno- and a ligand (C4BP)-based assay for free protein S. When the assay was performed at 37 degrees C, the levels of free protein S in plasma from protein S deficient patients were approximately twice those obtained at room temperature. The reason for this phenomenon was that plasmas from protein S deficient patients exhibited a time-, temperature-, and dilution-dependent increase in free protein S, which was more pronounced than corresponding dilution of the normal plasma that was used to create the standard curve. These findings demonstrate the importance of assay procedure and sample handling in assays for free protein S.
Aged↗
Multiple or sequential oestrogen receptor assays in breast cancer.
Multiple oestrogen receptor assays have been performed on primary, locally recurrent, and metastatic breast cancer in 62 patients. A change in receptor status was found in seven of 25 late recurrences (28%). It was noted that the level of oestrogen receptor was higher in lymph nodes than the primary tumour, particularly at the time of initial surgery. Other factors which might contribute to differences in receptor levels include site of metastases, method of handling, heterogeneity of tumours, and subsequent treatment. Such differences have important implications in management.
Antineoplastic Agents↗
Influence of handling procedures on rat plasma creatine kinase activity.
Ten-week-old Sprague-Dawley rats were held behind the front legs before and during anesthetic injection of sodium pentobarbital (group 1, 12 rats), or lifted by the base of the tail before and during injection (group 2, 12 rats). Creatine kinase (CK) values were higher (P = 0.004) in group 1 (median 564 IU/L) than in group 2 (median 272 IU/L). Handling rats by holding them behind the front legs may reduce the usefulness of CK activity as a measure of muscular disorders.
Alanine Transaminase↗
Impact of various handling and storage conditions on quantitative detection of hepatitis C virus RNA.
BACKGROUND/AIMS: Both HCV RNA viral load and HCV genotype have been described as important predicting factors determining the response to interferon in chronic hepatitis C. To investigate whether processing and storage conditions might influence the stability and could alter the concentration of the HCV RNA in serum, quantification of HCV RNA was performed by branched DNA assay. METHODS: We studied serum samples obtained from seven patients with histologically proven chronic hepatitis C. These were subjected to the following physical conditions: (1) immediate quantification, (2) storage at room temperature for 5 days, (3) storage at 4 degrees C for 5 days, (4) storage at -20 degrees C for 5 days, (5) storage at -80 degrees C for 5 days, (6) five freeze-thaw cycles, (7) blood unspun for 4 h at room temperature then centrifuged and stored at -80 degrees C for 5 days, (8) storage at 4 degrees C for 6 months, (9) storage at -20 degrees C for 6 months, (10) storage at -80 degrees C for 6 months. RESULTS: A loss of 100% HCV RNA titers was observed after storage at RT for 5 days and then storage at 4 degrees C for 6 months. A surprising decrease of HCV RNA titer (15.6%) was observed in sera stored for 5 days at -20 degrees C. Five freeze-thaw cycles resulted in a 16% decrease of the HCV RNA level. When centrifugation was performed after a 4 h delay at room temperature, a significant loss of HCV RNA titers of 29.5% was observed. Long-term stability (6 months) was observed at -80 degrees C with a slight loss of about of 10% HCV RNA titers, but a significant decrease in HCV RNA of 23% was observed at -20 degrees C. The reproducibility of the bDNA assay on five patient samples was performed eight times in duplicate and showed an average coefficient of variation of 9.1%. CONCLUSIONS: These data confirm the importance of storage and handling in measuring the amount of HCV RNA in clinical samples.
Blood Preservation↗
A method for obtaining repeatable measurements of the tensile properties of skin at low strain.
Initial attempts at measuring the low strain tensile properties of skin led to results which were not repeatable. It appeared that strains induced in the sample due to the handling needed to mount it in a simple clamp led to the low precision of the test. A new method which minimized the manipulation needed to mount a sample in a testing machine was then developed. Using this method the skin was aligned with a set of grips in a saline bath. When aligned, the saline was removed and the sample clamped in the grips without the need for additional handling. This approach produced a marked improvement in the repeatability of the tensile properties of skin at low strains.
Animals↗
Ensuring quality in long term environmental monitoring for chemical speciation.
The awareness regarding quality assurance (QA) and quality control (QC) for environmental monitoring has considerably increased in the past few years, especially with respect to the determination of chemical species, since errors occurring at various levels may considerably affect the interpretation of results (e.g. studies of toxic impact, geochemical pathways, etc.). QA for environmental analysis covers a broad range of activities from sample collection to laboratory work and the approach for ensuring quality data should be considered in a global context. However, whereas great emphasis has been placed on QA within the laboratory, there have been few systematic attempts to evaluate risks of discrepancies related, for example, to field manipulations. The situation is even worse when monitoring is applied to relatively new fields such as chemical speciation. This paper outlines some of the main aspects of quality control of environmental analysis, including the validation of methods, sampling and sample handling, storage, etc., with special emphasis on the monitoring of chemical forms of elements (e.g. species of tin, mercury, lead and selenium).
Chemistry Techniques, Analytical↗
A container for processing small volumes of cell suspensions for critical point drying.
The attachment of lymphocytes to glass or filters, in order to facilitate handling for processing prior to scanning electron microscopy, may introduce artefacts in surface topography. A container has therefore been adapted, from an embedding capsule, for the preparation of small volumes of cell suspensions.
Cytological Techniques↗
Method of cell handling affects leakiness of cell surface labeling and detection of intracellular keratins.
Keratins are a subgroup of cytoskeletal intermediate filament proteins found in most epithelial cells. Some reports have suggested that keratins may be found on the cell surface as well as their well-accepted cytoskeletal location. A major part of the evidence in the interpretation of cell surface expression of keratins is cell surface radioiodination. Here we show that lactoperoxidase-catalyzed iodination of colonic and breast tissue culture cells results in radiolabeling of the keratins when cells are manipulated. No labeling of keratins is detected when cells are labeled directly on the tissue culture dish. A similar result was obtained when intact cells were biotinylated using water-soluble sulfo-NHS-biotin. Partitioning of the keratins to a soluble and an insoluble pool after "cell surface" 125I-labeling showed that both pools became iodinated. Indirect immunofluorescence showed that binding of a panel of anti-keratin antibodies to intact epithelial cells occurs only on the cells that are more adherent, which are the cells that require longer manipulation to remove from the tissue culture dish. Taken together, our results indicate that the reported expression of cell surface keratins in some cells likely reflects intracellular keratins. In addition, the method of epithelial cell handling can dramatically alter the leakiness of cell surface iodination techniques.
Adenocarcinoma↗
[Clinicopathological discordance for lesions of the oral mucosa].
BACKGROUND: The oral cavity is the site of various oral mucosal lesions. The physical examination must be completed by a pathology examination in order to establish a final diagnosis. The objective of our study was to investigate the discordance between clinical and pathological findings concerning lesions oral mucosa. MATERIAL AND METHOD: This study was a cross-sectional epidemiological investigation involving 21 patients seen during a 16-month period. RESULTS: Statistical analysis demonstrated 17 cases of agreement (81%) and 4 cases of discordance (19%) between the clinical diagnosis and pathology diagnosis. DISCUSSION: Such discordance can be operator-dependent or related to sample procurement or handling.
Adolescent↗