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PubMed2002

Histochemical studies of the extracellular matrix of human articular cartilage--a review.

OBJECTIVE: This paper reviews the histochemistry of the extracellular matrix of human articular cartilage. No systematic review of histochemical knowledge and techniques in the study of articular cartilage has been published previously. METHODS AND RESULTS: Literature was searched in the Winspirs Medline database from 1960 to 2000. Only techniques applicable for bright field or polarization microscopy were considered. Unless otherwise noted, all applies to hyaline cartilage. The most widely used fixatives are adequate for routine staining of proteins, but proteoglycan fixation is problematic, and no one fixative can be recommended. Proteoglycan can be stained reliably but it is problematic that, at low substrate concentrations, these methods are not stoichiometric. Collagen can be stained efficiently, although attempts to differentiate collagen types have not been successful. CONCLUSIONS: Detailed studies of fixation and staining procedures should be carried out and standards for cartilage sampling, handling and evaluation agreed upon if results from different laboratories are to be compared.

Cartilage, Articular↗
PubMedDate not supplied

Effect of fluctuations in temperature encountered during handling and shipment of human cryopreserved semen.

The effect of temperature fluctuations which cryopreserved spermatozoa may undergo during routine shipping and handling was evaluated in sperm frozen with two cryoprotectants. Sperm frozen in TEST-yolk buffer maintained motility better than those frozen in glycerol solution in all studies. Sperm motility was significantly compromised in samples stored more than one day in dry ice, regardless of the cryoprotectant, and more than two days in a liquid nitrogen shipping dewar if frozen in glycerol solution. Sperm motility was not compromised following exposure to room temperature for up to 3 min if TEST-yolk buffer was used as the cryoprotectant, but was compromised following 1 min exposure using glycerol cryoprotectant. This study describes the limits of non-ideal conditions that spermatozoa may undergo during shipping and handling, and demonstrates the effects of the cryoprotectant on those limits.

Cryopreservation↗
PubMed1982

Maintenance of sterility in urinary drainage bags.

When the outlet tube of urinary drainage sets was kept full of 3 per cent hydrogen peroxide at all times, no evidence of bacterial growth in the urine of the collection bag was noted in at least 92.0 per cent of the patients and 93.3 per cent of the drainage periods in a group of 60 patients and 75 drainage periods. One patient had a positive urinary drainage bag culture and a positive bladder culture as a result of improper disconnection of the catheter, and four patients had bacterial growth from urinary drainage bag samples, two of which were possibly due to a faulty culture technique and one of which may have resulted from contamination of amphoteracin B irrigating solution. Hydrogen peroxide, 3 per cent, appears to approach 100 per cent effectiveness as an antibacterial barrier. Only one patient of 60 or 1.7 per cent had a urinary tract infection develop which was related to the improper handling of the drainage set.

Anti-Infective Agents↗
PubMed1999

Comparison of enteric neuronal morphology as demonstrated by Dil-tracing under different tissue-handling conditions.

The aim of this study was to determine locations and morphologies of enteric neurons innervating the small intestinal mucosa of the pig after application of the carbocyanine tracer Dil onto a single villus. The tissue was processed in two ways: incubation (1) of fixed material (postmortem tracing) for several months and (2) of living specimens within organotypic culture in vitro for several days (supravital tracing). In both procedures Dil-labelled neurons were found in the three ganglionated plexuses, the internal and external submucous plexus as well as the myenteric plexus. Postmortem tracing revealed different neuronal morphologies. Adendritic type II neurons were present in all three plexuses, type IV neurons with short, scarcely branched, polarly emerging dendrites were mainly found in the myenteric plexus and small dendritic neurons were mainly present in the internal submucous plexus. The latter may correspond to minineurons hitherto described only immunohistochemically. Tracing within tissue culture showed somata of neurons and, partly, proximal segments of processes to be labelled. Subsequent immunohistochemistry using general neuronal markers revealed some neurons to be adendritic type II neurons. Visualization of dendrites was less clear, hampering an accurate morphological classification of dendritic neurons. Our results suggest that neurons of all ganglionated enteric nerve plexuses of the pig participate in the innervation of the mucosa, and that postmortem tracing revealed enteric neuronal morphology more clearly than supravital tracing. Since the former method cannot be applied for deciphering the chemical coding of enteric neurons, combination of both methods will extend our knowledge of the morphological substrate for the intrinsic neuronal microcircuits in the gastrointestinal tract.

Animals↗
PubMed1986

Beta-nerve growth factor measurements in mouse serum.

Mouse serum beta-nerve growth factor (NGF) levels were measured using a newly developed competitive beta-NGF radioimmunoassay. The basal serum beta-NGF levels in male and female mice were consistently less than 2 ng/ml when these animals were maintained in individual cages for at least 7 days before they were killed. However, in male mice, serum beta-NGF levels were significantly elevated when they were housed 5 per cage. The rise in serum beta-NGF levels, presumably mediated by intermale aggression, was confirmed by grouping previously isolated mice together in one cage for 20 min before they were killed. In all aggressive male mice, serum beta-NGF levels were elevated by two orders of magnitude. The beta-NGF radioimmunoassay values were also validated by a neurite outgrowth bioassay system using the serum of aggressive male mice. In summary, both measurement techniques confirm that mouse serum beta-NGF levels undergo marked changes depending on animal handling conditions.

Aggression↗
PubMed1982

Limitations in protection afforded by gloves in laboratory handling of aflatoxins.

The permeability of latex and vinyl laboratory gloves by aflatoxins in chloroform or dimethyl sulfoxide (DMSO) has been investigated. Latex gloves provide good protection against permeation by aflatoxins in DMSO, but aflatoxins in chloroform permeate all types of laboratory gloves. If gloves are contaminated when aflatoxins in chloroform are handled, an immediate change of gloves is recommended.

Aflatoxins↗
PubMed1993

The stability of serum hepatitis C viral RNA in various handling and storage conditions.

To investigate whether certain handling and storage conditions of serum samples could alter the sensitivity and specificity of the hepatitis C virus (HCV) RNA assay, we studied serum samples obtained from five patients known to be positive for HCV RNA and two patients with autoimmune chronic active hepatitis. Samples were subjected to one of the following conditions: (1) immediate storage at -20 degrees C, (2) five freeze-thaw cycles, (3) storage at 4 degrees C for 5 days, and (4) storage at room temperature for 5 days. Detection of HCV RNA was performed by polymerase chain reaction. Titers of HCV RNA were determined by serial end point dilutions. We found that the titer of HCV RNA was reduced by only one logfold in samples subjected to conditions 2 through 4 in two of the five patients. False-positive results were not seen with the serum samples that were subjected to similar conditions from the two negative control patients. We conclude that serum HCV RNA is resistant to degradation under routine laboratory handling and storage conditions.

Blood Preservation↗
PubMed2006

Quality aspects of prenatal cytogenetic diagnosis: determining the effect of various factors involved in handling amniotic fluid and chorionic villus material for cytogenetic diagnosis.

OBJECTIVES: To investigate the effect of factors involved in cell culturing and slide preparation of amniotic fluid (AF) and chorionic villus biopsies (CVB) for prenatal cytogenetic diagnosis. METHODS: The effect on the outcome of our standard AF cell culture procedure of volume and appearance of the submitted AF specimen, gynaecologist performing the amniocentesis, week of gestation in which the specimen was taken and culture medium was retrospectively investigated. In a prospective study controlled experimental variation was introduced in composition of fixative, relative humidity, temperature and airflow during slide preparation from primary CVB and AF in situ cultures. For evaluation, analysis of regression or variance was used. RESULTS: Provided that at least 0.8 mL AF per culture dish was admitted, none of the investigated factors appeared as critical resulting in unacceptable variation in outcome. Variation in appearance of the AF had a relatively major impact: bloody or brown AF resulted in a 3 days longer culture time. To a limited degree, metaphase quality of AF and CVB cells was affected by composition of fixative, relative humidity, ambient temperature and airflow during slide preparation. CONCLUSION: Current prenatal cytogenetic practice as described here appears in general to be robust and reliable. The investigated conditions are not critical within the investigated range. Expensive measures for fine control of these conditions are, therefore, not required.

Amniotic Fluid↗
PubMed1999

Evaluating the potential occupational hazard of handling dental polymer products using the HET-CAM technique.

The irritation potencies of 8 dental polymer products, used as dental restorative materials, adhesives, or temporary constructions, were tested using the HET-CAM (hen's egg test-chorioallantoic membrane) technique. Liquid and powder components, and extracts of cured and freshly mixed non-cured materials of 5 glass ionomers, 1 bonding, 1 composite, and 1 cold-cured acrylate were examined. Results showed that the liquid component of all products had a strong irritation capacity but powder suspensions and extracts from cured and freshly mixed non-cured materials had no effect on the CAM. Thus, dental personnel who handle liquid and powder manually are exposed to components with a high irritation potential, in contrast to patients who are exposed to the cured and mixed non-cured materials with low irritation potential. This illustrates the importance of safe handling procedures and practices for dental personnel who handle non-cured polymers manually.

Animals↗
PubMed2000

An evaluation of sampling and laboratory procedures for determination of heterotrophic plate counts in dental unit waterlines.

BACKGROUND: The high numbers of heterotrophic microorganisms that have been cultured from dental unit waterlines (DUWs) have raised concern that this water may exceed suggested limits for heterotrophic plate counts (HPCs). The main purpose of this investigation was to examine HPC variability in DUWs and to examine in detail the effect of laboratory processing of water samples on HPC values. METHODS: Water samples were collected from dental offices either at the beginning of or during the clinic day and were transported to the laboratory, where they were analyzed. RESULTS: Measuring HPC levels within an office would involve testing all units, because significant differences were found between units connected to the same municipal water supply. Within a unit, the average microbial count from high-speed lines was approximately twice the average count from air/water lines. The laboratory processing of water samples significantly affected the numbers of heterotrophic microorganisms that were recovered. Incubation temperature, time and media, as well as neutralization of residual chlorine, all had significant effects on the HPC values. However, no significant differences in microbial counts were found between samples plated with the spread plate method on R2A agar and those plated with the pour plate method with Plate Count Agar. CONCLUSIONS: Dental organizations have suggested target limits in terms of numbers of heterotrophic microorganisms recovered in water from dental units, but standards for laboratory handling must be established as well. A protocol for sample collection and laboratory handling is proposed.

Bacteriological Techniques↗
PubMed1985

Facilitated determination of ionized calcium.

Using a calcium-containing heparin preparation for anticoagulation, we determined [Ca2+], the mean concentration of ionized calcium, in whole blood of 120 healthy blood-donors to be 1.23 (SD 0.04) mmol/L. Similarly, for 50 intensive-care patients selected without conscious bias, the correlation between [Ca2+] in serum (mean 1.15, SD 0.10 mmol/L) and in whole-blood samples anticoagulated with the same heparin preparation (mean 1.15, SD 0.09 mmol/L) was very good (r = 0.95). Storing samples anaerobically on ice for as long as 2 h did not alter whole-blood [Ca2+]. On the other hand, various concentrations of calcium-free heparin preparations all induced a significant decrease in measured [Ca2+]. By using whole-blood samples, rather than plasma or serum, for [Ca2+] determination with a calcium-selective electrode, repetitive measurements can be made with simple handling procedures, facilitating rapid implementation of appropriate therapeutic measures for critically ill patients.

Adult↗
PubMedDate not supplied

Anatomy, visualization and sampling of the biliary tree in animals and man.

Difficulties in obtaining proper bile samples are due to inaccessibility of the biliary tree and to distortions induced by sampling methods. One must be cognizant of the effects of diet on bile secretion and gallbladder motility. Experimental methods which interrupt the enterohepatic circulation or alter the intestinal migrating myoelectric complex induce spurious changes in bile flow and composition. Biliary tract pressure-flow relationships must be maintained or the gallbladder will be made functionless. Dead space errors lead to distortions unless studies are performed in the steady state, or dead space is measured and corrections are applied. Surgery has major effects on some parameters of interest, and animals should be allowed to recover when these are studied. The effect of the mixing of bile with other secretions in the duodenum must be considered when using bile-rich duodenal fluid. For some parameters of bile secretion, mixing is unimportant but for others, special precautions for handling bile and interpreting results are required.

Animals↗