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PubMed2000

Challenges in ophthalmic pathology: the vitreoretinal membrane biopsy.

The introduction of vitreoretinal microsurgery has produced a new type of biopsy; that of the vitreoretinal membrane. This review investigates methods by which these scar-like tissues are handled in the laboratory and explores the implications of the results of such evaluations. The study of vitreoretinal membrane biopsies has provided much information concerning the pathobiology of the various conditions which may give rise to the tissue as well as insights into how membranes themselves develop. Moreover, the application of new laboratory techniques is expected to enhance our understanding of the formation of vitreoretinal membranes, and lead to further advances in their surgical and medical management.

Biopsy↗
PubMed1988

Recommendations regarding handling of the necropsy in Rett syndrome.

In order to maximize biochemical, neurochemical, molecular, and pathologic information from patients with Rett syndrome, a uniform procedure has been developed for the conduct of the postmortem examination. Tissue should be prepared for freezing, for electron microscopy studies, and for standard histologic examination. For the purposes of uniformity, three central repositories for necropsy materials have been established and are available at all times. In the event of the death of a patient with Rett syndrome, parents who consent to necropsy should request that materials be handled according to the protocol and transported under appropriate conditions to the central repository. The National Neurological Research Bank (Los Angeles), the Brain Tissue Bank (Belmont, Mass), and the Department of Neuropathology at Johns Hopkins Hospital (Baltimore) have agreed to serve as repositories for tissues. A committee of the International Rett Syndrome Association medical advisory panel will monitor this process in cooperation with the directors of the respective tissue banks.

Ammonia↗
PubMedDate not supplied

Flexible automation of cell culture and tissue engineering tasks.

Until now, the predominant use cases of industrial robots have been routine handling tasks in the automotive industry. In biotechnology and tissue engineering, in contrast, only very few tasks have been automated with robots. New developments in robot platform and robot sensor technology, however, make it possible to automate plants that largely depend on human interaction with the production process, e.g., for material and cell culture fluid handling, transportation, operation of equipment, and maintenance. In this paper we present a robot system that lends itself to automating routine tasks in biotechnology but also has the potential to automate other production facilities that are similar in process structure. After motivating the design goals, we describe the system and its operation, illustrate sample runs, and give an assessment of the advantages. We conclude this paper by giving an outlook on possible further developments.

Artificial Intelligence↗
PubMed2003

The accuracy of guestimates.

At times a clinician must describe the size of a lesion when measurement presents difficulties. A 'guestimate' may then be recorded. We tested the accuracy of forensic clinicians (10 physicians and 1 nurse) in guestimating the sizes of ten test objects, comparing their performance with 13 individuals from other professions. The participants were permitted to handle the objects but not to use a measuring device. For three of the objects (a balloon, a scratch and a coin) the entire sample of participants significantly overestimated size, by 13-22%. Both participant groups overestimated the size of a scratch, the clinicians being more accurate but not significantly so. Guestimates should be avoided unless the use of a ruler or tape measure will be against the interests of the client.

Adult↗
PubMed1993

[Final goal and problems in clinical chemistry examination measured by advanced analytical instruments].

In the field of clinical chemistry of Japan, the automation of analytical instruments first appeared in the 1960's with the rapid developments in electronics industry. After a series of improvements and modifications in the past thirty years, these analytical instruments became excellent with multifunctions. From the results of these developments, it is now well recognized that automated analytical instruments are indispensable to manage the modern clinical Laboratory. On the other hand, these automated analytical instruments uncovered the various problems which had been hitherto undetected when the manually-operated instruments were used. For instances, the variation of commercially available standard solutions due to the lack of government control causes the different values obtained in institutions. In addition, there are many problems such as a shortage of medical technologists, a complication to handle the sampling and an increased labor costs. Furthermore, the inadequacies in maintenance activities cause the frequent erroneous reports of laboratory findings in spite of the latest and efficient analytical instruments equipped. Thus, the working process in clinical laboratory must be systematized to create the rapidity and the effectiveness. In the present report, we review the developmental history of automation system for analytical instruments, discuss the problems to create the effective clinical laboratory and explore the ways to deal with these emerging issues for the automation technology in clinical laboratory.

Autoanalysis↗
PubMed1985

Formation of free fatty acid and ceramide during brain handling: lability of sphingomyelin.

Intact brain and brain homogenates readily form free fatty acids and ceramides, even in the cold during subcellular isolation procedures. The fatty acid formation is slightly stimulated by chelators and might be due to phospholipid hydrolysis by lysosomal phospholipases. The ceramide formation is accompanied by loss of sphingomyelin and is apparently due to the action of neutral, metal ion-activated sphingomyelinase. The latter reaction is inhibited by EDTA whereas both degradative processes are inhibited by mercuriphenylsulfonate, the thiol-reacting inhibitor. Cerebroside does not seem to be a source of accumulated ceramide.

4-Chloromercuribenzenesulfonate↗
PubMed1995

Consensus on minimal criteria of clinical and neuropathological diagnosis of schizophrenia and affective disorders for post mortem research. Report from the European Dementia and Schizophrenia Network (BIOMED I).

The sophisticated analysis of and growing information on the human brain requires that acquisition, dissection, storage and distribution of rare material are managed in a professional way. In this publication we present the consensus of the European work group "European Dementia and Schizophrenia Network", granted by the BIOMED I project of the EU, on minimal neuropathological and clinical requirements to include brains of patients with schizophrenia and affective disorders in post mortem studies. The description of clinical prerequisites in different EU countries and institutions is followed by a consensus on tissue handling, a consensus on minimal neuropathological criteria and a consensus on minimal clinical diagnostic criteria including clinical vignette, family, social, education/professional and general medical histories, general physical history including neurostatus, neurological, psychiatric, medication and general pathological histories, psychostatus, laboratory tests and a history provided by family/health care giver questionnaire. This publication should give help to interconnect different European brain bank centers on a basis of standardized protocols.

Brain↗
PubMed2001

Measurement of ammonia in blood.

The measurement of ammonia, now known to be a normal constituent of all body fluids, is fraught with problems. An elevated ammonia level in blood (100 micromol/L or higher) is an indicator of an abnormality in nitrogen homeostasis. The collection, handling, storage, and analysis of blood samples, their limitations, and potential sources of error are discussed. New techniques that permit continuous or real-time estimates of systemic ammonia levels over a broad range are also discussed. The aim should always be to minimize the release of ammonia from the collected sample before analysis. Recommendations are made on the collection and processing of blood samples, for it is by standardization and rigid adherence to these techniques that the reliability of the test results will be improved.

Ammonia↗
PubMed1989

Problems in measuring human leucocyte cation content: effects of cell preparation and handling.

1. Previous published measurements of leucocyte cation content are inconsistent, with sodium concentrations having particularly high coefficients of variation. We have measured the effects of recovery time after isolation, different types of handling, and centrifugation on leucocyte sodium and potassium content. 2. Sodium content fell markedly during the first 30 min after isolation and was stable from 1 to 3 h of incubation. There was a small but significant rise in potassium content over the same time period. 3. During incubation, occasional gentle resuspension of the cells gave optimal sodium contents, whereas mixing by inversion caused large falls in sodium and potassium content. 4. Centrifugation of stable cells at 200 g for 6 min caused marked increases in sodium content.

Cell Separation↗
PubMedDate not supplied

Intestinal nutrient uptake measurements and tissue damage: validating the everted sleeves method.

The reliability of methods for nutrient uptake measurements across the intestinal epithelium relies on the integrity of the mucosal epithelium and the enterocytes. We tested effects of tissue handling during the "everted sleeves method" on the length of intestinal villi, the surface magnification, the circumference of the gut, and the thickness of the muscle layer in sunbirds (Nectarinia osea), chicken (Gallus gallus), and mice (Mus domesticus). The sunbird has thin and delicate intestinal villi that are greatly affected by the everted sleeves method. After eversion and incubation, villi lost 30% of their original length. The severe tissue damage coincides with uptake measurements for glucose that were an order of magnitude lower than in other nectar-feeding (nectarivorous) birds of similar body size. Tissue handling during the everted sleeves method had significant effects on morphometric parameters of chicken and mouse intestines, but on a light-microscopical level, the tissue integrity and the cytology of the enterocytes were not altered. Therefore, we think that the everted sleeves method renders reliable and reproducible measurements of nutrient uptake in those species. We conclude that a histological evaluation is necessary to assess the reliability of the method before it is applied to adults or to the developmental stage of any species.

Animals↗
PubMedDate not supplied

Ecological factors affecting the adrenocortical response to stress in chestnut-collared and McCown's longspurs (Calcarius ornatus, Calcarius mccownii).

Secretion of the steroid hormone corticosterone (CORT) in response to perturbations results in behavior patterns that can maximize survival. Three commonly cited hypotheses suggest that during breeding, there are advantages associated with suppressed (CORT) secretion in (1) the sex that is most important for parental care, (2) individuals in good body condition, and (3) species with a short breeding season and limited renesting opportunities. We addressed these hypotheses in two midlatitude breeders, chestnut-collared and McCown's longspurs, by assessing CORT secretion over a 1-h period of handling. These species have congeners that are exclusively arctic breeders, and this provides a unique opportunity to assess adrenocortical responsiveness both within a phylogenetic framework and across environmental variables. In both species, males and females showed similar CORT secretion patterns in response to handling, and body condition was unrelated to CORT secretion. Additionally, although these midlatitude breeders have more opportunities to attempt a renest than their arctic congeners, their hormonal response to stress was similar to that reported for their arctic congeners. We suggest that an attenuated stress response may relate to the severity of the breeding environment and a low likelihood of successful renesting, since these variables are common to both arctic and grassland habitats.

Analysis of Variance↗
PubMed2003

Mass spectrometric analysis of chemical warfare agents and their degradation products in soil and synthetic samples.

A packed capillary liquid chromatography-electrospray ionization mass spectrometry (LC-ESI-MS) method was developed for the identification of chemical warfare agents, their degradation products and related compounds in synthetic tabun samples and in soil samples collected from a former mustard storage site. A number of organophosphorus and organosulfur compounds that had not been previously characterized were identified, based on acquired high-resolution ESI-MS data. At lower sampling cone voltages, the ESI mass spectra were dominated by protonated, sodiated and protonated acetonitrile adducts and/or their dimers that could be used to confirm the molecular mass of each compound. Structural information was obtained by inducing product ion formation in the ESI interface at higher sampling cone voltages. Representative ESI-MS mass spectra for previously uncharacterized compounds were incorporated into a database as part of an on-going effort in chemical warfare agent detection and identification. The same samples were also analyzed by capillary column gas chromatography (GC)-MS in order to compare an established method with LC-ESI-MS for chemical warfare agent identification. Analysis times and full-scanning sensitivities were similar for both methods, with differences being associated with sample matrix, ease of ionization and compound volatility. GC-MS would be preferred for organic extracts and must be used for the determination of mustard and relatively non-polar organosulfur degradation products, including 1,4- thioxane and 1,4-dithiane, as these compounds do not ionize during ESI-MS. Diols, formed following hydrolysis of mustard and longer-chain sulfur vesicants, may be analyzed using both methods with LC-ESI-MS providing improved chromatographic peak shape. Aqueous samples and extracts would, typically, be analyzed by LC-ESI-MS, since these analyses may be conducted directly without the need for additional sample handling and/or derivatization associated with GC-MS determinations. Organophosphorus compounds, including chemical warfare agents, related compounds and lower volatility hydrolysis products may all be determined during a single LC-ESI- MS analysis. Derivatization of chemical warfare agent hydrolysis products and other compounds with hydroxyl substitution would be required prior to GC-MS analysis, giving LC-ESI-MS a definite advantage over GC-MS for the analysis of samples containing chemical warfare agents and/or their hydrolysis products.

Biodegradation, Environmental↗
PubMed2000

A systematic and quantitative analysis of PCR template contamination.

A quantitative and systematic analysis is provided for ubiquitously present template DNA interfering with the quantification of human DNA by PCR. Two sources contributing to DNA background were identified. The first one is interpreted as DNA present in chemicals and on equipment and the second as caused by operator handling. The amounts were equivalent to 2.5 and 8.9 pg per mL of sample, and the estimated frequencies of contamination were 65 and 35%, respectively, resulting in an effective limit of detection of 17.4 pg/mL. Below this level--named effective laboratory background--a result could not be considered as authentic. Knowledge of these parameters is important for laboratories that analyze minute amounts of human DNA by PCR for purposes such as quantification, typing, and sequencing.

DNA Fingerprinting↗
PubMed1997

Viability during storage and stability of plasmids during storage and subculturing in strains of Vibrio anguillarum.

The stability of plasmids, 3.3 kb - approximately 200 kb, in 8 strains of Vibrio anguillarum displaying different plasmid profiles and reactions with O-antisera was investigated over an 18 months period. All plasmid profiles proved to be resistant to storage at different temperatures but strains stored at 37 degrees C were only viable for a short period of maximum 2 months. Strains stored at 5 degrees C and 20 degrees C were viable for a longer period. Viable strains maintained their plasmid profile throughout the experiment, except in 2 cases where a 67 kb and a 200 kb plasmid were lost. Strains stored at -80 degrees C all remained viable and maintained their plasmid profile throughout the study. By subcultivating daily for up to 100 successive days, most strains maintained their plasmid profiles. Only 2 strains lost their plasmid. When picking 100 single colonies from agar plates, none of the colonies showed plasmid profiles deviating from the expected. The results suggest that plasmid profiles among V. anguillarium are very stable during subculturing, storage and laboratory handling using standard laboratory procedures, and thus, reliable for epidemiological investigations. In a second experiment, 2 pairs of 2 strains were grown together in mixed cultures. They were identical in all traits, except that one strain in each pair harboured the 67 kb pJM1-like virulence plasmid, whereas the other had lost this plasmid. The result showed that the growth rate was the same for strains with and without the plasmid, indicating that under laboratory conditions, this plasmid is neither a benefit nor the opposite for bacterial growth.

Animals↗
PubMed2001

Handling forensic necropsy cases.

Immediately record the findings. A tape recorder would be excellent for this. If none is available, findings should be written down as soon as possible before memory of the examination fades. It is important to record comments on every organ that you examined. Three years from now in a court deposition, it may be difficult for the practitioner to remember if he looked at the adrenal glands or if there was no gross evidence of hyperplasia. When writing a report, it is a good idea to describe the tissue changes rather than just to give an interpretation. An example of the latter would be, "The horse had a severe pneumonia." A better method might be, "The horse's lungs had a bilaterally symmetric pneumonia in the ventral portions of the anterior, middle, and diaphragmatic lung lobes. This pneumonia accounted for roughly 45% of the total lung area. These pneumonic areas were dark red, sunken, and firm with a dull granular pleura. When cut, there was a yellow exudate that exuded from the cut surface." This latter description gives good evidence of why it was believed that the lung was pneumonic and just how severe the change was. Several years from now, the latter description does more to refresh the memory of the case. Carefully label and preserve the specimens for later diagnostic tests. List all the fresh samples collected. Time spent on these details now can save you considerable embarrassment later. Following these suggestions should allow the practitioner to provide the best possible service to all parties involved.

Animals↗
PubMed1989

Sample preparation for the HPLC analysis of drugs in biological fluids.

Sample preparation for the analysis of drugs in biological fluids consists of a number of unit operations that are used for (i) release of the drug from a conjugate or biological matrix; (ii) removal of endogenous compounds that could interfere with the assay; and (iii) techniques for liquid handling. The trends in sample preparation that have occurred over the past 10 years in the authors' laboratory are discussed. In general, there has been a move from the traditional liquid-liquid extraction to methods using bonded-silica which permit rapid throughput and efficient extraction. Automation of sample preparation has seen further gains in productivity; however, the present generation of equipment lack the control and communication systems that are essential for the development of the automated integrated laboratory of the future.

Automation↗
PubMed1982

[System of temporary blocking of transporting device of the universal automat of the AT-4 type for histological handling and staining of tissues].

A rapid transfer of the box with tissues from vial to vial by the AT-4 automate (10 seconds) results in premature contamination of the subsequent reagent with previous one causing excessive use of scarce and expensive reagents. A system of temporary blocking of the transporting device is described which permits one to arrest the box above the vial for 30 sec in which time all the reagents flows back into the vial from the box. The use of the blocking of the transporting device for 360 sec in each cycle of tissue staining (12 vials) does not reduce the working capacity of the AT-4 apparatus but markedly saves the amount of reagents used and improves the quality of treatment.

Histological Techniques↗
PubMedDate not supplied

Processing fine needle aspiration biopsies for electron microscopic examination. Experience implementing a procedure.

While processing needle rinses from fine needle aspiration biopsy material for electron microscopy, we experienced difficulties not fully outlined previously. Problems in preservation were traced to the time elapsed before glutaraldehyde was added to the needle rinse, which had been held in a physiologic transport medium; a tendency toward hypotonicity of the commercially prepared transport medium; and vigorous handling of the needle rinse. When these problems were dealt with, preservation improved dramatically. The problem of scanty material available for ultrastructural evaluation was dealt with by decreasing the number of smears made and filtering the needle rinse to capture tissue fragments. Implementation of the procedure required leadership, good communication, motivated cytotechnologists and cytopathologists, and continuous supervision and monitoring. In cases with adequately cellular well-preserved material available for electron microscopy, ultrastructural evaluation made a contribution of clinical value to the diagnosis in 48% of cases.

Biopsy, Needle↗