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Comparative effects of autologous and homologous seminal plasma on the viability of largely extended boar spermatozoa.

Sperm handling, associated to artificial reproduction technologies (ART) such as in vitro fertilization (IVF) or the use of flow cytometry for cell analysis or sorting imposes volumetric extension of the sperm suspension and decreases sperm viability, presumably because of the removal of seminal plasma (SP) components. This study evaluated whether a 10% v/v of autologous SP (retrieved from the same donor boar) or homologous SP (e.g. from any of the four fertile boars included, other than the one providing the spermatozoa) would differently affect the viability of boar spermatozoa subjected to large extension in a simple saline medium [phosphate-buffered saline and 0.1% ethylenediaminetetraacetic acid (EDTA), PBSm] to a concentration of 0.3 x 10(6) spermatozoa/ml and incubated for 2 h at 30 degrees C. Sperm viability was monitored as membrane integrity [using the fluorophore carboxyfluorescein diacetate (C-FDA) and propidium iodide (PI)], mitochondrial function (using the fluorophore R-123) and motility characteristics [using Computer Assisted Sperm Analysis (CASA)]. Substraction of the SP and extension followed by incubation in PBSm significantly (p < 0.05) decreased sperm viability, which could be restored by addition of autologous SP. Furthermore, exposure of the extended spermatozoa to homologous SP (from any other individual boar) significantly (p < 0.05) varied with the source of the sire; some boars exerting beneficial effects (even surpassing the effects of the autologous SP; p < 0.05) while at least one boar negatively (p < 0.05) influencing the viability of the incubated spermatozoa. It is concluded that SP should be present when incubating highly extended spermatozoa. As a result of the obvious differences among boars, it would be advantageous to examine the ability of SP to maintain sperm viability prior to the use of SP pools during sperm handling in vitro.

Animals↗

Lag times between blood sampling, spinning and plasma glucose estimation.

This study investigated the effect of lag times between blood sampling and glucose analysis on plasma glucose results from 6 volunteers. Our aim was to determine whether glucose tolerance test protocols should include instructions on the handling of blood between sampling and analysis. Plasma glucose levels remained stable for all lag times between spinning and analysis. With a lag time between blood sampling and spinning, plasma glucose levels did not remain stable, and a significant lowering of plasma glucose was found in the first 2 hours of lag. With increased lag time there was no further decrease in plasma glucose levels. Glucose tolerance test protocols should include clear guidelines on the handling of the blood samples between collection and analysis, and the spinning down of samples needs to be prioritized.

Adult↗

Follicular flushing at oocyte retrieval: a reappraisal.

A survey of Australian assisted reproduction technique (ART) units revealed many (> 50%) use flushing of the follicle in addition to direct aspiration of the fluid during oocyte retrieval. The rationale is that flushing offers an advantage to the patient, with a larger number of oocytes being collected and thus a higher potential for pregnancy Following a complication in a patient, the medical staff determined that flushing might have been implicated. While this was later shown to be unfounded, the unit changed the method of oocyte collection, thus providing an opportunity historically to evaluate the differences between aspiration of follicular fluid alone or with additional flushing of each follicle. Thus the aim of this analysis was to test the hypothesis that aspiration alone does not effect the outcomes of ART with respect to oocyte numbers collected, their quality and subsequent fertilisation, or ultimate pregnancy rate in a large patient group (n = 2378). During the review period (1991-1993) the manufacturer of the aspiration needles; the pump and pressure used for aspiration; the staff involved in all procedures; and the premises, equipment and media used for oocyte collection and culture remained constant. Similarly the two patient groups did not differ in their demographics and physical characteristics. There was no difference (p > 0.5) in the number of oocytes collected, the number of embryos created or the pregnancy rate for that treatment cycle. There was also no significant difference between the two groups in fertilisation rates, irrespective of the type of treatment being used in vitro fertilisation (IVF), gamete intrafallopian transfer (GIFT) or intracytoplasmic sperm injection (ICSI)). The data presented in this paper are not prospective. They use historical comparison and are confounded by minor changes in ovarian stimulation protocols, but the conclusions are still valid. These data suggest there is no difference in the outcome of ART (from oocyte collection through to pregnancy rate) whether or not aspiration of follicles is accompanied by flushing. First principles of surgery advocate the shortest possible operating time, the simplest procedure and minimum amount of tissue handling as maxims for reducing complication. Therefore, as a routine, flushing would seem superfluous in ART.

Female↗

[Homogenous low-density lipoprotein assay in type III hyperlipoproteinemia].

To investigate whether homogenous analysis of serum low-density lipoprotein cholesterol(LDL-C) was applicable in type III hyperlipoproteinemic subjects, 3 reagents were used to estimate LDL-C in 2 cases with apoE 2/2 phenotype. Each measurements were compared to LDL-C levels by ultracentrifugation(density: 1.019-1.063). LDL-C levels by homogenous analysis with any of the reagents were higher than those by ultracentrifugation, and the difference was varied between the reagents: +14-24% by LDL-EX, +29-55% by Cholestest LDL, and +89-115% by Determiner LDL-C. The cross-reaction by the reagents to intermediate density lipoprotein(IDL) and very low density lipoprotein(VLDL) cholesterol was studied in a case; 22% of cholesterol in these fractions was measured by LDL-EX, 32% by Cholestest LDL, and 110% by Determiner LDL-C. A part of the sample serum was stored at -60 degrees C for several days and melted for re-analysis of homogenous LDL-C levels. After the freezing process, IDL fraction pattern on electrophoresis had been changed and homogenous LDL-C levels by all reagents were different from the measurements of native serum. These results indicated that using reagents with low cross-reaction to IDL and VLDL fractions and careful sample handling are important in homogenous LDL-C analysis in type III hyperlipoproteinemia.

Aged↗

Quantitation of estrogen receptor in seventy-five specimens of breast cancer: comparison between an immunoassay (Abbott ER-EIA monoclonal) and a [3H]estradiol binding assay based on isoelectric focusing in polyacrylamide gel.

Quantitation of estrogen receptor has been performed in cytosol prepared from 75 specimens of breast cancer tissue from patients who had not received hormonal therapy. The study was performed in order to compare an immunoassay (Abbott Laboratories, North Chicago, IL) with our currently used method for estrogen receptor analysis based on isoelectric focusing of [3H]estradiol-receptor complex in polyacrylamide gels. Using linear regression analysis, a regression coefficient (slope) of 1.30 and a correlation coefficient of 0.75 were calculated. The differences in results between the two methods are probably partly explained by the fact that the ligand-based method only measures unoccupied receptor, whereas the immunoassay detects the total amount of receptor, resulting in generally slightly higher concentrations with the latter method. However, in five of 75 specimens the ligand-based method gave a considerably higher concentration of estrogen receptor. This was most probably explained by partial proteolysis resulting in the formation of receptor fragment(s), which was undetectable with the immunoassay but detectable with the ligand-based method. These observations underline the importance of careful handling of specimens during the whole immunoassay procedure.

Acrylic Resins↗

Transport and storage of infusion supplies and blood products.

Home care nurses face many challenges in transporting and storing medications and blood products in their vehicles and in patients' homes. Unlike climate-controlled institutions, products subject to the cold of winter and the heat of summer can easily be damaged, which can be harmful to the patient. Additionally, several regulations and protocols demand that products be cared for in a certain manner, stored in the proper container, and labeled appropriately. The following information presents the current practices in storing and transporting these products safely. It can be used as a guide for policy and procedure development and a review for clinicians in assuring safe handling of these products from office to home to lab.

Blood Preservation↗

Auxotab--a device for identifying enteric bacteria.

A multitest system called the Auxotab that uses ten dehydrated reagents on a paper card has been evaluated with 417 known stock cultures of Enterobacteriaceae. In double-blind studies with the Auxotab, 87% of the strains tested were correctly identified. Results of this study indicate that there is a need for modification of the product in regard to ease of handling, time required for use, and accuracy of identification of enteric bacteria.

Bacteriological Techniques↗

Extraction of cyanobacterial endotoxin.

To simplify our efforts in acquiring toxicological information on endotoxins produced by cyanobacteria, a method development study was undertaken to identify relatively hazard-free and efficient procedures for their extraction. One article sourced and two novel methods were evaluated for their ability to extract lipopolysaccharides (LPSs) or endotoxins from cyanobacteria. The Limulus polyphemus amoebocyte lysate (LAL) assay was employed to compare the performance of a novel method utilizing a 1-butanol-water (HBW) solvent system to that of Westphal's (1965) phenol-water system (HPW) for the extraction of endotoxin from various cyanobacteria. The traditional HPW method extracted from 3- to 12-fold more endotoxin from six different cyanobacterial blooms and culture materials than did the novel HBW method. In direct contrast, the novel HBW method extracted ninefold more endotoxin from a non-microcystin producing Microcystis aeruginosa culture as compared to the HPW method. A solvent system utilizing N,N'-dimethylformamide-water (HDW) was compared to both the HPW and HBW methods for the extraction of endotoxin from natural samples of Anabaena circinalis, Microcystis flos-aquae, and a 1:1 mixture of Microcystis aeruginosa/Microcystisflos-aquae. The LAL activities of these extracts showed that the novel HDW method extracted two- and threefold more endotoxin from the Anabaena sample that did the HBW and HPW methods, respectively. The HDW method also extracted approximately 1.5-fold more endotoxin from the Microcystis flos-aquae sample as compared to both the HBW and HPW methods. On the other hand, the HBW method extracted 2- and 14-fold more endotoxin from the Microcystis flos-aquae/Microcystis aeruginosa mixture than did the HPW and HDW methods, respectively. Results of this study demonstrate that significant disparities exist between the physicochemical properties of the cell wall constituents not only of different cyanobacterial species but also of different strains of the same cyanobacterial species, as showing by the varying effectiveness of the solvent systems investigated. Therefore, a sole method cannot be regarded as universal and superior for the extraction of endotoxins from cyanobacteria. Nevertheless, the ability of the novel HBW and HDW methods to utilize easily handled organic solvents that are less hazardous than phenol render them attractive alternatives to the standard HPW method.

Biological Assay↗

Identification of crystals in synovial fluids and joint tissues.

The identification of crystals in synovial fluids and joint tissues is the most rapid and accurate method of diagnosing the common forms of crystal-associated arthritis. Although there are numerous methods available for identifying and characterizing crystals in biologic specimens including x-ray crystallography and Fourier transform infrared spectroscopy, in practice, polarizing light microscopy is used almost exclusively for articular crystals. Unfortunately, problems with reliability and reproducibility undercut the usefulness of this simple procedure. This article highlights recent developments in the field and discusses the importance of identifying synovial fluid crystals, proper handling of specimens, and the appropriate use of available technologies for crystal identification.

Aged↗

Protocol for the pathologic examination and tissue processing of the mammographically directed breast biopsy.

Achieving the maximum yield of breast cancers detected by mammography has required certain changes in tissue handling and examination of the mammographically directed breast biopsy. This new radiographic technique, increased use of breast-conserving surgical approaches for the treatment of breast cancer, more enlightened and demanding patients, and increasing medical-legal exposure have all contributed to changes in the way surgical pathologists should process and sample breast biopsy specimens.

Biopsy↗

Measures of thrombosis and fibrinolysis.

Our recent understanding of acute coronary syndrome as an atherothrombotic process has led to research efforts in the development of markers of thrombosis and fibrinolysis for risk prediction in cardiovascular heart disease. Although American Heart Association/American College of Cardiology guidelines recommend fibrinogen as a category I risk factor and also suggest factor VII, plasminogen activator inhibitor-1, tissue-type plasminogen activator, and von Willebrand factor as other potentially clinically useful markers, these tests have not come into routine clinical use. Their development as predictors of risk may be hampered by inconsistent laboratory methodology, which causes difficulty in comparing result interpretation with published trial studies. This article presents the history of development for these tests, proper laboratory handling, the best trial data that present evidence of their accuracy, and current guidelines for clinical use.

Biomarkers↗

Extremity dosimetry for radiation workers handling unsealed radionuclides in nuclear medicine departments in India.

In India, for the past five decades, whole body radiation dose of radiation workers has been monitored by means of film and thermoluminescent dosimeter (TLD) badges worn on the body. However, there are no provision/regulatory requirements to monitor doses received at the extremities, i.e., to fingers. Finger dose monitoring is essential for controlling the extremity dose limits for occupational personnel handling unsealed radioactive sources. In order to estimate the doses received in various types of procedures using unsealed sources, finger dose monitoring was carried out in 54 major institutions in the country using a specially designed plastic finger ring embedded with a TLD. The maximum finger dose of occupational workers involved in handling Tc in such activities as extraction and radiopharmacy work is 0.35 mSv GBq; during injection of radiopharmaceuticals and scintigraphy, the doses were observed to be 1 and 0.95 mSv GBq, respectively. Similarly, while handling F-FDG, the maximum doses received during dispensing, injection, and scintigraphy were 0.098, 0.324, and 0.56 mSv GBq, respectively. The maximum radiation dose received during Re/Re balloon angioplasty and while handling Sm was 3.92 and 6.5 mSv GBq, respectively. All the doses recorded were well within the prescribed limit. However, monitoring of these doses periodically would help in compiling the feedback regarding the work practices followed in institutions handling radioisotopes in the country and would also help in maintaining a record of safe work procedures while handling radioisotopes.

Equipment Design↗

Coagulation tests in differential diagnosis.

Bleeding disorders pose a special problem for the practising physician haematologist or laboratory coagulation specialist. Great concern is placed on specimen integrity, either during handling or in storage. Heparin contamination, activated specimens and factor VIII lability are common issues.

Blood Coagulation Disorders↗

The pathologic handling of skeletal tumors.

Because bone harbors a wide variety of benign and malignant pathologic entities, most of which are uncommon, pathologists feel uneasy when faced with many of these lesions. Accurate diagnoses of these lesions require correlation of the radiologic and clinical findings with the pathologic features. At the time of biopsy, placing a lesion into one of five pathologic groups based on the predominant cell or matrix type present (osteoid, chondroid, giant cell, fibrous, small cell) aids in analyzing the specimen. Current management of malignant bone tumors includes diagnosis on often tiny biopsy samples followed by preoperative chemotherapy and finally limb-sparing resection of the involved bone. Adequate evaluation of these resections requires extensive examination with grading of the tumor necrosis and careful attention to the resection margins.

Adolescent↗

Artifactual hydropic degeneration in skin biopsy specimens immersed in saline: a light and electron microscopic study.

An artifact, consisting of hydropic degeneration of the basal cells and subepidermal bulla formation, is described in skin-punch biopsy specimens immersed in normal saline. The histologic appearance and ultrastructural features of this artifact are discussed. Proper handling of skin specimens intended for immunofluorescent studies is stressed. Similarities are noted between the histologic and ultrastructural findings of this artifact and of epidermolysis bullosa simplex and pathologic hydropic degeneration, as is seen in lupus erythematosus.

Arteriosclerosis↗