Comparison of Conventional Methods, the R/B System, and Modified R/B System as Guides to the Major Divisions of Enterobacteriaceae

Primary grouping of more than 2,000 members of Enterobacteriaceae, freshly isolated from clinical specimens, was performed on the basis of eight conventional laboratory tests used routinely in this laboratory and by the R/B system. Results indicate that both systems perform well in this first step in the identification of various medically significant species of the family. Difficulties encountered with certain reactions of the R/B system were corrected with a physical modification of the tubes. An additional 100 representatives of Enterobacteriaceae were compared with conventional methods and the modified R/B system. Results indicate improvement in observing and interpreting findings with the R/B system.     

Identification of bacterial strains by laboratories participating in the Deutsches Krebsforschungszentrum quality assurance programme

The quality assurance programme (QAP) of the Deutsches Krebsforschungszentrum (DKFZ) is a proficiency testing system developed to service the laboratory animal discipline. QAP comprises the quarterly distribution of two bacterial strains originating from various species of animals for identification to the species level and antibiotic susceptibility testing. We compared identification results reported by QAP participants over the years 1996-2004 with those obtained by the Dutch Bacterial Diagnostics reference laboratory on 68 samples comprising 71 bacterial strains and a fungus. Significant differences were found in the frequency of reported and correct identifications when bacteria were assigned to different groups based on morphology by Gram stain and on origin (animal versus environmental, rodent and rabbit versus other animal species, pathogen versus non-pathogens). Rodent and rabbit pathogens yielded 73% correct identifications, and with all bacterial strains only 60% of the identifications were correct. We assume that most QAP participants were from laboratory animal diagnostic laboratories. If this is true, the capabilities of laboratories in the laboratory animal discipline to correctly identify bacterial species are well below what are considered acceptable limits for human diagnostic laboratories. The distribution of cultured bacteria circumvents the most difficult step in the microbiological monitoring of animals, namely primary culture from clinical samples. We propose to set up a QAP that comprises the distribution of specimens mimicking clinical samples normally submitted to laboratory animal diagnostic laboratories.     

检验医学专业临床实习管理存在的问题及对策

临床实习是医学理论和临床实践工作有机结合的连接点,是医学生走向临床工作必不可少的重要环节。临床实习教学是医学生教育中一个重要的组成部分,是从课堂到临床、从理论到实践、从理性到感性的重要过程,也是巩固基础知识、理论联系实际、走向独立工作岗位的重要一步。在临床实践过程中,教职人员要培养学生获取、分析和处理疾病信息的能力,正确书写病历、熟练掌握诊疗技术以及与患者及家属和谐沟通的能力。医学检验学是运用现代物理化学方法、手段进行医学诊断的一门学科,通俗的说,就是利用仪器、试剂、CT扫描等检测人的各项指标是否正常,为临床诊断提供依据。因此,加强临床检验医学专业的实习管理工作至关重要。本文从建立合理的教学管理体系、加强临床实习考核制度、完善实习管理、加强教师队伍建设等几个方面,探讨临床检验学实习管理存在的问题,并提出相应的对策,为高等医学院校教学管理工作提供思路。     

Development and evaluation of a multi-applicator system of endocavitary radiothermotherapy of gynecologic tumors

An applicator system for the radiothermotherapy of gynecological tumors has been developed. The concept is based on combining high-dose rate afterloading therapy with local hyperthermia (27.12 MHz). The main applicator, the rf-gamma applicator consists of a guide tube for the gamma source, an electrode system and a cooling system. Isodoses and isotherms can be adapted to the individual anatomical-pathological situation presenting. For therapy planning the interaction between the applicators and the perfused tissue was investigated in a theoretical FEM model. In the first step, the SAR-function in the vicinity of the rf-applicator was determined by means of a 2D FEM calculation. In a second step, the 3D temperature fields was determined using linear shape functions. The results of these calculations showed that in every case the hot spots shifted from the applicator surface into the depths of the tissue. With the aid of an infrared camera and a split phantom the calculations were examined in a homogeneous non-perfused tissue model. Thermometry confirmed the accuracy of the results obtained. The radiothermotherapy system described here was tested in animal experiments, and is presently being used in a clinical pilot study.     

Aerotolerance of human clinical isolates of Prevotella spp

AIMS: To determine the influence of oxygen exposure and growth stages on oxygen tolerance for clinical and reference specimens of the genus Prevotella. METHODS AND RESULTS: Tolerance to oxygen exposure was evaluated along growth stages for a total of four Prevotella isolates constituted by two strains tested soon after isolation from periodontally compromised patients and two reference strains kept frozen (-86 degrees C) in our laboratory collection. Additionally, the recently recovered isolates were also assayed after 4 months exposed to oxygen during laboratory handling. On solid medium, recently recovered Prevotella specimens proved to be less tolerant to oxygen exposure soon after isolation than when exposed for 4 months to oxygen during laboratory handling. Oxygen resistance of reference strains maintained in the laboratory collection showed to be the highest when compared with the clinical isolates both before and after oxygen exposure. When oxygen tolerance was tested during growth, bacterial cells from the exponential phase were more tolerant. Differences were observed in protein patterns between clinical isolates soon after recovery and after laboratory handling as determined by SDS-PAGE of crude cell-free extracts. However, SOD activities were similar in all bacterial samples tested. CONCLUSIONS: The ability to adaptively change oxygen tolerance was observed for pathogenic specimens of Prevotella. This property may be considered a virulence factor. SIGNIFICANCE AND IMPACT OF THE STUDY: Adaptative aerotolerance of Prevotella as a factor for virulence and survival.     

Evaluation of the BBL Minitek System for the Identification of Enterobacteriaceae

Thirty-one different substrate disks were tested in parallel with comparable, prepared media (BBL) against a minimum of 300 cultures of Enterobacteriaceae. An overall correlation of 98% was observed with all the disks tested. In addition, the system was used to identify 461 fresh isolates of Enterobacteriaceae in parallel with conventional media using the schema used at the Veterans Administration Hospital, Baltimore. An overall correlation of 97% was observed. Minitek is a time and space saving system. It is accurate and easily adapted to the clinical laboratory. A wide variety of substrates are available, allowing most laboratories to use their own schema. The long shelf life of most disks is a definite advantage.     

Automated DNA mutation analysis by single-strand conformation polymorphism using capillary electrophoresis with laser-induced fluorescence detection

Automation is essential for rapid genetic-based mutation analysis in clinical laboratory to screen a large number of DNA samples. We propose in this report an automatic process using Beckman Coulter P/ACE™ capillary electrophoresis (CE) with laser-induced fluorescence (LIF) system to detect a single-point mutation in the codon 12 of human K-ras gene. Polymerase chain reaction (PCR) using a fluorescently labeled reverse primer and a plain forward primer to specifically amplify a selected 50 bp DNA fragment in human K-ras gene. The amplified DNA is placed on the sample tray of the CE system with a pre-programmed step for single-strand conformation polymorphism (SSCP) analysis. Sample injection and denaturation processes are performed online along with separation and real-time data analysis. The concept of automation for rapid DNA mutation analysis using CE-LIF system for SSCP is presented.     

Evaluation of a new algorithm to determine the hip joint center

Accurately locating the hip joint center is a challenging and important step in many biomechanical investigations. The purpose of this study was to test the accuracy and robustness of a "pivoting" algorithm used to locate the hip center. We tested the performance of this algorithm with data acquired by manipulating a ball and socket model of the hip through several motion patterns. The smallest mean errors of 2.2+/-0.2 mm occurred with a circumduction motion pattern, while the largest errors of 4.2+/-1.3 mm occurred with single-plane motion (e.g., flexion/extension). Introducing random noise with an amplitude of 30 mm increased the errors by only 1.3+/-0.5 mm with a circumduction motion pattern. The pivoting algorithm performs well in the laboratory, and further work is warranted to evaluate its performance in a clinical setting.     

Clinical performance of a system for semiautomated chromosome analysis.

Until recently equipment for automated chromosome analysis has not been used for routine purposes in clinical cytogenetic laboratories. During a 3 1/2-year period the chromosome laboratory of Rigshospitalet has tested the Magiscan chromosome system under routine conditions and performed the first evaluation of its clinical performance. The system consists of an image processor with a light pen for manual interaction connected to a hard-copy printer and a microscope with a TV camera and a motorized scanning stage for eight slides. Automated metaphase finding takes place without operator assistance. An operator is involved in the analysis after the metaphases are located. Using two of these complete systems, we have performed a total of 4,691 chromosome analyses comprising a count of 10 metaphases, of which three were "eyeball" karyotyped and one was "machine" karyotyped. Presently, two-thirds of our prenatal analyses (amniotic-cell cultures) are carried out with these two machines. A third Magiscan system without scanning stage is used as a "karyotyping-only" system to produce hard-copy karyograms in those cases in which metaphases are manually located and counted in the microscope. Since the end of 1984, 4,773 additional machine karyograms have been produced with this system. With a complete system, a prenatal analysis can be carried out in an average of 35 min. The average time for a machine karyotype is 7 min. Since 1984 the productivity of the laboratory has increased 17%-20% without enlarging the staff.     

Comparison of five methods for determination of total plasma protein concentration

Quantitation of exact total protein content is often a key step and is common to many applications in general biochemistry research and routine clinical laboratory practice. Before embarking on any type of protein analysis, particularly comparative techniques, it is important to accurately quantitate the amount of protein in the sample. In order to assess the quality of total protein estimation results, five methods were tested and were applied to the same pooled plasma sample. For this aim, Bradford (Coomassie Brilliant Blue), Lowry (Folin-Ciocalteau), Biüret, Pesce and Strande (Ponceau-S/TCA), and modified method of Schaffner-Weismann (Amido Black 10B) were used. The last two methods employ simultaneous precipitation of proteins with the acid containing dye solutions followed by dissolution of precipitate in a NaOH solution. It is shown that each assay has advantages and disadvantages relative to sensitivity, ease of performance, acceptance in literature, accuracy and reproducibility/coefficient of variation. All of the methods tested show a CV % < 6. Besides pooled plasma, a known concentration of human serum albumin was also analyzed and discussed by means of standardization of plasma total protein content.     

Laboratory technicians; the Clinical Laboratory Law and its meaning to private physicians

The present laws and regulations relating to clinical laboratories in California are the outcome of over a quarter century of cooperative development. The medical profession, public health department, laboratory workers, and the legislature have worked together in this development.At first the system of certifying technicians and laboratories was on a voluntary basis. The clinical laboratory law in effect legalized and made generally applicable a system which had already been accepted voluntarily. The application of the clinical laboratory law provides physicians a reasonable assurance that competence and reliability will prevail in clinical laboratory operation. Of great importance is the conduct of proper training programs by approved laboratories. Since modern medical practice is so dependent on accurate clinical laboratory work it is essential that special effort be directed by physicians toward influencing young people to enter the profession of medical technology.     

Compact optical microfluidic uric acid analysis system

We designed, fabricated and tested a novel compact fluorescence analysis system for quantification of uric acid (UA) in clinical samples at the point-of-care. To perform an analysis, diluted saliva, urine or blood samples are simply placed in a disposable thin-film sample holder using a dropper. A new enzyme immobilization technique was developed to retain within the sample holder two enzymes and a molecule, which transforms into a fluorescer in amounts depending on the UA concentration. The small instrument (7.5 cm × 5 cm × 5 cm) into which the sample holder is placed for analysis contains an LED, a narrow-band filter and an amplified photodiode. The analysis time is 30s, and the dynamic range of the system is 4-400 μM of UA. The calibration curve for transparent saliva and urine was made using solutions of UA. The calibration curve for opaque blood was obtained with spiked samples of blood. The three different types of clinical samples were collected from three subjects and simply diluted before their measurements. Analysis with our instrument yielded UA concentrations within the expected concentration ranges. Development of instruments based on the current laboratory prototype is expected to result in products for clinical trials and point-of-care.     

The role of quality control in a skin bank: tissue viability determination.

New surgical procedures requiring viable skin have increased rapidly over the last few years. The cell viability assessment in allograft skin is a major step forward in burn treatment, since it is well-known that taking is correlated with grafted tissue viability. Various methods, both qualitative and quantitative, are currently used. Although qualitative assays (histomorphology, immunocytochemistry) are routinely performed in our laboratory, there arose a need to set up a standardised quantitative assay in an attempt to obtain a cut-off value so that the skin sample could be determined valid or not for grafting. Therefore, two different tetrazolium salt compounds MTT and WST-1, were compared in order to determine their efficacy in the evaluation of tissue viability. Several experimental conditions were analysed: 1- cellular cultures of keratinocytes and fibroblasts, 2- fresh skin tissue samples, 3- the same specimen tested daily for at least 2 weeks, 4- after cryopreservation and thawing. Viable cells were analysed by the cleavage of tetrazolium salts to formazan by cellular enzymes. The formazan dye produced by metabolically active cells was then quantified by measuring the absorbance of the dye solution at the appropriate wavelength. It was seen that WST-1 is easier to handle, more stable, has a wider line arrange, accelerated colour development and is more sensitive than MTT on fresh specimens and cell suspension. However, after 72 hours of storage at 4°C, most of the WST-1 tested specimens no longer gave any absorbance signal, whilst MTT specimens were seen to give a signal for more than two weeks. Moreover, after thawing WST-1 tested samples were almost negative,whilst MTT samples continued to give strong signals. In conclusion, WST-1 assay offers rapid and precise results as to the cell viability of fresh allografts and cell cultures, whilst the MTT method is much more useful in establishing viability after long conservation and cryopreservation. In our clinical experience, allografts transplanted at 72 hr post-harvesting or after cryopreservation showed a mean of take more than of 80%, demonstrating that the MTT system is more reliable for the determination of allograft viability. Studies are ongoing with larger clinical cohorts to establish the precise cut-off value for skin graft validation. This revised version was published online in July 2006 with corrections to the Cover Date.     

Applications of a signal averager for neurophysiological investigations in clinical function laboratories using a general signal-processing unit

A general 'coherent signal averager' software package which can be run on a small laboratory computer is presented as an application of a new approach to medical instrumentation. The combination of the minicomputer, preprocessing hardware and the above-mentioned software yields a flexible multipurpose averaging system for electrophysiological signals. The possibilities of the system are discussed with reference to visual evoked potential measurements in a clinical function laboratory.     

A Laboratory Information Management System (LIMS) for a high throughput genetic platform aimed at candidate gene mutation screening

High throughput mutation screening in an automated environment generates large data sets that have to be organized and stored reliably. Complex multistep workflows require strict process management and careful data tracking. We have developed a Laboratory Information Management Systems (LIMS) tailored to high throughput candidate gene mutation scanning and resequencing that respects these requirements. Designed with a client/server architecture, our system is platform independent and based on open-source tools from the database to the web application development strategy. Flexible, expandable and secure, the LIMS, by communicating with most of the laboratory instruments and robots, tracks samples and laboratory information, capturing data at every step of our automated mutation screening workflow. An important feature of our LIMS is that it enables tracking of information through a laboratory workflow where the process at one step is contingent on results from a previous step. AVAILABILITY: Script for MySQL database table creation and source code of the whole JSP application are freely available on our website: http://www-gcs.iarc.fr/lims/. SUPPLEMENTARY INFORMATION: System server configuration, database structure and additional details on the LIMS and the mutation screening workflow are available on our website: http://www-gcs.iarc.fr/lims/     

Communication and the laboratory physician

A clinical laboratory documentation system is described, suitable for community hospitals without computer services. The system is cumulative and is designed to provide the laboratory physician with the clinical information necessary for intelligent review and comment on the laboratory''s findings. The mode of presentation of requests to the laboratory and lay-out of the reports to the clinicians are designed to make the two-way communication as close and personal as possible; to encourage the selection of those investigations likely to prove rewarding, and to discourage unnecessary investigation. The possibility of important data escaping notice is minimized.The system is economical in capital equipment, labour and supplies.     

Understanding behavioral responses of fish to pheromones in natural freshwater environments

There is an abundance of experimental studies and reviews that describe odorant-mediated behaviors of fish in laboratory microcosms, but research in natural field conditions has received considerably less attention. Fish pheromone studies in laboratory settings can be highly productive and allow for controlled experimental designs; however, laboratory tanks and flumes often cannot replicate all the physical, physiological and social contexts associated with natural environments. Field experiments can be a critical step in affirming and enhancing understanding of laboratory discoveries and often implicate the ecological significance of pheromones employed by fishes. When findings from laboratory experiments have been further tested in field environments, often different and sometimes contradictory conclusions are found. Examples include studies of sea lamprey (Petromyzon marinus) mating pheromones and fish alarm substances. Here, we review field research conducted on fish pheromones and alarm substances, highlighting the following topics: (1) contradictory results obtained in laboratory and field experiments, (2) how environmental context and physiological status influences behavior, (3) challenges and constraints of aquatic field research and (4) innovative techniques and experimental designs that advance understanding of fish chemical ecology through field research.     

Die Kommunikation von Sialis (Megaloptera) durch Vibrationssignale

A newly discovered system of communication used by Sialidae while mating is described here. By means of this system males and unmated females of Sialis lutaria and S. fuliginosa are able to find each other and to communicate. Three kinds of signals are to be distinguished: (a) Rhythmic vibrations of the abdomen (♂+♀) allow mutual approach and recognition of species and sex; (2) prolonged, unstructed vibrations (♂ only), and (3) tapping of abdomen and wings on the ground is used by ♂♂ of S. lutaria as feedback for ♀♀ after their approach and by ♂♂ and ♀♀ of S. fuliginosa in the first step of their approach. The vibrations caused by movements of the abdomen are passed on via legs on to the ground and transmitted there only. The receptor is located within the legs. Observations in nature are being tested by experiments in the laboratory.     

Standard Test Set and a Computer-generated Diagnostic Register for Gram negative Fermentative Rods

A simple two-step method for the identification of Gram negative fermentative rods is based on interpretation of test results by means of a computer-generated diagnostic register. The first step is based on 10 tests, the second on six. Of 1451 strains routinely isolated in clinical laboratories 84% were identified in the first step. If the second step was included, the efficacy of the system rose to 99.17%. Disagreements with the conventional method in the first step are discussed.     

Comparison of five methods for determination of total plasma protein concentration

Quantitation of exact total protein content is often a key step and is common to many applications in general biochemistry research and routine clinical laboratory practice. Before embarking on any type of protein analysis, particularly comparative techniques, it is important to accurately quantitate the amount of protein in the sample. In order to assess the quality of total protein estimation results, five methods were tested and were applied to the same pooled plasma sample. For this aim, Bradford (Coomassie Brilliant Blue), Lowry (Folin–Ciocalteau), Biüret, Pesce and Strande (Ponceau–S/TCA), and modified method of Schaffner–Weismann (Amido Black 10B) were used. The last two methods employ simultaneous precipitation of proteins with the acid containing dye solutions followed by dissolution of precipitate in a NaOH solution. It is shown that each assay has advantages and disadvantages relative to sensitivity, ease of performance, acceptance in literature, accuracy and reproducibility/coefficient of variation. All of the methods tested show a CV % < 6. Besides pooled plasma, a known concentration of human serum albumin was also analyzed and discussed by means of standardization of plasma total protein content.