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PubMed1994

Scoring of radiation-induced micronuclei in cytokinesis-blocked human lymphocytes by automated image analysis.

The micronucleus assay in human lymphocytes is, at present, frequently used to assess chromosomal damage caused by ionizing radiation or mutagens. Manual scoring of micronuclei (MN) by trained personnel is very time-consuming, tiring work, and the results depend on subjective interpretation of scoring criteria. More objective scoring can be accomplished only if the test can be automated. Furthermore, an automated system allows scoring of large numbers of cells, thereby increasing the statistical significance of the results. This is of special importance for screening programs for low doses of chromosome-damaging agents. In this paper, the first results of our effort to automate the micronucleus assay with an image-analysis system are represented. The method we used is described in detail, and the results are compared to those of other groups. Our system is able to detect 88% of the binucleated lymphocytes on the slides. The procedure consists of a fully automated localization of binucleated cells and counting of the MN within these cells, followed by a simple and fast manual operation in which the false positives are removed. Preliminary measurements for blood samples irradiated with a dose of 1 Gy X-rays indicate that the automated system can find 89% +/- 12% of the micronuclei within the binucleated cells compared to a manual screening.

Algorithms↗
PubMed2006

Clinical impact of a new automated system employed for peripheral blood stem cell collection.

At the moment, PBSC collections can be performed using semi-automated or automated cell separator devices. The automated methods offer the advantages of a decreased working load for dedicated personnel and high standardization of the collection procedure. Herein we report our single institutional experience in 80 PBSC collections employing the new automated COM.TEC Fresenius autoMNC program that provides the ability to predict the total number of CD34(+) cells collected, based on the pre-leukapheresis CD34(+) cell count in peripheral blood. Fourty-eight patients and 21 healthy donors were mobilized with chemotherapy + G-CSF or G-CSF alone, respectively. Eighty leukapheresis collections were performed starting with a CD34(+) cell count in peripheral blood at least of 20/microL. Collection parameters and related side effects were evaluated. The mean CD34(+) cell collection efficiency in patients and donors was 81.8% (sd 27.6) and 95.1% (sd 15.6), respectively. The mean difference between real and predicted CD34(+) cells was +30.2% (sd 92.9) for patients and +4.6% (sd 30.3) for donors. The mean leukapheresis bag volume was 240.7 ml (sd 67.3) and 310.3 ml (sd 86.8) with a mean HCT of 10.9% (sd 34.4) and 9.2% (sd 3.9) for patients and donors, respectively. The automated PBSC collection with the new program COM.TEC Fresenius autoMNC demonstrated a very high CD34(+) cell collection efficiency. Moreover, the ability to predict the CD34(+) cell yield permits improved management of the leukapheresis collection, with the only disadvantage of larger leukapheresis volume and higher hematocrit.

Adolescent↗
PubMed1992

A fully automated method for mononuclear bone marrow cell concentration.

We describe our experience in processing 40 bone marrow aspirates harvested for autotransplantation from patients with several hematological diseases using the CS-3000 blood cell separator. The bone marrow of the first 30 patients was processed by a semiautomated method, and a fully automated procedure was used for the remaining 10 cases. Both procedures were developed in our laboratory and yielded a similar average mononuclear cell recovery of 87.78% and 86.98%, respectively, and similar nucleated cell recovery (27.39% and 27.11%). The cloning efficiency of hematopoietic progenitor cells, measured as the total CFU-GM colony recovery in the in vitro cultures, did not differ between processed and recovered mononuclear cells. On the other hand, all the patients with transplants showed complete hematologic recovery, and the time to engraftment was similar to that described for other procedures. The automated procedure resulted in an average red cell removal of 97.81%, similar to the semiautomated procedure (94.19%), though with a narrower range (96.31-98.6% vs. 80.34-98.34%). The time taken to process a similar amount of bone marrow cell suspension was very different for each method: 1 hour for the fully automated vs. 2 1/2 hours for the semiautomated method to process 1,000 ml. Furthermore, the semiautomated procedure required the addition of homologous or irradiated plasma in a laminar air flow chamber, while the automated method is performed in a closed sterile system. We conclude that our procedure using the CS-3000 processor is an efficient method for fully automated large-scale processing of human bone marrow cells.

Adolescent↗
PubMed2001

CAFASP2: the second critical assessment of fully automated structure prediction methods.

The results of the second Critical Assessment of Fully Automated Structure Prediction (CAFASP2) are presented. The goals of CAFASP are to (i) assess the performance of fully automatic web servers for structure prediction, by using the same blind prediction targets as those used at CASP4, (ii) inform the community of users about the capabilities of the servers, (iii) allow human groups participating in CASP to use and analyze the results of the servers while preparing their nonautomated predictions for CASP, and (iv) compare the performance of the automated servers to that of the human-expert groups of CASP. More than 30 servers from around the world participated in CAFASP2, covering all categories of structure prediction. The category with the largest participation was fold recognition, where 24 CAFASP servers filed predictions along with 103 other CASP human groups. The CAFASP evaluation indicated that it is difficult to establish an exact ranking of the servers because the number of prediction targets was relatively small and the differences among many servers were also small. However, roughly a group of five "best" fold recognition servers could be identified. The CASP evaluation identified the same group of top servers albeit with a slightly different relative order. Both evaluations ranked a semiautomated method named CAFASP-CONSENSUS, that filed predictions using the CAFASP results of the servers, above any of the individual servers. Although the predictions of the CAFASP servers were available to human CASP predictors before the CASP submission deadline, the CASP assessment identified only 11 human groups that performed better than the best server. Furthermore, about one fourth of the top 30 performing groups corresponded to automated servers. At least half of the top 11 groups corresponded to human groups that also had a server in CAFASP or to human groups that used the CAFASP results to prepare their predictions. In particular, the CAFASP-CONSENSUS group was ranked 7. This shows that the automated predictions of the servers can be very helpful to human predictors. We conclude that as servers continue to improve, they will become increasingly important in any prediction process, especially when dealing with genome-scale prediction tasks. We expect that in the near future, the performance difference between humans and machines will continue to narrow and that fully automated structure prediction will become an effective companion and complement to experimental structural genomics.

Automation↗
PubMed2001

A highly automated 96-well solid phase extraction and liquid chromatography/tandem mass spectrometry method for the determination of fentanyl in human plasma.

A high-throughput bioanalytical method based on automated sample transfer, automated solid phase extraction, and fast liquid chromatography/tandem mass spectrometry (LC/MS/MS) analysis, has been developed for the determination of the analgesic fentanyl in human plasma. Samples were transferred into 96-well plates using an automated sample handling system. Automated solid phase extraction (SPE) was carried out using a 96-channel programmable liquid-handling workstation using a mixed-mode sorbent. The extracted samples were then dried down, reconstituted and injected onto a silica column using an aqueous/organic mobile phase with tandem mass spectrometric detection. The method has been validated over the concentration range 0.05-100 ng/mL fentanyl in human plasma, based on a 0.25-mL sample size. The assay is sensitive, specific and robust. More than 2000 samples have been analyzed using this method. The automation of the sample preparation steps not only increased the analysis throughput, but also facilitated the transfer of the method between different bioanalytical laboratories of the same organization.

Analgesics↗
PubMed1995

Automation of a high-performance liquid chromatography-based enzyme assay: evaluation of inhibition constants for human immunodeficiency virus-1 protease inhibitors.

Enzyme-based assays are commonly employed in clinical and pharmaceutical laboratories to aid in quantitation of organic substances. Many enzyme assays are tedious, requiring the addition of reagents at multiple time intervals. The HPLC-based analysis of reaction products requires an additional step of vialing the samples and placing them in the autosampler. Such time-consuming, repetitive procedures are ideally suited for automation. We automated an HIV protease assay for the purposes of screening compounds as inhibitors of HIV protease and determining inhibition constants. Automation was accomplished by interfacing a robotic sample processor from a Gilson Model 232/401 biocompatible automatic sample processor and injector, with a Hewlett Packard HPLC. We used this configuration to automate the following steps: (a) preparation of serial dilutions of inhibitor, (b) enzyme assay setup, and (c) quantitation of products of enzyme assays. The resulting automated method produced inhibition constants that were of comparable accuracy and precision to those determined by manual methods.

Amino Acid Sequence↗
PubMed2000

Automation of industrial bioprocesses.

The dramatic development of new electronic devices within the last 25 years has had a substantial influence on the control and automation of industrial bioprocesses. Within this short period of time the method of controlling industrial bioprocesses has changed completely. In this paper, the authors will use a practical approach focusing on the industrial applications of automation systems. From the early attempts to use computers for the automation of biotechnological processes up to the modern process automation systems some milestones are highlighted. Special attention is given to the influence of Standards and Guidelines on the development of automation systems.

Automation↗
PubMed2001

Multicenter evaluation of an automated system using selected bacteria that harbor challenging and clinically relevant mechanisms of resistance to antibiotics.

A multicenter study was carried out to evaluate the performance of a new commercial automated system in comparison with that of the reference agar dilution method. Ten clinical microbiology laboratories tested a collection of 61 strains of gram-negative bacilli (49 Enterobacteriaceae and 12 Pseudomonas aeruginosa), and 6 other laboratories tested a collection of 55 strains of gram-positive cocci (10 enterococci and 45 staphylococci) against 10-20 antimicrobial agents. The strains were selected on the basis that they harbored challenging and characterized mechanisms of resistance. In comparison with the agar reference method, the automated system gave an overall essential agreement (+/-1 dilution) of 94.5%, 93.5%, and 97% for the gram-negative bacilli, enterococci, and staphylococci, respectively. According to the interpretive standards of the National Committee for Clinical Laboratory Standards, the category agreement ranged from 96 to 96.4% for the three sets of organisms. The accuracy of the automated system, as determined by the kappa test, ranged from 0.80 to 0.88, reflecting an almost perfect agreement with the reference technique. Very major, major, and minor errors obtained with the automated system were 0.3%, 2.9%, and 6.6% for gram-negative bacilli, 3.4%, 0%, and 5% for enterococci, and 1%, 1.6%, and 2.7% for staphylococci, respectively. The high rate of very major errors in enterococci was mostly due to a single strain of multidrug-resistant Enterococcus faecium, which was found susceptible to several antibiotics in a majority of participant laboratories. The use of a heavy inoculum and of a broth test medium by the automated system might account for a better expression of certain resistance mechanisms, including beta-lactamases, as compared to the agar dilution reference method. The interlaboratory reproducibility was acceptable, as shown by the narrow dispersion of MICs and by the results of quality control.

Anti-Bacterial Agents↗
PubMed1996

Research on flexible automation and robotics for plant production at Rutgers University.

This is an overview of research activities in the areas of flexible automation and robotics (FAR) within controlled environment plant production systems (CEPPS) in the Department of Bioresource Engineering, Rutgers University. In the past thirty years, our CEPPS research has dealt with the topics including structures and energy, environmental monitoring and control, plant growing systems, operations research and decision support systems, flexible automation and robotics, and impact to natural (i.e. surrounding) environment. Computer and modeling/simulation techniques have been utilized extensively. Mechanized systems have been developed to substitute human's physical labor and maintain uniformity in production. Automation research has been directed towards adding, to the mechanized systems, the capabilities of perception, reasoning, communication, and task planning. Computers, because of their programmability, provide flexibility to automated systems, when incorporated with generic hardware devices. Robots are ideal hardware tools to be employed in flexible automation systems. Some technologies developed in our CEPPS research may be readily adaptable to Closed Bioregenerative Life Support Systems (CBLSS).

Artificial Intelligence↗
PubMed1996

Mission simulation as an approach to develop requirements for automation in Advanced Life Support Systems.

This paper examines mission simulation as an approach to develop requirements for automation and robotics for Advanced Life Support Systems (ALSS). The focus is on requirements and applications for command and control, control and monitoring, situation assessment and response, diagnosis and recovery, adaptive planning and scheduling, and other automation applications in addition to mechanized equipment and robotics applications to reduce the excessive human labor requirements to operate and maintain an ALSS. Based on principles of systems engineering, an approach is proposed to assess requirements for automation and robotics using mission simulation tools. First, the story of a simulated mission is defined in terms of processes with attendant types of resources needed, including options for use of automation and robotic systems. Next, systems dynamics models are used in simulation to reveal the implications for selected resource allocation schemes in terms of resources required to complete operational tasks. The simulations not only help establish ALSS design criteria, but also may offer guidance to ALSS research efforts by identifying gaps in knowledge about procedures and/or biophysical processes. Simulations of a planned one-year mission with 4 crewmembers in a Human Rated Test Facility are presented as an approach to evaluation of mission feasibility and definition of automation and robotics requirements.

Automation↗
PubMed2006

Implementation of a rapid and automated high performance liquid chromatography method development strategy for pharmaceutical drug candidates.

A comprehensive, fully automated strategy is demonstrated for HPLC-UV chromatographic method development using ChromSword optimization software. The strategy involves: (1) the automated screening of various column and mobile phase combinations, (2) rational selection of the best starting conditions; and (3) subsequent automated method development to generate optimized separation methods. Pharmaceutical compounds were applied to solve problematic drug impurity separations. ChromSword software automates the screening, optimization, and documentation steps thus reducing the method development time. The strategy was compared to a manual method development approach showing the automated method strategy affords better selectivity in a shorter time.

Automation↗
PubMed2004

Assessment of automated DNA extraction coupled with real-time PCR for measuring Epstein-Barr virus load in whole blood, peripheral mononuclear cells and plasma.

BACKGROUND: Epstein-Barr virus (EBV) DNA load monitoring in blood has been shown to be essential for the diagnosis of EBV-associated diseases. However, the methods currently used to assess EBV DNA load are often time-consuming and require prior blood separation. OBJECTIVES: The aim of this study was to evaluate the relative diagnostic value of EBV DNA load monitoring in whole blood, peripheral blood mononuclear cells (PBMCs) and plasma after automated DNA extraction using the MagNA Pure extractor followed by LightCycler real-time quantitative PCR (LC-PCR). STUDY DESIGN: First, EBV DNA load was assessed retrospectively after automated or manual extraction on 104 PBMC specimens. Second, EBV DNA load was determined prospectively with the automated extraction procedure in the whole blood, PBMCs and plasma of 100 samples from patients with EBV-related diseases (group 1, n = 20), HIV-seropositive individuals (group 2, n = 66), and healthy EBV carriers (group 3, n = 14). RESULTS: A good correlation was observed between automated and manual extraction on 104 PBMC specimens (r = 0.956; P < 0.0001). In the prospective study, 67 samples were positive in both whole blood and PBMCs, with a good correlation between EBV DNA loads in whole blood and PBMCs (r = 0.936; P < 0.0001). Only 18/100 samples were positive in plasma. Higher viral loads were regularly observed in the three blood compartments from group 1 than from groups 2 and 3. CONCLUSION: This study demonstrated that an automated extraction of EBV DNA is easier to perform in whole blood or plasma than in PBMCs and facilitates the standardisation of EBV DNA measurement by real-time quantitative PCR. The quantitative detection of EBV DNA load in whole blood appeared more sensitive than in plasma for infectious mononucleosis in immunocompetent patients, probably because of a rapid loss of plasmatic EBV DNA. In transplant patients, EBV DNA load monitoring in whole blood and in plasma turned out to be equivalent in terms of feasibility and accuracy for the early diagnosis of post-transplant lymphoproliferative diseases (PTLDs).

AIDS-Related Opportunistic Infections↗
PubMed2005

Measurement of total and differential white blood cell counts in synovial fluid by means of an automated hematology analyzer.

We developed a rapid and accurate method for quantifying total and differential white blood cell (WBC) counts by pretreating synovial fluid with hyaluronidase and using an automated hematology analyzer. Forty-seven samples of synovial fluid that had been placed in blood-collection tubes containing ethylenediamine- N,N,N',N' -tetraacetic acid as an anticoagulant were treated with hyaluronidase at 37 degrees C for 10 minutes, and then the total and differential WBC counts were determined by means of the automated hematology analyzer. Results were compared with those achieved by traditional manual counting methods. For the automated method, the coefficient of variation values for within-run precision of the WBC count were 5.27%, 3.56%, and 3.01% at 0.54, 1.12, and 2.05 x 10(9) /L, respectively; the run-to-run coefficient of variation values was less than 10.0%. The total and differential WBC counts obtained by this automated method showed good correlation with those obtained by the hemocytometer method ( r = .998; P < .0001; regression formula, y = 0.986 x - 0.072). Bland-Altman plots indicated no significant discrepancy between the methods. Our evaluation supports the use of this automated hematology analyzer method to measure total and differential WBC counts, which should aid clinical diagnosis.

Arthritis, Infectious↗
PubMed2005

Automated selection of beam orientations and segmented intensity-modulated radiotherapy (IMRT) for treatment of oesophagus tumors.

BACKGROUND AND PURPOSE: For some treatment sites, there is evidence in the literature that five to nine equi-angular input beam directions are enough for generating IMRT plans. For oesophagus cancer, there is a report showing that going from four to nine beams may even result in lower quality plans. In this paper, our previously published algorithm for automated beam angle selection (Cycle) has been extended to include segmented IMRT. For oesophagus cancer patients, we have investigated whether automated orientation selection from a large number of equi-angular input beam directions (up to thirty-six) for IMRT optimisation can result in improved lung sparing. MATERIALS AND METHODS: CT-data from five oesophagus patients treated recently in our institute were used for this study. For a prescribed mean PTV dose of 55 Gy, Cycle was used in an iterative procedure to minimise the mean lung dose under the following hard constraints: standard deviation for PTV dose inhomogeneity 2% (1,1 Gy), maximum spinal cord dose 45 Gy. Conformal radiotherapy (CFRT) and IMRT plans for a standard four field oesophagus beam configuration were compared with IMRT plans generated by automated selection from nine or thirty-six equi-angular input beam orientations. Comparisons were also made with dose distributions generated with our commercial treatment planning system (TPS), and with observations in the literature. RESULTS: Using Cycle, automated orientation selection from nine or thirty-six input beam directions resulted in improved lung sparing compared to the four field set-ups. Compared to selection from nine input orientations, selection from thirty-six directions did always result in lower mean lung doses, sometimes with even fewer non-zero weight beams. On average only seven beams with a non-zero weight were enough for obtaining the lowest mean lung dose, yielding clinically feasible plans even in case of thirty-six input directions for the optimisation process. With our commercial TPS we observed the same contra-intuitive, unfavourable results as reported in the literature; nine field equi-angular IMRT plans had substantially higher mean lung doses than plans for the conventional four field set-ups. For all cases, the Cycle plans generated from nine equi-angular input directions were superior compared to similar plans generated with our commercial TPS. CONCLUSIONS: For the studied oesophagus cancer patients the best plans for IMRT were obtained with Cycle, using automated beam orientation selection from thirty-six input beam directions. The lowest mean lung doses could be obtained with, on average, a selection of only seven beams with non-zero weight.

Algorithms↗
PubMed2006

A new automated and high-throughput system for analysis of the forced swim test in mice based on magnetic field changes.

INTRODUCTION: MicroAct (Neuroscience, Tokyo, Japan) was originally developed as a new system for an automated analysis of scratching behaviors in mice. This system is based on the detection of electric current in the coils according to the movement of the magnets implanted in the hind paws. We applied and improved this system to establish an automated analysis system of forced swimming behaviors in mice, which is used as an animal model of depression. METHODS: One day before the test, male CD-1 mice were attached with a wire ring to their hind paws under inhalation anesthesia. After attaching a small magnet to both of the wire rings, each animal was placed for 6 min in a glass cylinder filled with water, which is surrounded by a coil. The swimming behaviors of the mouse were analyzed for the measurement of duration of immobility, a major marker of depression in rodents, by the detection system (MicroAct). The duration of immobility was also determined by manual measurement using the swimming behavior-recorded videotapes produced at the same time as the automated analysis. RESULTS: The difference of the duration of immobility between naïve mice and mice with the rings was not significant. The dose-response effect of imipramine (tricyclic antidepressant, 0, 7.5, 15, 30 and 60 mg/kg, p.o.) on the duration of immobility in the last 4-min of the 6-min testing period determined by MicroAct was similar to that assessed by the manual measurement. These data from the two different methods were significantly correlated (r=0.8805). Moreover, throughput of the automated analysis was 15 times more efficient than that of the manual analysis. DISCUSSION: These results suggest that the automated analysis system of forced swimming of mice using MicroAct can be used as a high-throughput method to examine antidepressive activity of a compound with objectivity and reliability.

Animals↗
PubMed2000

Automating solid-phase extraction: current aspects and future prospects.

This paper reviews current trends and techniques in automated solid-phase extraction. The area has shown a dramatic growth the number of manuscripts published over the last 10 years, including applications in environmental science, food science, clinical chemistry, pharmaceutical bioanalysis, forensics, analytical biochemistry and organic synthesis. This dramatic increase of more that 100% per year can be attributed to the commercial availability of higher throughput 96-well workstations and extraction plates that allow numerous samples to be processed simultaneously. These so-called parallel-processing workstations represent the highest throughput systems currently available. The advantages and limitations of other types of systems, including discrete column systems and on-line solid-phase extraction are also discussed. Discussions of how automated solid-phase extractions can be developed, generic approaches to automated solid-phase extraction, and three noteworthy examples of automated extractions are given. The last part of the review suggests possible near- and long-term directions of automated solid-phase extraction.

Automation↗
PubMed1996

Automated screening of cervical cytology specimens.

After more than four decades of research into automation of the process of screening Papanicolaou (Pap) smears, attempts to develop commercially viable automated screening machines have increased in recent years. These developments have been made possible in part because of the improving price-to-performance ratios in computers and other electronics. Although the Pap smear has been responsible for a very significant decrease in mortality of cervical cancer over the past 40 years, concern has arisen over false-negative cases, with their effects on patients, and the associated legal liability, particularly in the United States. In addition, shortages of cytotechnologists, which have been exacerbated by new regulations limiting the number of slides that may be examined per day, have caused concern about handling the workload, which will probably increase as more individuals gain access to preventive health care. Automated screening machines can potentially allow detection of abnormal cases that are missed with conventional screening, although they may substantially increase the cost of Pap smears. The use of automated screening machines represents a change in the way cervical cytology specimens are processed, and with some machines, a significant change in the operation of the cytology laboratory. Current methods for processing and evaluating Pap smears have not changed significantly for the past four decades. This review discusses some of the principles of operation and practical aspects of automated screening machines.

Automation↗
PubMed2001

The feasibility of short automated static perimetry in children.

OBJECTIVE: To evaluate the feasibility of short automated static perimetry using tendency-oriented perimetry in the pediatric population. DESIGN: Prospective observational case series. PARTICIPANTS: Fifty normal children age 6 through 12 years. TESTING: Subjects underwent testing with the Octopus TOP-32 program on the Octopus 1-2-3 automated perimeter. Testing was performed in a typical clinical setting without adaptations to the perimeter, prolonged training, or the use of custom seating. Each eye was tested twice. MAIN OUTCOME MEASURES: Ability to complete automated static perimetry tests with both eyes. Mean sensitivity, mean defect, and loss of variance; gray scale and numeric representations of the field; duration of each test and of the entire session; subjective assessment of each test as normal or abnormal; calculation of test specificity. Comparisons by age and test number were performed. RESULTS: All subjects successfully completed all four tests. The mean duration for each test was 2:30+/- 0.23 minutes. The average time for the whole session, including training, testing both eyes twice, and rest periods, was 25.8+/-4.87 minutes. Improvement in the specificity of the test (fewer abnormal tests in normal children) occurred in direct relation to subject age (R = 0.5). CONCLUSIONS: Automated static perimetry using short, tendency-oriented programs can be successfully performed in normal children age 6 through 12 years in a typical clinical setting. Age was the best predictor of the mean sensitivity, reproducibility, and accuracy of the test, with the most reliable results obtained after 7 years of age. In children 6 to 7 years old, significant interindividual variability was present, and testing success was more dependent on the child's maturity and ability to concentrate. Short automated perimetry seems to be a promising tool for the evaluation of peripheral vision in pediatric patients.

Automation↗